PREPARATION AND QUALITY EVALUATION OF

GINGER WINE

by

Pushpa L. Rai

Central Department of Food Technology Institute of Science and Technology

Tribhuvan University, Nepal November, 2009

ii

Preparation and Quality Evaluation of Ginger Wine

A dissertation submitted to the Central Department of Food Technology

in Tribhuvan University in partial fulfillment of the requirements

for the degree of M. Tech. in Food Technology

by

Pushpa L. Rai

Central Department of Food Technology Institute of Science and Technology

Tribhuvan University Dharan, Hattisar, Nepal

November, 2009

iii

Tribhuvan University

Institute of Science and Technology

Central Department of Food Technology Central Campus of Technology, Dharan

Approval Letter

This dissertation entitled Preparation and Quality Evaluation of Ginger Wine

presented by Pushpa L. Rai has been accepted as the partial fulfillment of the

requirements for the M. Tech. in Food Technology.

Dissertation Commettee

1. Head of Department __________________________

(Assoc. Prof. Dhan B. Karki)

2. External Examiner __________________________

(Prof. Dr. Ganga P. Kharel)

3. Supervisor __________________________

(Assoc. Prof. Dhan B. Karki)

4. Internal Examiner __________________________

(Lecturer Babita Adhikari)

Date: 24th November, 2009

iv

Acknowledgements

I would like to express my hearty sense of gratitude to Assoc. Prof. Dhan Bahadur Karki,

Chief of Central Department of Food Technology, Hattisar, Dharan. He encouraged me to

carry out this dissertation and provided valuable insights to coordinate the sources of

information and to proceed the research work. I would like to express my gratitude to him

for his moral and technical support with frequent inspiration and supervision.

I extend my profound gratitude to Assoc. Prof. Basanta Rai, CCT, Hattisar, Dharan, for

providing me GenStat software programme. I am also grateful to Mr. Bhaskar Mani

Adhikari , Mr. Ghanendra Gartaula, Mr. Santosh Singh, Mrs. Meera Shrestha, Mr. Madhav

Prasad Tiwari, Mr. Dipesh Basyal for providing me constant support during my

dissertation work. I express my hearty gratitude to Mr. Sujan Shrestha, Manager, Makalu

Wine Industries (P.) Ltd., Basantapur, Tehrtathum, for providing me true wine yeast for

this dissertation.

I would like to express my hearty thanks to my teachers, all friends, and staff of library

and laboratory, for their direct and indirect co-operation, and suggestions. I express my

affectionate thanks to D. Katuwal, Dharan-11, Sunsari, for providing me constant

inspirations.

At last, I owe my deepest gratitude to my respected parents and my family for making me

able to stand in this position where I am now.

________________

Pushpa L. Rai

24th November, 2009

v

Abstract

Effect of mash TSS (16, 20 and 24oBrix), ginger amount (1, 1.5 and 2% m/v) and yeast

types (Saccharomyces cerevisiae and Saccharomyces ellipsodeus) on the chemical and

sensory qualities of the wines were studied. The wine was clarified using bentonite

suspension (5% m/v). Fermentation mash containing 10% raisin, 20oBrix TSS, 4.5 pH and

1% ginger (m/v) was found to be optimum for wine fermentation using baker’s yeast (S.

cerevisiae) at room temperature (28-30oC).

The average pH, TSS (oBrix), alcohol content (%v/v), total acidity (as g lactic acid/L),

fixed acidity (as g lactic acid/L), volatile acidity (as g lactic acid/L), reducing sugar (g/L),

esters (as mg ethyl acetate/L alcohol) and total aldehydes (mg acetaldehyde/L alcohol) of

the ginger wines fermented by wine and baker’s yeasts were found to be 4.1 and 4.2, 5.8

and 6.2, 7.8 and 8.71, 7.56 and 7.92, 5.4 and 5.88, 2.16 and 2.04, 5 and 6, 45.5 and 40.23

and 30.61 and 23.16 respectively. TSS, total acidity, fixed acidity, volatile acidity, esters

and total aldehydes were not significantly different but pH, alcohol and reducing sugar

were significantly different (p<0.05). Sensory analysis showed that taste and mouth feel

were not significantly different but smell, color and overall acceptance scores were

significantly higher in wine fermented by true wine yeast compared to baker’s yeast.

Bentonite was found to be most effective at the rate of 0.5g/L for the clarification of

ginger wine. The ginger wine could be prepared from the mash having 10% raisin, 20oBrix

TSS, 1% ginger (m/v) and 4.5pH by using baker’s yeast in comparable quality to that

fermented by true wine yeast.

Contents

Approval Letter .................................................................................................................. iii Acknowledgements ............................................................................................................. iv Abstract ................................................................................................................................ v 1. Introduction ..................................................................................................................... 1

1.1 General introduction .................................................................................................... 1 1.2 Statement of problem ................................................................................................... 2 1.3 Significance of the study ............................................................................................. 2 1.4 Objective of the study .................................................................................................. 3 1.5 Limitations ................................................................................................................... 3

2. Literature review ............................................................................................................. 4 2.1 Introduction of Ginger ................................................................................................. 4 2.2 Varieties of ginger cultivated in Nepal ........................................................................ 4 2.3 Composition of ginger ................................................................................................. 5 2.4 Ginger and its health benefits ...................................................................................... 5 2.5 Historical background of alcoholic beverage .............................................................. 6

2.5.1 Brief description of alcoholic beverages .............................................................. 7 2.5.2 Classification of alcoholic beverages ................................................................... 8

2.6 Traditional alcoholic beverages of Nepal .................................................................. 11 2.6.1 Jand ..................................................................................................................... 11 2.6.2 Rakshi ................................................................................................................. 11 2.6.3 Toddy/ Tadi ........................................................................................................ 11

2.7 History of wine making ............................................................................................. 12 2.8 Classification of wine ................................................................................................ 13 2.9 General cultural conditions for fermentation ............................................................. 15

2.9.1 pH ....................................................................................................................... 16 2.9.2 Temperature ........................................................................................................ 16 2.9.3 Sugar concentration ............................................................................................ 16

2.10 Wine yeast ............................................................................................................... 17 2.11 Alcoholic fermentation ............................................................................................ 17

2.11.1 Biochemistry of alcohol fermentation .............................................................. 18 2.11.2 Malo-lactic fermentation .................................................................................. 19

2.12 Technology of wine production ............................................................................... 19 2.12.1 Selection of raw material .................................................................................. 21 2.12.2 Blending/ Crushing ........................................................................................... 21 2.12.3 Sulphiting/ Preservatives .................................................................................. 21 2.12.4 Yeast ................................................................................................................. 21

2.12.4.1 Inoculum Development and Pitching ........................................................ 23 2.12.4.2 Fermentation .............................................................................................. 23 2.12.4.3 Factors influencing fermentation ............................................................... 25

2.12.4.3.1 Yeast culture ....................................................................................... 25 2.12.4.3.2 Sugar and its concentration ................................................................. 25 2.12.4.3.3 Sulphur-dioxide .................................................................................. 26 2.12.4.3.4 Acids and pH ...................................................................................... 26 2.12.4.3.5 Temperature ........................................................................................ 27 2.12.4.3.6 Minerals and growth factors ............................................................... 28 2.12.4.3.7 Oxygen ................................................................................................ 28

vii

2.12.4.3.8 Ethanol toxicity ................................................................................... 28 2.12.5 Problems during fermentation .......................................................................... 29

2.12.5.1 Stuck fermentation ..................................................................................... 29 2.12.5.2 Production of off-characters ...................................................................... 29 2.12.5.3 Methanol production and its quality .......................................................... 30 2.12.5.4 Activity of undesirable microorganisms .................................................... 30

2.13 Racking .................................................................................................................... 30 2.14 Clarification and fining ............................................................................................ 31 2.15 Stabilization of wine ................................................................................................ 31 2.16 Maturing and aging of wine ..................................................................................... 31 2.17 Bottling .................................................................................................................... 32 2.18 Pasteurization ........................................................................................................... 32 2.19 Finishing .................................................................................................................. 33 2.20 Storage and ageing of wine ..................................................................................... 33 2.21 Wine made from different raw materials ................................................................. 34 2.22 Fining agent and its types ........................................................................................ 35

2.22.1 The Proteins ...................................................................................................... 35 2.22.2 The Earths ......................................................................................................... 36

2.22.2.1 Bentonite and its properties ....................................................................... 36 2.22.2.2 Preparation of bentonite ............................................................................. 37

2.22.3 Synthetic Polymers ........................................................................................... 38 2.22.4 The Colloids ..................................................................................................... 38

2.22.4.1 Natural Polysaccharides ............................................................................ 38 2.22.5 Alternative Methods of Metal Depletion .......................................................... 39 2.22.6 Activated Carbon .............................................................................................. 39 2.22.7 Silica Suspension .............................................................................................. 39

2.23 Components of wine ................................................................................................ 40 2.23.1 Ethanol .............................................................................................................. 40 2.23.2 Methanol ........................................................................................................... 40 2.23.3 Higher alcohols (Fusel oils) .............................................................................. 41 2.23.4 Carbonyl compounds ........................................................................................ 41 2.23.5 Esters ................................................................................................................ 42 2.23.6 Acids ................................................................................................................. 42 2.23.7 Glycerol ............................................................................................................ 43 2.23.8 Minerals ............................................................................................................ 43 2.23.9 Pectins and gums .............................................................................................. 43 2.23.10 Water and sugar .............................................................................................. 44

2.24 Yield ........................................................................................................................ 44 2.25 Wine defects and spoilage ....................................................................................... 44 2.26 Wine and its health benefits ..................................................................................... 45 2.27 Brief Introduction of Ginger wine ........................................................................... 46

3. Materials and methods .................................................................................................. 47 3.1 Raw Materials ............................................................................................................ 47 3.2 Optimization of TSS and amount of ginger in the fermentation mash ...................... 47

3.2.1 Preparation of mash ............................................................................................ 47 3.2.2 Pitching and agitation ......................................................................................... 48 3.2.3 Fermentation ....................................................................................................... 48 3.2.4 Racking, pasteurization and bottling .................................................................. 48 3.2.5 Quality analysis .................................................................................................. 50

viii

3.3 Selection of the best yeast ......................................................................................... 50 3.4 Clarification of ginger wine using Bentonite ............................................................ 50 3.5 Analytical methods .................................................................................................... 50 3.6 Quality analysis of prepared wines ............................................................................ 51

3.6.1 Sensory evaluation .............................................................................................. 51 3.6.2 Statistical analysis ............................................................................................... 51

4. Results and discussion ................................................................................................... 52 4.1 Effect of TSS and ginger amount on the chemical and sensory quality of ginger wine ......................................................................................................................................... 52

4.1.1 Effect on chemical characteristics ...................................................................... 52 4.2 Effect of yeast culture on chemical and sensory properties ...................................... 57

4.2.1 pH ....................................................................................................................... 58 4.2.3 Alcohol content ................................................................................................... 58 4.2.4 Total acidity ........................................................................................................ 59 4.2.6 Volatile acidity ................................................................................................... 60 2.4.7 Reducing sugar ................................................................................................... 60 4.2.8 Esters .................................................................................................................. 61 4.2.9 Total aldehydes ................................................................................................... 61

4.3 Sensory Evaluation .................................................................................................... 61 4.3.1 Smell ................................................................................................................... 62 4.3.2 Taste .................................................................................................................... 62 4.3.3 Mouth feel ........................................................................................................... 62 4.3.4 Color ................................................................................................................... 63 4.3.5 Overall acceptance .............................................................................................. 63

4.4 Effect of bentonite on the clarification of ginger wine. ............................................. 63 5. Conclusion and recommendation ................................................................................. 65

5.1 Conclusions ............................................................................................................... 65 5.2 Recommendations ..................................................................................................... 65

6. Summary ........................................................................................................................ 66 References ........................................................................................................................... 68 Appendices .......................................................................................................................... 74

ix

List of fables and figures

List of tables Table2.1 The chemical composition of ginger……………………………………..…5

Table2.2 Classification of distilled and un-distilled alcoholic beverages produced from

different raw materials……………………………………….……………....9

a. Cereal Grains as raw materials……………………………....……………..9

b. Vegetables as raw materials……………………….…..…………....9

c. Fruit juice as raw materials……………………….….…………….10

d. Other Sources of raw materials…………...…………….................10

Table 2.3 Classification of wines………………………………………………….…...15

Table 2.4 Typical ranges of application of fining agents………………………………36

Table 3.1 Composition of different mashes…………………………..…………..…….47

Table 4.1 Chemical composition of the ginger wines……………...…………………..57

Table A.1 TSS reduction during fermentation of mashes………………………………73

Table A.2 Data obtained from analysis of samples of wines…………………………...73

Table A.3 Specimen cards for sensory evaluation by hedonic rating…………………..74

Table A.4 ANOVA table for chemical properties of ginger wines fermented by baker’s

yeast………………….…..……………………..……………………………75

Table A.5 ANOVA table for sensory characteristics of ginger wines fermented by

baker’s yeast……………….........…………..……………………………….76

Table A.6 Sensory evaluation scores of the wine samples…………................………...77

Table A.7 Chemical composition of the ginger wines fermented by true wine and baker’s

yeasts…………..…….………………………………………………………78

Table A.8 t-test table for chemical properties of the wines……....……………..………78

Table A.9 t-test table for sensory properties of the wines………………...…………….79

Table A.10 ANOVA table for turbidities of ginger wines………………...……………..79

Table A.11 Effect of bentonite on the clarification of ginger wine………..…..….……...80

Table A.12 Average chemical analysis of prize-winning high quality wines……….…...80

Table A.13 Major Wine producing countries of the world-1996………….………...…...81

Table A.14 Composition of some wines………….....……………….….……………….82

x

List of figures Figure 2.1 Simplified pathway of alcohol synthesis by yeast………….………………...18

Figure 2.2 The malo-lactic pathway……………………………………………………...22

Figure 3 Outline of Red Table Wine production……………………………………….22

Figure 3.1 Fermentation of ginger wine………………………………………………….48

Figure 3.2 Preparation of ginger wine……………………………………………………49

Figure 3.3 Clarification of ginger wine using bentonite………………………………….50

Figure 4.1 Effect of initial TSS on the TSS of the ginger wines …………………………52

Figure 4.2 Effect of ginger amount on the TSS of the ginger wines……………………...53

Figure 4.3 Effect of initial TSS on the alcohol content of the ginger wines……...………54

Figure 4.4 Effect of ginger % on the alcohol content of the ginger wines…………...…...54

Figure 4.5 Effect of initial TSS on the sensory properties………………………..………55

Figure 4.6 Effect of ginger amount on the sensory properties…………………..………..55

Figure 4.7 Effect of yeast type on the sensory properties of ginger wines fermented by true

wine and baker’s yeasts……………………………………………..…………61

Figure 4.8 Effect of bentonite on the clarification of ginger wine…………..…………….63

Part I

Introduction

1.1 General introduction

Alcoholic beverages are among the most popular and appreciated food products all over

the world (Ray et al., 2005). Large numbers of distilled and un-distilled alcoholic products

are enjoyed in different geographical regions throughout the world (Jones, 1985). Wine is

the end product of partial or complete alcoholic fermentation of the juice of grape (Prescott

and Dunn, 1987). It is also made from a variety of fruits, such as grapes, peaches, plums or

apricots etc. and saps of different palm tree (Okafor, 1972). Wine is an un-distilled

beverages having 6-20% ethanol by volume (Pearson, 1976). Wine represents a safe and

healthful beverage. It also provides calories and vitamins. Generally wine is made from

grapes. The grapes are crushed to squeeze out the juice and then are left for some time to

ferment (Amerine et al., 1972).

After this first step of winemaking, the primary fermentation stage that usually takes

around one to two weeks while yeast transforms majority of the sugars in the grape juice to

ethanol, which is alcohol. The resulting liquid is then transferred to several vessels for

secondary fermentation when the remaining sugar is slowly converted to alcohol and the

wine gets clearer in color. Some amount of the wine is then placed in oak barrels to age

before bottling that adds aromas to the wine.

Most of the wines, however, are placed inside bottles and shipped right away that can be

opened starting from after few months to twenty years for top wines. It is important to note

though that only a small percentage of wines will be tastier after five years, compared to

after one year. Wild yeast and other microorganisms are present on the skin of the grapes

and these pass into the juicy pulp (known as must) when the fruit is crushed. These are

destroyed by adding sulfur dioxide (or KMS) in the required quantity. Nowadays other

fruits are also used for winemaking. Many spices are used to flavor the wine (Manay and

Shadaksharaswamy, 1987).

Ginger wine is an alcoholic beverage made from a fermented blend of ground ginger

(Zingiber officinale Rosco.) and raisins fermenting by the yeast, Saccharomyces cerevisiae.

2

It is a popular beverage in Europe. The word drink is primarily a verb, meaning to ingest

liquids. Ginger is usually used to flavor a wine. It has many health benefits. Ginger wine

can be consumed by blending with whisky, brandy or rum.

The first documented appearance of Ginger wine occurred with the foundation of 'The

Finsbury Distilling Company' based in the City of London in 1740.

1.2 Statement of problem

Ginger is becoming the major cash crop for the mid-hill Nepalese farmers. Salyan, Palpa,

Tanahu, Syanja, Kaski, Nawalparasi, Bhojpur and Ilam are the leading districts for ginger

production. Most of the ginger is used as spices. The mother ginger root is harvested in

Ashad and Shrawan months. It is humid and heavy rainy season. So, most of the ginger is

decayed due to moist weather in a short period of time. There is no proper transportation

facility for marketing. There is no good market for the ginger. Nepalese farmers are not

getting a good profit by ginger production. It is very hard to achieve the returns of their

investment. So, if ginger is used for the production of ginger wine, farmers would get good

market for their ginger and their socio-economic status will be changed. Ginger has many

health benefits too. So, it would be a very valuable if we utilize ginger for making different

products such as wine, juice, candy, brandy that preserve for several months to years.

1.3 Significance of the study

My proposed work will be focused on the preparation of a good quality ginger wine by

using wine and baker’s yeasts. Ginger can also be used to prepare dry ginger candy. But it

is not an easy method. It requires a suitable dry weather and takes long time to prepare. It

can not be prepared in all seasons. Sugar is not easily available in the rural areas. Ginger

wine can be prepared easily by using baker’s yeast in rural areas which is as comparable to

the ginger wine made by using true wine yeast. It saves the ginger from decaying. If ginger

is used in winemaking, the people of Salyan, Palpa, Tanahu, Syanja, Kaski, Nawalparasi,

Bhojpur and Ilam will raise their economic status by the production of ginger wine and

brandy. Since ginger can be produced in large quantities in hill regions, we can utilize it

effectively for ginger wine production. Ginger wine is consumed in large quantity in

European and other countries. The ginger wine can be exported to those countries and

earned foreign currency. It helps to increase national income in our country.

3

1.4 Objective of the study

The overall objective of the study is to prepare a good quality ginger wine.

The specific objectives of the study are as follows:

I. To determine the optimum amount of ginger and sugar in fermentation mash for

winemaking.

II. Quality comparison of ginger wines prepared by true wine yeast and baker’s yeast.

III. Clarification of ginger wine by using bentonite.

IV. To determine the physicochemical properties of the ginger wine

V. To evaluate sensory characteristics of ginger wines fermented by true wine and

baker’s yeast.

1.5 Limitations

I. The suitable temperature could not be adjusted. Wine fermentation requires 15-

20oC for good quality wine. Ginger wine was prepared at higher temperature

(i.e.28-30oC) than desired temperature due to technical constraints.

II. Clarification could be done by other fining agents but only bentonite was used due

to time constraints.

III. The prepared ginger wine could not be aged properly due to time constraints.

Ageing is an essential requirement for the good organoleptic qualities of wine.

Part II

Literature review

2.1 Introduction of ginger

Ginger (Zingiber officinilae Rosc.) is an herbaceous perennial plant of the family

Zingiberaceae which consists of 47 genera. The genus Zingeber consists of 80-90 species

among them Officinale is cultivated one (Borget, M. 1989). Ginger is becoming the major

cash crop for the mid-hill Nepalese farmers. It is grown successfully from Terai (100

meters above sea level) to mid hills (1500 meters above sea level).

Ginger is one of the oldest spices to be supposedly native to South East Asia, but like

many other tropical plant of economic importance, its exact origin is uncertain. It is

mentioned in early literature of China and India. The adventurer, Marco Polo, in recording

to his travels during the 13th and 14th centuries, noted that ginger was being cultivated in

South China and Malabar Coast of India (Leverington, 1983).

2.2 Varieties of ginger cultivated in Nepal

Ginger is one of the important spices as well medicinal plants in the country. It is

becoming the major cash crop of the mid-hill farmers of Nepal. Salyan, Palpa, Tanahu,

Syanja, Kaski, Nawalparasi, Bhojpur and Ilam are the leading districts for ginger

production.

As ginger rarely set seeds, the general mode of propagation is asexual. This leads to little

variation between forms grown over a wide geographical area (Lawrence, 1984). Therefore

the classification of cultivars of ginger is done according to their germplasm collected area,

such as Calicut, Cochin, Reo de Generio, Salyan, Ilam etc.

In Nepal, locally available ginger has two varieties- fibrous (NASE) and non fibrous

(BOSE). The germplasm collected from Salyan, Bhojpur and Ilam fall under the ‘BOSE’

variety and these are considered the best in quality (Sharma, 1997).

5

2.3 Composition of ginger

The ginger rhizome contains a mixture of an essential oil, a fixed oil, pungent compounds,

starch and other saccharides, proteins, cellulose, waxes, coloring matter, trace minerals etc.

Starch is the most abundant of these components (Jogi et al., 1972). Chemical composition

of ginger varies with varieties, climatic condition, soil condition, fertilizer used etc. Further

there is a great effect of maturity, handling, storage, drying and other processing methods

on the chemical composition of ginger. The chemical composition of green ginger is given

in table 2.1.

Table 2.1 The chemical composition of ginger

Component Value Watt and Merril, 1975 Value Swaminathan, 1974

Moisture (g) 80.89 87

Protein (g) 2.3 1.4

Fat (g) 0.9 1

Fibre (g) 2.4 1.1

Carbohydrate (g 12.3 9.5

Calcium (g) 0.02 0.023

Phosphorus (g) 0.06 0.036

Iron (g) 2.6 2.1

Carotene (mg) 40 40

Thiamine (mg) 0.06 0.02

Niacin (mg) 0.6 0.7

Riboflavin (mg) 0.03 0.04

Ascorbic acid (mg) 6 4

2.4 Ginger and its health benefits

Ginger has been revered for its medicinal and culinary benefits for centuries. The

underground stem known as the rhizome contains the most medicinal benefits of the plant.

The volatile oils of the ginger plant gives ginger its characteristic odor and taste. It is best

to use ginger in its fresh form to obtain the most health benefits from its use. Ginger has

the following health benefits:

6

1. Ginger can help to alleviate diarrhea, aid digestion and reduce flatulence. It also helps

to relieve the nausea associated with morning sickness and motion sickness. Ginger

also helps to neutralize stomach acid that can cause upset and diarrhea.

2. Ginger has natural anti-inflammatory properties. It helps to reduce the inflammation

associated with arthritis.

3. Ginger is a natural decongestant and antihistamine. It helps to relieve the congestion of

colds, and reduces fever as well.

4. Ginger may help to prevent the formation of blood clots by relaxing the muscles around

blood vessels. Ginger is also a natural blood thinner.

5. Ginger can help to lower cholesterol and prevent blood platelets from clumping

together. It also stimulates the circulatory system.

6. Ginger may also be beneficial in the prevention of heart disease and cancer, as well as

in the treatment of diabetes. Research continues to determine the effectiveness of

ginger in these areas as well as other health conditions.

(Source: http://www.ehow.com/facts_4924826_health-benefits-ginger.html)

2.5 Historical background of alcoholic beverage

Alcoholic beverages are among most popular and most appreciated food products all over

the world. Alcohol was discovered in 8327 B.C. on a warm afternoon by “Grog” who

returned to his cave and drank the fermented milk of a coconut that had been cracked and

left out in the sun. Beer and berry wines were made for the first time in 6400 B.C. while

Grape wines were made in 300-400 B.C. (Ray et al., 2005). Large numbers of distilled and

un-distilled alcoholic products are enjoyed in different geographical regions throughout the

globe. Alcoholic beverages are believed to have originated in Egypt and Mesopotamia

some 6000 years ago (Jones, 1985).

Despite this early application of microbiology, the ability of microorganisms to stimulate

the biochemical changes was demonstrated several years later. Gay Lussac first identified

alcoholic fermentation in 1810, but at that time yeast was not recognized as a causative

organism. Schwan in 1835 demonstrated that yeast could produce alcohol and carbon

dioxide when introduced in sugar-containing solution. He termed yeast Zuckerpilz meaning

sugar fungus from which the name Saccharomyces originated (Prescott and Dunn, 1987).

Saccharomyces group possesses almost all the credits of producing alcoholic beverages

(Tannanhill, 1937).

7

The production and consumption of alcoholic beverage is one of the man’s oldest

activities. Today brewing, wine making and distilling are of major commercial importance

in many non-Islamic countries and, through taxation, can be an important source of

government revenue (Vernam and Sutherland, 1994).

There are different types of alcohols. Some are used in chemistry and industry, e.g.

isopropyl and methyl alcohol. Another type of alcohol, also known as ethanol has been

consumed by human beings for its intoxicating and mind-altering effects. The term

‘alcohol’, unless specified otherwise, refers to ethanol or ethyl alcohol.

2.5.1 Brief description of alcoholic beverages

There are many types of alcoholic beverages. They are briefly described as;

Wine: Wines are the oldest of the alcoholic beverages made by fermentation of grape

juice. Wine, strictly speaking, is a product of vine, but often includes all fermented liquors

obtained from different fruit juices (fruit wines). Wines differ greatly in their characters,

because grapes grown in different regions differ in composition, particularly in their

volatile components which contribute to flavor and bouquet and in the method used for

wine making (Amerine et al., 1972).

Wine is the end product of partial or complete alcoholic fermentation of the juice of

grape (Prescott and Dunn, 1987). It is also made from a variety of fruits, such as grapes,

peaches, plums or apricots etc. and saps of different palm tree (Okafor, 1072).

Wine is an un-distilled beverages having 6-20% ethanol by volume (Pearson, 1976).

Most of the natural wines contain 8-10% alcohol. Fortified wines contain about 20%

alcohol, which is sufficiently high to kill the microorganisms that attack natural wines.

Wines containing less than 14% alcohol are table wines, whereas those containing more are

dessert wines. The term wine is broadly used to include any properly fermented juice of

ripe fruits. The names of the fermented products are different according to the types of

fruits used. For example: the product obtained from the grape juice is known as wine,

similarly product from apple juice and pear pulps are known as cider and perry respectively

(CFRL, 1984).

The most common wines are produced from grapes. The soil in which the grapes are

grown and the weather conditions in the growing season determine the quality and taste of

the grapes which in turn affects the taste and quality of wines. When ripe, the grapes are

crushed and fermented in large vats to produce wine.

8

Beer: Beer is also made by the process of fermentation. A liquid mix, called wort, is

prepared by combining yeast and malted cereal, such as corn, rye, wheat or barley.

Fermentation of the liquid mix produces alcohol and carbon dioxide. The process of

fermentation is stopped before it is completed to limit the alcohol content. The product so

produced is called beer. It contains 4 to 8 percent of alcohol.

Whisky: Whisky is made by distilling the fermented juice of cereal grains such as corn, rye

or barley. Scotch whisky was originally made in Scotland. The word “Scotch” has become

almost synonymous with whisky of good quality.

Rum: Rum is distilled beverage made from fermented molasses or sugarcane juice and is

aged for at least three years. Caramel is sometimes used for coloring.

Brandy: Brandy is distilled from fermented fruits juices. Brandy is usually aged in oak

casks. The color of brandy comes either from the casks or from caramel that is added.

Gin: Gin is a distilled beverage. It is a combination of alcohol, water and various flavors.

Gin does not improve with age, so it is not stored in wooden casks.

Liqueurs: Liqueurs are made by distilling sugar and flavoring such as fruits, herbs or

flowers to brandy or to a combination of alcohol and water. Most liqueurs contain 20-65

percent alcohol. They are usually consumed in small quantities after dinner.

2.5.2 Classification of alcoholic beverages

There are different types of distilled and un-distilled congeneric alcoholic beverages all

over the world according to source of raw materials; some are listed in the following table

2.2.

9

Table 2.2 Classification of distilled and un-distilled alcoholic beverages produced from

different raw materials.

a. Cereal Grains as raw materials.

Source Name of fermented beverage Name of distilled beverages

Barley Beer, Barley wine Scotch whisky, Irish whiskey

Rye Rye beer kvass Rye whiskey, Roggenkon (Germany)

Corn Chichi, Corn beer Bourbon whiskey, Vodka

Sorghum Burukutu (Nigeria), Pito

(Ghana)

Maotai, Gaoliang, types of Baijiu

(China)

Wheat Wheat beer Wheat whisky

Rice Huangjiu, Choujiu (China), Sake,

Sonti, Makkoli,

Rice baijiu (China), Shochu and

Awamori (Japan)

Millet Millet beer(Sub-Saharan Africa),

Tongba (Tibet)

b. Vegetables as raw materials.

Source

Name of

fermented

beverage

Name of distilled beverage

Juice of ginger

root

Ginger beer

(Botswana)

Potato and/ or

Grain

Potato beer Vodka: Poland and Germany, Aquavit or Brannvin:

Sweden, Akvavit: Denmark

Beets Pink vodka/ Woman’s vodka/ Girlie vodka (Russia)

10

c. Fruit juice as raw materials.

d. Other Sources of raw materials.

Source Name of fermented beverage Name of distilled beverage

Sap of

palm

Coyol wine (Central America), Tembo

(Sub-Saharan Africa), Toddy in

Nigeria, Tadi (Nepal)

Arrack

Honey Mead, Teg (Ethiopia) Distilled mead (“mead brandy” or

“honey brandy”)

Pomace Pomace Wine Raki (Turkey), tsikoudia (Greece),

grappa (Italy), Trester (Germany),

marc (France)

Milk Kumis or Kefir Araka

Source: htto://en.wikipedia.org/wiki/Alcoholic_beverage

Source Name of fermented

beverage Name of distilled beverage

Juice of grapes Wine, grapes wine Brandy, Cognac (France)

Juice of apples (“Hard”) Cider,

Apfelwein

Applejack (or apple brandy), Calvados,

Cider, Lambic

Juice of pears Perry, or Pear cider,

Poire (France)

Pear brandy, Eau-de-Vie (France)

Juice of sugarcane,

or molasses

Basi, Betsa- betas

(regional)

Rum (Caribbean), Pinga or Cachaca

(Brasil), Aguardiente, Tequila, Mezcal

Juice of agave Pulque Tequila, Mezcal

Juice of plums Plum wine Slivovitz, Tzuica, Palinca

Juice of pineapples Tepache (Mexico)

Juice of Bananas Urgwagwa (Uganda,

Rwanda)

11

2.6 Traditional alcoholic beverages of Nepal

Alcoholic beverages have played an important role in human spiritual and cultural life both

in Eastern and Western societies. Unlike in Europe and the Middle East, where indigenous

alcoholic beverages are produced primarily from fruit, alcoholic beverages are produced

from cereals in the Asia-Pacific region, and serve as an important source of nutrients.

European beer uses barley malt as the primary raw material, while Asian beer utilizes rice

with molded starters as the raw material. Beverages vary from crystal-clear products to

turbid thick gruels and pastes. Clear products which are generally referred to as

Shaosingjiu in China, Chongju in Korea and Sake in Japan, contain at least 15% alcohol

and are designated as rice-wine, while turbid beverages, such as Takju in Korea and Tapuy

in the Philippines which contain less contain less than 8% alcohol along with suspended

insoluble solids and live yeasts, are referred to as rice-beer (Haaed, 1999).

2.6.1 Jand

Jand is an alcoholic beverage (un-distilled) indigenous to Nepal. It is prepared by solid-

substrate fermentation of starchy cereals like corn, rice, wheat and millet. Murcha, a starter

culture, is used as the inoculum in traditional fermentation. Murcha contains saccharifying

molds, lactic acid bacteria and fermenting yeasts. Jand is therefore the result of concerted

action of these microorganisms on the cooked cereal (Rai, 2005).

2.6.2 Rakshi

Raksi (also spelt rakshi, rukhsi) is an un-aged congeneric spirit obtained by pot distillation

of the slurry of jand. The product likens whiskey and has highly varying alcohol contents

(K.C. et al., 2004), generally between of 15 and 40% (Subba et al., 2005). Several basic

researches have been done on raksi production from different cereals using murcha starter

as well as wine cultures isolated thereof (Rai, 1984) but there seems to be general lack of

attention towards process development such as preparation of good starter culture,

increasing efficiency of traditional distillation apparatus, and separation of fients and

foreshots for improving quality of raksi.

2.6.3 Toddy/ Tadi

It is a fermented sap of palm trees by natural contamination. In Nepal naturally fermented

palm sap used as alcoholic beverage is called “Tadi”. Traditionally, sap is collected

12

overnight in clay pots (with bottom containing a crust of microorganisms formed from the

previous fermentation), from the slit made at the top portion of the tree trunk. The tapped

sap, which is trickles down into the collection pot, is inoculated and fermentation sets

immediately. The sap is converted into sweet Tadi by the fermentation. This product is

white and effervescent (Dhakal, 2007).

2.7 History of wine making

As stated by sir John Malcohn in his first account of Persia during the regime of king

Jamshed, Viticulture flourished and it is he who is credited with the dictionary of

fermentation (Andrew, 1980). History of wines has left its traces in Near East, particularly

Mesopotamia (Iraq, Iran territory), later– in Persia (Iran), Egypt, Ancient Greece, Roman

Empire. Think of Greek classical pottery and Dionysus cavorting with his satyrs and

maenads and you will get a clue of the ancient history of wine that created immortal

legends. Egyptian history of wines origin in Nile delta– the fertile land where grapes grew

and white wine made from what is today called the Muscat grape of Alexandria. It is not

surprising that the early Egyptians attributed this drink with the god Osiris and used it

during funerary rituals.

Since Roman times, wine (potentially mixed with herbs and minerals) was assumed to

serve medicinal purposes as well. It was not uncommon to dissolve pearls in wine for

better health. Cleopatra created her own legend by promising Marc Anthony she would

"drink the value of a province" in one cup of wine, after which she drank an expensive

pearl with a cup of wine. From Rome winemaking greatly prospered under the Catholic

Church who held widespread influence over Christian Europe. Eventually, winemaking

capability and practiced extended to far-flung places like England who enjoyed wine

varieties of Sherry, Port and Madeira. Christian monks of France and Northern Italy kept

records of their winemaking practices and grape cultivation. By 1800, France would be

recognized as the best of the wine-producing regions of the world.

(Source: http.//www.metalimagination.com/winemaking.html).

The Pheonicians from Lebanon introduced the wine and its secrets to the Romans and

Greeks who subsequently propagated wine making and even dedicated a God to wine the

Roman Bacchus and the Greek Dionysus. Fermented beverages have been produced since

the Paleolithic period probably at first by accident from honey. Later, cereals were used

13

and then grapes and various fruits. During the Neolithic period, wines from fruits, and

especially from grapes, were more popular in Greek and Roman territory (Dhakal, 1988).

Heating wine to produce a caramelized or baked odor was known in the Roman period. In

the 19th century, it was first used in the Madeira Islands (Johnson, 1974).

Crude methods of clarification, preventing spoilage, and treating spoiled wines were

developed by the Romans (Kirk, 1969). Wine is probably the most widespread and

historically significant beverage starting from ancient times. Wine is the drink of kings,

just as it is the beverage of choice for ordinary people. Wine has played a major role in the

rise and fall of countless individuals, nations and even civilizations. History of wine is very

long, interesting and intricate at the same time; nevertheless, classification of wine is no

less capturing and complicated as its history.

Grape wine is found widely distributed throughout the world. The most important species

Vitis vinifera is believed to have been brought by man from Southern Russia to Asia

Minor. Europe is obviously the most important wine-producing area with more than 75%

on average and over 68% of wine production comes from European countries, with France

and Italy capturing nearly 45% of total production (Amerine et al., 1967). For the mass

production in wineries, new methodologies and technologies were implemented so called

modern wineries. (Source: http://www. Byronwines.com/iw_facilities.asp)

Modern wineries are automatic and computerized and are capable of producing 3-4

million liters of wine with only handful of people (Birch and Lindley, 1985). More

recently, the use of tower fermentation (Berry and Watson, 1987) with timer and

programmer for the production of both wine and cider has been demonstrated.

In Nepal, there are only three wineries, one in Basantapur, Tehrathum (Makalu wine

industries (P.) Ltd.) and others in Jomsom, Mustang and Pokhara (Hill Hut Winery) to

produce raspberry wine, cider and different fruit wines respectively.

2.8 Classification of wine

Types of wines are normally classified by vinification method, by taste, by vintage, by

wine style, and/or by quality. Vinification refers to how the wine is made. Vinification

wine classification refers to three major categories: table wines, sparkling wines, and

fortified wines. Types of wine can also be classified by taste. Table wines, for instance, are

classified by character as dry (not sweet), semidry, semisweet; sweet wines are classified

as dessert wines. Apart from palate, types of wines can also be distinguished by sugar and

14

alcohol percentage. Dry wine contains 2-3% of sugar and about 10% of alcohol– such wine

is the lightest. Semisweet wines have sugar 5-6% and alcohol 13-14%, while semidry

wines are a little bit sweeter than semisweet ones. Dessert or sweet wines contain the

highest percentage of sugar and alcohols than other types of wine– about 14-16%, and 16%

of alcohol. Table wines are also further classified by color, as red, white, or rose (pink). In

addition to this wine classification, wines may also be classified according to specific

flavors, types of grape they are made of and origins where this grape grew.

Table wines, also called still or natural wines, are consumed mostly with food, they tend

to compliment the meal. Table wines contain less than 14% alcohol. White dry wine is

usually served with seafood, fish, cheese, or nuts. Red dry wine is served with meals of

meat and vegetables that are roasted, stewed, smoked, etc. Fortified or dessert types of

wine, such as sherry or vermouth, are most commonly drunk before or after meals and are

served with various cakes, pastry, chocolate, fruits, etc. Fortified wines are also frequently

used in cooking. Concerning sparkling wines, for example champagne, is distinguishable

by its effervescence and is drunk for the most part on festive occasions such as weddings,

birthdays, and during the holidays.

Wines are usually named either by their grape variety or by their place of production.

Generally speaking, European wines are named both after the place of production (e.g.

Bordeaux, Rioja, Chianti, Cotnari) and the grapes used (e.g. Pinot, Riesling, Chardonnay,

Merlot). Wines from everywhere except Europe are generally named for the grape variety.

Whether you prefer vintage wine or not, whatever the classification of wine you like, wine

is an ideal gift for any special occasion.

Wines can be classified on various bases viz., (i) color, (ii) relative sweetness, (iii)

effervescence, (iv) alcohol content, and (v) the system used by Wine Advisory Board,

USA. However, the basic groups of wines are most easily distinguishable for the consumer.

They are (i) table wines, (ii) sparkling wines, and (iii) fortified wines. A summary of the

classification scheme is given in table 2.3.

15

Table 2.3 Classification of wines.

Basis of classification Class/ type Description Example

Color Red wine Contains the red coloring matter of skin,

pulp and seeds.

Burgundy

White wine Does not contain the red coloring matter,

pulp and seeds.

Rhine wine

Pink wine Low concentration of red coloring

matter is maintained.

Rose

Relative sweetness Sweet wine Contains up to 7% sugar. Sherry

(sweet)

Dry wine Contains less than 0.12% sugar. Sherry (Dry)

Alcohol content Natural Contains 8.5-16% alcohol by volume (%

abv)

Table wines

Fortified Contains 17-21% abv. Sherry

Effervescence Still Does not contain CO2 Chianti

Sparkling Contains CO2 (Natural or Artificial ) Champagne

Wine Advisory Board,

USA

Dessert wine Contains sugar; taken after meal Sherry

(Sweet)

Appetizer

wine

Dry; fortified; taken before meal Sherry (Dry)

Sparkling

wine

Contains CO2 Champagne

Red-table

wine

Natural; red in color Chianti

White-table

wine

Natural; pale yellow to straw color Rhine wine

Note: There is considerable overlapping of wine types in the classification shown above.

For example, a Red Table wine can at the same time is sweet, sparkling, fortified, or

natural. Similarly, a fortified wine can be sweet, sparkling, red, or white (Rai, 2002).

2.9 General cultural conditions for fermentation

Cultural condition refers to the environment of yeast i.e. fermentation media on which the

propagation of yeast as well as final quality of wine is largely depended (Varnam and

16

Sutherland, 1994). Following are the few parameters, which determine cultural condition

of the fermentation media.

2.9.1 pH

The optimum pH for wine production varies from types of the selected fruit but generally

3.8-4.5 is supposed to be optimum. At higher pH, the concentration of glycerine is

increased during fermentation whereas at lower pH, there is a noticeable effect of log phase

(Prescott and Dunn, 1987).

2.9.2 Temperature

The optimum temperature for the fermentation is dependent upon the types of wines

produced. For white wine, the temperature is 10-15oC and that for the red wine is 20-30oC

(Prescott and Dunn, 1987). There is possibility of ‘stuck’ fermentation if it is carried at

higher temperature. On the other hand, low temperature may delay onset of fermentation.

At high temperature, the loss of alcohol and aroma substance takes place. Also, a large

amount of by-product like glycerol, acetaldehyde may be formed. An imbalance of these

constituents can be very detrimental to wine quality. It has been reported that at higher

temperature the formation of higher alcohol decreases (Peynand and Gumiberteau, 1962).

The fermentation temperature for most white wines is in the range of 18oC to 24oC and

there is little interest in fermenting at higher temperatures due to the progressive loss of

volatiles under these conditions. The contribution of the fermentation temperature to white

wine aroma is directly related to the retention of grape-based aromas and formation of the

group of volatile byproducts referred to as fermentation bouquet (Boulton et al., 1997).

There are additional effects of fermentation temperature on the formation of glycerol

(Ough and Amerine, 1965) and the higher alcohols (Ough et al., 1966). The advantage of

lower fermentation temperature are the fresher and fruitier character of wine, smaller losses

of ethanol and less danger of producing volatile acidity (Prescott and Dunn, 1987).

2.9.3 Sugar concentration

The ‘must’ having very high sugar concentration imparts high osmotic pressure, which in

turn has a negative effect on yeast cells, since both growth of yeast and fermentation

activity are lowered. The tolerance of higher sugar concentration varies according to the

yeast species (Prescott and Dunn, 1987).

17

2.10 Wine yeast

Wine yeast is the member of the Saccharomyces cerevisiae group. The name originates

from the Greek word sakchar means sugar and mykes means fungus, referring to the strong

sugar fermenting properties of the genus in general. Although, Hansen regarded them as a

separate species, they are more ellipsoid in shape than the round or oval cells of brewery

and bakery yeasts. Hansen restricted the name S. ellipsoideus to them. In the nomenclature

of Dutch school, these yeasts are classified as a variety of S. ellipsoideus and consequently

named S. cerevisiae var. ellipsoideus (Austin, 1968).

Good wine yeast is one which will impart a vinous or fruit like flavor, will ferment sugar

to a low content producing 14-18% alcohol, and is characterized by remaining in

suspension during fermentation and than agglomerating to yield a coarse granular sediment

that settles quickly and is not easily disturbed in racking (Pederson, 1971).

Good wine yeast should have the following four properties:

1. High alcohol tolerance, i.e. the yeast should continue to ferment despite the

increasing concentration of the alcohol, giving stronger, drier wines with up to 16%

alcohol (v/v), or even up to 18% (v/v) where the yeast is fed by periodic additions

of sugar in small amounts.

2. Good degree of agglutination, i.e., the tendency of the yeast to flocculate into small

lumps that give a cohesive sediment as fermentation ceases, so that racking is

simple and the wine clears easily.

3. Steady, persistent fermentation capacity; this leads to wines of better quality than

when the fermentation falls away after a tempestuous start.

4. Absence of unpleasant flavors generated by dead and dying cells (Austin, 1968).

2.11 Alcoholic fermentation

There are different kinds of alcohols, but when the term is used loosely as by winemakers,

it invariably applies to the potable alcohol called ethyl alcohol or ethanol. It mixes easily

with water in any proportion and where quantities are mixed there is a contraction in

volume. It has a low boiling point, 78.4oC, compared with water. It burns easily in air, so

that oxidation is possible and then gives a blue, smokeless flame, producing water and

CO2. Ethyl alcohol is produced by the zymase complex of enzymes in yeast (Austin,

1968). There are three main classes of alcoholic beverages; wines, malted beverages and

18

distilled liquors (Lal et al., 1987). The essential step in all the fermentation processes is the

conversion of glucose into alcohol by yeast (Manay and Shsdasharaswamy, 1987).

The intermediate products are methyl glyoxal (CH3:OCH:O), Acetaldehyde (CH3CHO)

and pyruvic acid (CH3COCOOH).

Alcoholic fermentation is simply the production of alcohol by using carbon and nitrogen

substrate (Kaushik and Yadav, 1997). Sugar and nitrogen compounds are the principal

substrates for alcohol fermentation (Prescott and Dunn, 1987).

2.11.1 Biochemistry of alcohol fermentation

Alcoholic fermentation is an anaerobic process (i.e. takes place in the absence of air).

Microorganisms utilize the carbohydrate present in the materials to obtain energy for

growth and metabolic activities, leading to the formation of alcohol. Monosaccharides

(hexoses) are directly fermented. The flow of carbon in ethyl alcohol formation takes place

via the well known Embden-Meyerhof-Parnas pathway (Patel, 1999). The formation of

alcohol from sugar is accomplished by yeast enzymes which are contributed by the

growing yeasts. S. ellipsoideus is the true wine yeast. The organism uses EMP pathway,

generating two ATPs per mole of glucose converted to ethanol, plus CO2. Ethanol, which

is the end product, is primary metabolite. In an industrial fermentation, the basic strategy is

to maintain Crabtree effect during the fermentation. A truncated form of the metabolic

pathway for ethanol synthesis is given in Fig. 2.1.

Fig. 2.1 Simplified pathway of alcohol synthesis by yeast.

+C6H12O6 C2H5OH 2CO2

2ADP2ATP

Glucose 2[1, 3-di P glycerate]

4 ADP

4 ATP

4 Pyruvate

2 Acetaldehyde

2 [NAD + H+] 2 [NAD]

Alcohol dehydrogenase2 Ethanol CO2

19

2.11.2 Malo-lactic fermentation

It refers to secondary fermentation in which lactic acid bacteria are allowed to metabolize

malic acid to lactic acid and carbon dioxide. This fermentation is particularly useful if the

titrable acidity of wine is to be reduced. Wines with low levels of acidity should be

protected from malo-lactic fermentation: wine quality decreases if the acid level falls too

low. Malo-lactic fermentation can be easily prevented by early racking, cool storage, and

maintaining 100 p.p.m. or more of SO2. On the other hand, if such fermentation is desired,

it can be facilitated by leaving the wine on the lees (yeast sediments) for prolonged periods

at higher temperatures. This storage causes lysis of yeast cells and releases amino acids and

other nutrients needed for the growth of the ‘contaminant’ lactic acid bacteria.

Malo-lactic fermentation has an important bearing in the quality of wine. It is a natural

way of reducing acidity in wine. Besides, the fermentation also results in wines with

greater softness and mellowness. The bacteria implicated for malo-lactic fermentation are

Leuconostoc oenos, Lactobacillus, and Pediococcus, the first one being the most important

(Rai, 2005). The biochemistry of fermentation is given in figure 2.2.

Figure 2.2 The malo-lactic pathway.

2.12 Technology of wine production

Winemaking starts during the time of harvest when grapes are selected and placed in

containers. After harvesting, the grapes are crushed to squeeze out the juice and then are

left for some time to ferment. The winemaking technology of red and white wines also

differs. If red wine is desired, the skins are left to soak in the juice for a while so that the

wine would take the skin’s color. In order to make white wine, the juice is extracted with

COOH

COOH

CH2

COOH L-malic acid

L-malate dehydrogenase Pyruvate + CO2

Malo-lactic enzyme

L-lactose dehydrogenase

CH3CH2COOH

20

minimal contact from the grape skin. After this first step of winemaking, the primary

fermentation stage that usually takes around one to two weeks while yeast transforms

majority of the sugars in the grape juice to ethanol, which is alcohol.

According to winemaking technology, the resulting liquid is then transferred to several

vessels for secondary fermentation when the remaining sugar is slowly converted to

alcohol and the wine gets clearer in color. Sweet wines are created by allowing some

residual sugar to remain before or after fermentation or by adding another alcoholic

beverage to kill the yeast before fermentation is completed. Some amount of the wine is

then placed in oak barrels to age before bottling that adds aromas to the wine. Most of the

wines, however, are placed inside bottles and shipped right away that can be opened

starting from after a few months to twenty years for top wines. It is important to note

though that only a small percentage of wines will be tastier after five years, compared to

after one year.

Wild yeast and other microorganisms are present on the skin of the grapes and these pass

into the juicy pulp (known as must) when the fruit is crushed. These are destroyed by

adding sulfur dioxide (or KMS) in the required quantity. If the sugar content is low,

sucrose is added to the desired strength and the pH is adjusted to 3.2 to 3.4 by the addition

of tartaric acid. Next, the must is inoculated with a wine culture of actively growing yeast

(S. ellipsoideus). The temperature and duration of fermentation depend upon whether dry

or sweet wine is required. Fermentation usually lasts 4-10 days.

When fermentation is complete, the clear wine is siphoned from the yeast sediment into

barrels (racking) and the wine is allowed to age. During this period, secondary

fermentation takes place and wine also losses its raw and harsh flavor and mellows down.

During this period of maturation, clarification takes place in natural way. It can also be

achieved by fining and filtration. Next, the wine is bottled and allowed to mature; the time

of this maturation extends to a number of years depending upon the quality desired (Manay

and Shadaksharaswamy, 1987).

21

2.12.1 Selection of raw material

A suitable raw material is chosen to function as a substrate. Compared to cereals, fruit

juices are more readily utilizable substrate by yeasts for the alcoholic fermentation. The

later is also a suitable media for the yeast to grow (Varnam and Sutherland, 1994). Good

raw material for fermentation should be clean, sound, mature, impart from any taste and

odor and good source of carbon and Nitrogen with sufficient amount of fermentable sugar.

(Source: http/www.austwine/0089a/rm.html).

2.12.2 Blending/ Crushing

This step is carried out to extract the juice from the fruit. It has been suggested that the

process should be very gentle (Vernam and Suthearland, 1994). If the blending and

crushing machine is constructed of mild steel or cast iron then iron causes ‘ferric casse-

cloudiness’ of wine due to iron; actually iron will react with the tannin of the juice to form

ferric-tannin complex. Bronze equipment is also used but may cause dissolution of copper

and tin from bronze equipment and it will affect the color. Usually, stainless steel is used

for the crushing machine. Water may be added during blending/crushing for smoothness of

operation.

2.12.3 Sulphiting/ Preservatives

The antiseptic and antioxidant properties of sulfur dioxide are taken advantage of both in

connection with treatment of musts prior to fermentation and later in the winemaking

process. The dosage of SO2 usually ranges between 100 and 200 p.p.m. (Douglas and

Considine, 1982). SO2 is added before the fermentation process to prevent air oxidizing the

juice and converting the alcohol into vinegar. The air has bacteria principally Acetobacter

i.e. it is alive in the presence of air of oxygen, takes of the oxygen from the must to let the

wine yeast which is anaerobic condition convert the fruit sugar into alcohol. SO2 also

forms a coating on the surface of juice to prevent the air entering the juice (Andrew, 1980).

2.12.4 Yeast

Wine yeasts are the member of Saccharomyces and consequently of great individual

importance (Austin, 1968). A good quality of wine yeast should have the following

characters (Vernam and Sutherland, 1994):

• Introduction of flocculation and reduction of H2S production.

22

• Reduction of higher alcohol production.

• Improvement of fermentation efficiency.

• Resistance of ethanol.

• Resistance of killer activity.

Fig. 2.3 Outline of Red Table Wine production

Red table wine

Bottling, Labelling, Casing

Secondary Fermentation and Filling

Racking, Blending, Fining, Malo-lactic fermentation

Filtration and Tartarate Stabilization

Polishing

Pasteurization

SO2 (75 ppm)

Yeast

Press wine

Propagation

Free-run wine

Primary Fermentation

Drawing off and Pressing

Purple grapes

Destemming

Crushing

Must

Must Treatment

Pomace

SO2 (75-125 ppm)

23

2.12.4.1 Inoculum Development and Pitching

Sufficient quantity of inoculums (pitch) is developed before the preparation of must. The

developing medium should have law sugar concentration so that the ‘Pasteur Effect’ is

maintained and maximum growth is necessary for the respiration of growing yeast cells.

The medium should, preferably, be the juice of the same fruit so that the yeast is adopted

with the fruit juice composition. Pitching is done when the culture of the pitch is at its

optimum stage of growth. Vigorous agitation is done after pitching to help distribute the

culture and also to help in their initial growth (Karki, 2001).

2.12.4.2 Fermentation

In alcoholic fermentation by yeast, which is an anaerobic process, sugar (or glucose) is the

substrate and alcohol is produced as the product along with carbon dioxide. According to

the Gay-Lussac’s equation, theoretical yield of 51.1% alcohol (ethanol) and 48.9% CO2 of

the weight of the sugar fermented, is possible. This is biologically unobtainable and

possible only in absence of yeast growth and loss of alcohol as vapor (Karki, 2001). The

yield of alcohol varies from 47.87 to 48.12% and of CO2 from 47.02 to 47.68% of the

weight of sugar fermented (Gvaladez, 1936). Fermentation is the soul (heart) of wine

making. All the desirable reactions take place during this step, so most of wine makers pay

strict attention to this stage. Fermentation is the process of adding wine yeast (technically

termed as S. ellipsoidues) to fresh juice to convert the natural sugar to ethyl alcohol. In this

process, CO2 is simultaneously released making fermentation violent at first and then slow.

The yeast added is 1-3% of the volume of the juice. Generally, 14 days is required for

complete alcoholic fermentation.

Most of the fermentation takes place in three stages.

• An initial stage during which time the yeast cells are multiplying.

• A very vigorous stage accompanied by bubbling and marked rise in temperature.

• Quiet fermentation that can proceed for quite along time at a lower and lower rate.

Fermentation time may range from 2-20 days depending upon numerous variables- types

and condition of fruits, type of wine made, and climatic conditions. Among others

temperature is quite critical to the fermentation process (Douglas and Considine, 1982).

The optimum temperature for fermentation of red wine is higher than that of white wine.

The optimum temperature is believed to be 21.1-27.4oC (Johnson and Peterson, 1974). At

temperature above 90oF (32.2oC), it is likely that wine flavor and bouquet will be injured.

24

High temperature also encourages heat tolerant bacteria to produce acid, mannitol and off

flavor (Douglas and Considine, 1982).

At the usual total sugar content of 19-23%, alcoholic fermentation proceeds rapidly and,

with alcohol tolerant strains of yeast, to completion, producing about 10-12.5% alcohol (by

volume) (Johnson and Peterson, 1974). If sugar content is greater than 23%, the high sugar

content may inhibit fermentation and the rate of fermentation, form glycine for example,

but is primarily derived from hydrolysis of naturally occurring pectin. The amount of

higher alcohols produced is less when ammonium phosphate is added prior to

fermentation. At very low concentration, the higher alcohols may play a desirable role in

sensory quality (Amerine et al., 1967). The oxidative conditions during fermentation favor

higher alcohol production (Guymon et al., 1961). Glycerol production is favored by low

temperature, high tartaric content and by addition of SO2. Most of the glycerol develops in

the early stages of fermentation. Most enologists consider that glycerol is of considerable

sensory importance because of its sweet taste and its oiliness (Gentillini and Cappelleri,

1959).

Acetaldehyde is a normal by-product of alcoholic fermentation. Acetaldehyde retention is

much greater when SO2 is added before the fermentation (Keilhofer and Wurding, 1960).

The primary source of acetaldehyde is from enzymatic process, i.e., in the presence of

yeast (Kielhofer and Wurding, 1960). Acetaldehyde reacts with ethyl alcohol to form

acetal, a substance with a strong aldehyde like odor, found very little in wines.

The tartaric, malic and citric acids of the must are found in the resulting wines but in

decreased amounts. They are important constituents of wine not only for their acid taste but

also they protect the wine from spoilage, maintain the color, and are themselves sometimes

attacked by microorganisms. Malic acid disappears during alcoholic fermentation to the

extent of 10 to 30%.

Succinic acid is a product of alcoholic fermentation. Lactic acid has a slight odor and is a

weak acid. It is a constant by-product of alcoholic fermentation, 0.04 to 0.75 g/L. Carbonic

acid constitutes a very special case for both still and sparkling wines. It has no odor and

very little taste. But it does have a feel and disengagement of the bubbles from wine

probably brings more oxygen away from the surface of wine (Amerine et al., 1967).

The end of fermentation is signaled by a clearing of the liquid, by a vinous taste and aroma,

and by a drop in temperature, and can be confirmed by checking degrees balling (sugar

residual) (Douglas and Considine, 1982).

25

2.12.4.3 Factors influencing fermentation

Various factors influence the course and consequence of the fermentation. These are

briefly discussed below.

2.12.4.3.1 Yeast culture

Yeast culture plays important role in winemaking. The pattern and end products of

alcoholic fermentation are greatly affected by the type of yeast culture utilized. The natural

wine culture and wine culture also produce wine with differing chemical constituents,

which may alter the organoleptic qualities of wines. Natural yeast flora of wine (i.e. raisin

culture) is a mixed culture, containing the wine strain of wine yeast, Saccharomyces

cerevisiae var. ellipsiodeus. It is said that the mixed flora produce wines with motalre

complex distribution of aroma components than wine fermented with S. cerevisiae (Reed,

1987).

Several species of Saccharomyces are the more important yeasts for winemaking, and

they constitute the true wine yeasts. The physiological properties of the yeast culture are of

importance in winemaking (Prescott and Dunn, 2004). The strain of yeast to be used for

alcohol fermentation should possess the following selective features:

1. Should be a sufficient strain. In other word, it should produce a large quantity of

alcohol.

2. It should be a fast growing strain.

3. It should have a high tolerance to alcohol, as well as to osmotic pressure.

4. It should possess uniform and stable biochemical properties.

2.12.4.3.2 Sugar and its concentration

Generally, monosaccharides are the normal and preferred substrates for the alcoholic

fermentation by yeast. The concentration of sugar determines the rate of fermentation.

High sugar concentration exerts high osmotic pressure which has negative effect on yeast

cells since both growth and fermentation activities are lowered. The inhibition is further

increased by ethanol which is formed during fermentation. The tolerance of high sugar

concentrations differs for various yeast species (Prescott and Dunn, 2004). The optimum

sugar concentration for maximum speed of fermentation is fairly low, perhaps only 1 or 2

percent. The maximum alcohol content in a single fermentation is obtained with musts of

from 25 to 35 percent sugar. The maximum alcohol content obtainable in normal winery

26

practice is about 16%; however, this varies with strain of yeast, temperature, conditions of

aeration, and method of conducting fermentation. The rate of fermentation is rapid below

25% sugar concentration of the must. Above 25% sugar concentration, the fermentation

gets retarded and at even higher concentration i.e. about 70%, most wine yeast will not

ferment the sugar. The inhibitory effect of high sugar is partly owing to the osmotic effect

(Amerine et al., 1967).

2.12.4.3.3 Sulphur-dioxide

The biological effect of SO2 comprises an inhibition of undesirable the microorganisms of

the must including acetic acid bacteria and lactic acid bacteria, which affect the course of

fermentation and quality of wine negatively. The chemical effect of SO2 is to bind

acetaldehyde which is formed during fermentation and which has undesirable organic

properties. Further, SO2 is added to prevent oxidative reactions of enzymatic or non-

enzymatic nature. For must with low total acidity, it is often desirable to use So2 to inhibit

malo-lactic fermentation. The SO2 is also desirable to suppress undesirable yeast species

such as Kloeckera apiculata and Metschnikowia pulcherrima on the must to retain good

wine quality. SO2 has also a direct effect on the course of the fermentation by

Saccharomyces yeasts. It delays the onset of fermentation, and the lag period is longer the

greater the amount of H2SO3 added to the must (Prescott and Dunn, 2004). However in

spite of its some desirable properties, SO2 is never added in base wine for brandy

manufacture due to its adverse effects on the quality of volatile acids is a advantage of

fermentations under CO2. Also, the metabolism of lactic acid bacteria is not inhibited

which may result malo-lactic fermentation and its undesirable consequences (Koch et al.

1953). In alcoholic media, the inhibition of yeast growth due to CO2 pressure is increased.

At relatively low CO2 concentrations of 0.6 to 1.8 g per liter, the growth is inhibited.

However, this effect depends also on the original yeast cell counts in the medium, and this

is also true at higher CO2 concentrations (Haubs et al., 1974).

2.12.4.3.4 Acids and pH

Yeasts are not very sensitive to the amounts of fixed organic acid but there may be some

effect of organic acids on the by-product of alcoholic fermentation (Amerine et al., 1967).

Microbiologically, the low pH (4.5 or below) is considered a favorable and selective factor

for wine fermentation. Most undesirable bacteria are inhibited at lower pH. For yeast, a pH

27

range between 3 and 6 is most favorable for growth and fermentation activity. A change in

pH can affect the formation of fermentation by-products. For instance, at higher pH values

the concentration of glycerine is increased. There is a positive relationship between the pH

of the must and the formation of pyruvic acid (Rankine, 1967A).Within the range from pH

3 to 4, there is a noticeable effect on lag phase and fermentation activity. At higher pH

values, the lag phase is reduced and fermentation activity increased (Ough, 1966A, B). The

effect of pH on growth and fermentation activity depends also on the concentration of

sugar and ethanol (Neish and Blackwood, 1951; Trautwein and Wassermann, 1931).

2.12.4.3.5 Temperature

The optimum temperature for fermentation by most wine yeasts is between 71.6oF and

80.6oF (Schanderl, 1959).However, temperature has many other effects besides its direct

effect on yeast growth and activity. These are due to losses of alcohol and aromatic

constituents at higher temperatures and to the by-products formed as well as to direct

effects on the efficiency of fermentation. Temperature affects yeast and consequently the

course of wine fermentation considerably, and a number of factors should be considered

for the selection of proper temperature. The fermentation metabolism of yeast can be

carried out within a rather large temperature range. Maximum value for S. cerevisiae is

near 40-45oC (White and Munas, 1951) and minimum temperature approaches to 0oC

(Osterwalder, 1934; Saller, 1955). Temperature also affects the formation of by-products.

With higher temperature within the 15-35oC range the concentration of glycerin, acetone,

2, 3-butanediol and acetaldehyde increase (Lafone, 1955; Rankie and Bridson, 1971).

Similarly, formation of acetic acid and other volatile acids, pyruvic acid and 2-ketoglutaric

acid also increase with increase in temperature. Whereas the formation of higher alcohols

decreases with increase in temperature. Whereas the formation of higher alcohols decreases

with increased the fermentation temperature having a maximum concentration at 20oC

(Dittrich, 1977). The temperature sensitivity of yeast is also affected by the ethanol formed

during fermentation.

2.12.4.3.6 Nitrogen

Yeasts readily assimilate amino acids but proteins can be hydrolyzed and used for cell

growth. Between 50 to 70% of total nitrogen of musts can be assimilated by yeasts

(Tarantola, 1955). The quantity of nitrogenous substances is entirely adequate for a

28

vigorous fermentation, and under normal circumstances there is even excess. Generally,

nitrogen is not required for grape juice, but fruit musts are often deficient in nitrogen. So,

urea or ammonium phosphate must be added. Nitrogen source addition is used to check the

formation of fusel oil, which is formed due to the action of yeast on amino acids. The

member countries of European economic country permit the addition of up to 30 gm/hL of

ammonium phosphate or ammonium sulfate to provide additional nitrogen in readily

assimable form (Reed, 1987).

2.12.4.3.6 Minerals and growth factors

The normal course of alcoholic fermentation requires magnesium, potassium, zinc, cobalt,

iodine, iron, calcium, copper and anions of phosphorous and sulfur. For growth alone

yeasts require copper, iron, magnesium, potassium, phosphorus, and sulfur. Adequate

amounts are supplied by grape and fruit juices. The presence of excessive iron (over

6p.p.m.) or copper hinders the fermentation of sparkling wines (Schanderi, 1959). Some

desirable growth factors for yeast are biotin, inositol, nicotinic acid, pentothenic acid, p-

aminobenzoic acid, pyridoxine and thiamine (Karki, 2001).

2.12.4.3.7 Oxygen

Oxygen is necessary for the maximum growth of yeast. But alcoholic fermentation is best

in an anaerobic condition. Here less of sugar is used by the yeast in respiration and there is

no oxygen to interfere with enzymatic activity. Aeration during normal fermentation may

results in wine with higher aldehyde content and darker color (Amerine et al., 1967).

2.12.4.3.8 Ethanol toxicity

Yeasts show some adaptive responses to the challenge of ethanol toxicity. The adaptive

behavior of yeast to ethanol response is also found in the relative resistance of the cells to

ethanol formed during fermentation, in contrast to the high sensitivity of cells to ethanol

supplementation. The most important response is that to the major toxic effect, the

disruption in the membrane permeability and change in fluidity. Increased ethanol

concentration can cause a decrease in yeast viability, reduction in yeast growth and

reduction in the rate of ethanol production. Slapack et al. reported that wine range of

cellular function is affected by ethanol, including inhibition of solute uptake, denaturation

and inhibition of glycolytic enzymes, uncoupling of oxidative phosphorylation and the

29

induction of peptic yeast strains. Saccharomyces cerevisiae may tolerate up to 17% by

volume ethanol concentration (Ribereau-Gayon and Peynaud, 1960).

2.12.5 Problems during fermentation

Different problems may arise during fermentation such as stuck fermentation, off-flavors

production, methanol production, and contamination with undesirable microorganisms.

There are two general classes of problems for winemakers that can arise during the

alcoholic fermentation: sluggish or stuck fermentations and off-flavor production.

2.12.5.1 Stuck fermentation

Premature arrest of alcoholic fermentation is an occasional but continuing problem for

wine makers. It may manifest itself as sluggish activity during mid and later phases of

alcoholic fermentation. Whereas in other cases cessation of fermentation activity may be

abrupt. In either case, the resultant wine may have perceivable (an often objectionably

high) levels of sugar with decreased production of alcohol. The causes of sluggish and

stuck fermentations include fermentation at temperature extremes, nutritional deficiencies,

osmo-regulation, ethanol toxicity, and in low-temperature fermentation, long-term

anaerobiosis. Such problem is generally treated with addition of deficient nutrition such as

nitrogenous compounds e.g. di-ammonium phosphate, and provision of optimum

temperature along with aeration for the reactivation of yeast (Kenneth, 1997).

2.12.5.2 Production of off-characters

Saccharomyces strains have also been implicated in the production of certain volatile

phenols in wine (Chatonner et al. 1993). During fermentation, production of off-characters,

off flavors or off aroma compound detract from overall wine quality. An important class of

spoilage compounds is sulfur-containing volatiles which have very unpleasant odors.

Although they are produced in traces, they cause damaging effect on the quality. Foremost

among the sulphur containing volatiles is hydrogen sulfide (H2S) (Baulton, 1997). To

prevent this problem very careful control of fermentation conditions is worthwhile.

30

2.12.5.3 Methanol production and its quality

Small amount of methanol is inevitably present in brandies, but if significant quantity is

formed during fermentation, large accumulation may result in distilled product which may

be detrimental to health. Methanol is produced by the demethylation of pectin present in

the substrate by fruit pectin esterase enzymes. Yeast does not form an enzyme capable of

hydrolyzing pectin and consequently the reaction does not commonly occur in fruit

fermentation but the fruit itself contains this enzyme. Methanol is very toxic to humans

with a fatal internal dose of 60-250 ml. The methanol production can be prevented by

inactivating the fruit pectin esterase enzymes before fermentation with heat treatment and

by obstructing the contamination of the fermentation with molds and bacteria.

2.12.5.4 Activity of undesirable microorganisms

Low pH values of the must are inhibitive for most of the bacteria but two types of bacteria

viz. lactic acid bacteria and acetic acid bacteria, which may start malo-lactic fermentation,

metabolizing the malic acid to lactic acid and CO2. The malo-lactic fermentation is

desirable in some cases, but generally it lowers the acidity causing rise in pH and

susceptibility of wine to further spoilage. Similarly, lactic acid bacteria ferment the sugar

into lactic acid and other volatile acids, causing an undesirably high acidity. Acetic acid

bacteria are aerobic and if they get favorable condition, they grow on the must and change

the produced ethanol into acetic acid (Reed, 1987).

2.13 Racking

After completion of fermentation, the wine must be separated from the dead cells, which

decomposes and give off flavors and odors to wine (Andrew, 1980). This dead yeast settle

at the bottom of the fermentation vessel and the wine is carefully transferred (siphoned) to

other vessel without disturbing the dead yeast leaving some wine at the bottom called lees.

The advantages of racking are:

• It helps removing CO2.

• It raises O/R potential, which retards the formation of H2S.

• It clarifies the wine.

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2.14 Clarification and fining

Clarification by conventional racking process is a long process. To hasten this, certain

agents commonly called fining agents are added during racking. Fining is a traditional

method of bringing about clarification. Fining agents may be used during ageing as well.

They not only clarify the wine (by physical adsorption) but also help to remove excess

tannins (Rai, 2005). The purposes of clarification and fining during wine processing

include removal of excessive levels of certain wine components, achieving clarity, and

making that clarity stable especially from a physiological viewpoint. The materials used for

these reasons are collectively referred to as fining agents. Examples of such fining

reactions are: This is a process of converting cloudy wine into clear wine. This may be

done by adding gelatinous substances such as icing glass, egg white, bentonite and tannin.

Pectin hydrolyzing enzymes are also used in the clarification of wine (Andrew, 1982).

After clarification, the wine is passed through fine filters for filtration. The pad filters are

most common. In order to increase filter life, diatomaceous earths are added to wine during

filtration. These mix with mucilaginous materials and maintain the capacity of the filter for

longer times i.e. increase filter capacity. Recently membrane filters have been widely

employed for wines. These have uniform but small pore size so that a very large percentage

of the filter-surface is available for filtration. They also greatly reduce the number of

bacteria (Johnson and Peterson, 1974).

2.15 Stabilization of wine

Because of the unknown nature of the wine, it is generally good practice to stabilize them

against microbiological changes by use of antiseptics such as sorbic acid or its potassium

and sodium salts in amounts ranging from 300 to 1000 p.p.m. An alternative is to

pasteurize the wines after bottling. Another alternative may be to flash pasteurize, fill into

clean bottles, and seal using clean closures (Chan, 1983).

2.16 Maturing and aging of wine

This is one of the most interesting and one of the most important, yet one of the most

complex processes of wine making. This takes place naturally by retaining the wine in oak

barrel for one or two years to gain maturity and pick up soft and mellow characters from

the oak wood. Andrew (1980) found that maturation can be artificially induced by

32

agitation, heating, refrigeration and electrical impulses. The bouquet and aroma of wine are

developed during aging (Banwart, 1987).

Aging of wine for long periods of months to years produces desirable changes in body

and flavor of wine. In addition, malic acid of grape juice is fermented by lactobacilli during

aging to give lactic acid and carbon dioxide and also decrease the acidity (Sivasankar,

2005).

Aging is one of the most interesting and important yet one of the most complex process

in winemaking. Newly fermented wine is cloudy, harsh in taste, yeasty in odor and without

the pleasing bouquet that develops later in its history (Amerine et al., 1967). The wine is

aged to reduce the acidity and to develop a characteristic bought. The main acid in most

wines is tartaric acid but, in some red wines, malic acid is present in a high concentration.

In these, secondary malo-lactic fermentation by lactic acid bacteria converts malic acid to

lactic acid to reduce the acidity and to improve the flavor and aroma. Lactic acid bacteria

produce small amount of aldehydes and lactic and acetic acids, which give the product a

characteristic aroma and flavor (Fellows, 1990). Aging of wines improves the flavor and

bouquet due to oxidation and formation of esters. These esters of higher acids formed

during aging give the ultimate pleasing bouquet to the well aged wine. Aged wine may be

polished by filtration to give a clear, bright appearance prior to bottling (Desrosier and

Desrosier, 1978).

2.17 Bottling

This is done before the blended wine has lost its bouquet, fineness, quality and color.

Bottles are cleaned and dried with hot air. Cool and dry weather is chosen for this purpose.

Bottles are closed with a fine, soft supple cork applying pressure with the finger. Corks are

finally sealed with Spanish wax (Andrew, 1980).

2.18 Pasteurization

Pasteurization is the process used to kill microorganisms present in the wine so that

fermentation is stopped. Pasteurization is applied in one of the three ways:

1. By flash pasteurizing and returning to the storage tank.

2. Flash pasteurizing into the final bottles and

3. Pasteurization by heating the filled and sealed bottles.

33

The time temperature relationship for pasteurization of wine is: vegetative yeast cells are

killed at about 40oC while yeast spores are only killed at 57oC (Desrosier and Disrosier,

1978). The quality of some wine is reduced by pasteurization while that of other may be

improved. Pasteurization inactivates the enzymes but injures the quality of the product

(Johnson and Peterson, 1974).

2.19 Finishing

The traditional method of finishing the wine was to turn the bottles on end, place them in

racks at about 45o angle and turn them to the left and right daily to get the yeast deposit

into the neck of the bottle and on the cork. The process is called riddling “reumage”. The

temperature of the whole bottle is then reduced to about 30oC to 40oC. The neck of the

bottle containing the yeast deposit is then frozen (by placing in brine or other freezing

solution). When the cork is removed, the solid plug containing the yeast is ejected. This is

called disgorging (Johnson and Peterson, 1974).

2.20 Storage and ageing of wine

Actually racked wine contains some suspended particles. Racked wine is flash pasteurized

in order to coagulate the suspended particles. After pasteurization it is kept at room

temperature for 1-2 days, then at -3 to -4oC for 2-5 days. Then it is filtered in the cold state

(-3 to -4oC) and transferred to storage tank. Wines are aged in bottles, barrels, tanks or

puncheons. The tank may be wood, concrete or metal ((Dhakal, 1988).

A wine cellar should be maintained at a uniform temperature of 60oF and a humidity of

50%. When stored, each bottle of wine must be laid in a horizontal position so that the

wine keeps the cork moistened. The room should be darkened, free from dirt, and

mechanical or sound vibrations (Smith and Milner, 1974).

The purposes of storage and ageing are:

• For the development of body, flavor and bouquet,

• To aid the clarification.

Ageing of wines improves the flavor and bouquet due to oxidation and formation of

esters. These esters of higher acids formed during ageing give the ultimate pleasing

bouquet to well aged wine. Aged wine may be polished by filtration to give a clear, bright

appearance prior to bottling (Desrosier, 1982). The period of ageing depends upon quality

of wine. For example- dry wines are aged for 2 years, and fine wines for 5 years.

34

2.21 Wine made from different raw materials

Wine can be prepared from different raw materials. It can be made from different fruits

(grapes, apple, pears, bael, guava, banana, pineapple, pumpkins, etc.) and roots (ginger,

potato, calocassia, etc.). Many researchers have prepared wines from different raw

materials in CCT, Hattisar, Dharan. Dhakal, (1988) prepared wines from ginger and banana

having varied recipes fermented by yeast isolated from murcha. TSS and pH were

optimized for banana and ginger wines. Banana wine was found best having 17oBrix TSS

and 4.5 pH. Similarly, ginger wine having 22oBrix and 5pH was found best. The alcohol

content of ginger and banana wines was found to be 7.06% and 8.39% (v/v) respectively.

He found that 17-20% sugar concentration, 100-200 p.p.m. SO2, 25-30oC temperature and

4.5-5pH were suitable for appreciable starter activity.

Shakya, (2002) prepared bael wines from bael pulps obtained by hot and cold extractions.

The bael wine prepared from hot extracted pulp was found to be better than the cold

extracted. Bael wine from mash of 25% pulp content was the best. Fining agents tannin and

gelatin produced exceptional clarity. The bael wine contained 11%(v/v) alcohol, 0.12%

volatile acidity, 0.49% fixed acidity, 9.5g/100L methanol, 225 mg/L esters, 280 mg/L

aldehydes and 193 g/100L total higher alcohol.

Gubhaju, (2006) prepared wine from Rhododendron flower using wine yeast. Effect of

fresh and dried flower on quality of wine was studied. She found that use of raisin and

brown sugar did not improve the quality of wine. TSS of 20oBrix and pH of 4.5 were found

optimum for the preparation of Rhododendron wine using dry flower and S. cerevisiae.

Average alcohol content and esters of the prepared wine were found to be 11.03% (v/v)

and 3.81 mg/L respectively.

Dhakal, (2007) prepared wine from palm sap using baker’s yeast. Wine made from mash

with 4.5 pH and 20oBrix was found to be most acceptable. Total solids (%m/v), sp. gr. (at

25oC), alcohol content (%m/v) and ash content (%m/v) were found to be 0.13, 0.9529, 36.5

and 0.005 respectively. Similarly, total aldehydes (as g acetaldehyde), esters (as g ethyl

acetate), fusel oil (as g amyl alcohol), total acidity (as g lactic acid), volatile acidity (as g

acetic acid) and fixed acidity (as g lactic acid) were found to be 0.525, 44.1, 84.46, 281,

162 and 37.47 per 100 L of ethanol respectively.

35

2.22 Fining agent and its types

The material which is used to achieve clarity of wine is called fining agent. The fining

agent has several adsorption sites on each head or molecule and a number of solute

molecules are either adsorbed to its surface or exchanged into its inferior. Several of the

agents currently in use (such as the proteins and the gums) are colloidal in nature and as the

adsorption occur, resulting in precipitation of the solute/agent complex from the solution.

The amount of solute removed by a certain addition of an agent will depend on the

solute/agent pair as well as the concentration of the solute in the wine and the quantity of

the agent added. The fining agents can be classified into the following groups:

2.22.1 The Proteins

The purpose of adding a protein preparation to wine is to soften or reduce the wine’s

astringency or reduce its color by the adoption and precipitation of polymeric phenols and

tannins. Although it is rarely practiced today, white wines can be clarified by adding a

protein followed by tannin due to the co-precipitation that occurs. All of these proteins

come from natural sources usually in a partially purified form. The four most commonly

used proteins for wine precipitation are casein, gelatin, albumin, and isinglass. Their

properties are summarized in Table 2.4.

36

Table 2.4 Typical ranges of application of fining agents.

Agent Common Range of Application (mg/L)

White Table Wine Red Table Wine

Casein 60 to 120 60 to 240

Albumin N/A 30 to 240

Isinglass 10 to120 30 to 240

Gelatin 15 to 120 30 to 240

Bentonite (Na form) 120 to 720 N/A

Silica Sol 40 to 200 40 to 200

PVPP 120 to 240 120 to 480

Agar/ Alginate 120 to 480 120 to 480

Activated Carbon 120 to 600 120 to 480

(Source: Boulton et al., 1997)

2.22.2 The Earths

There are a number of clays: silica, alumina matrix with exchangeable cations, bentonite

etc. The clays (silica, alumina matrix) have been considered as alternatives to bentonite and

these include kaolin, Spanish earth. They generally have a lower adsorption capacity and

therefore are not preferred in winemaking applications.

2.22.2.1 Bentonite and its properties

Bentonite is widely used for the adsorption of proteinaceous material from wines.

Bentonite was originally introduced as a means of clarifying wines and vinegars (Saywell,

1934A.B) and its application to inducing heat stability in white wines came several years

later. Bentonites are mined from several areas of world and come in different levels of

purity, particle size, adsorption and swelling capacity. Bentonite is natural clay that is

classified as a montmorillonite, with a general composition of the form: Mg, Ca, Na,

Al2O3.5SiO2.nH2O (Siddiqui, 1968). The source of the bentonite influences its properties

slightly and the main difference lie in the proportion of Mg++, Ca++, and Na+ in the

lattice. Bentonite has a structure which expands after contact with water and preparations

have optimum adsorption after two days of soaking. The limited cation exchange capacity

37

poses particular problems with the removal of negatively charged and neutral protein

fractions from wines. Bentonite is essentially inert with respect to the phenolic components

in wine except for cationic anthocyanins. There is little effect of temperature on the

adsorption (Jacob, 1968; Blade and Boulton, 1988). Bentonite may indirectly bind phenols

that have complexed with proteins. Bentonite may affect red wine color by binding with

positively-charged anthocyanin monomers and may result in color decrease depending

upon the age of the wine.

Bentonite may also remove more color in younger wines because of the greater action on

the colloidally colored material found in the younger wines (Bergeret, 1963). Addition of

bentonite to red wines at levels of 6 to 12 g/hL (0.5 to 1 lb/1000 gal) improves membrane

filterability due to reduction in colloidally suspended particles. Bentonite fining of juice

may remove peptides and some amino acids, potentially affecting rate and completion of

fermentation. Bentonite fining is known to indirectly prevent or impede formation of

copper, and possibly iron casse, this is probably due to removal or reduction in levels of

proteins and peptides known to be involved in the formation of haze and precipitate.

2.22.2.2 Preparation of bentonite

Preparation of bentonite greatly impacts its activity toward proteins. In solution, bentonite

swells to many times its dehydrated dimensions. Its activity is much like that of a

multiplateted, ong-chain, linear, negatively-charged molecule (Singleton, 1967). During

the hydration phase, charged platelets repel each other and begin to separate. Water

molecules partially neutralize and separate exposed surfaces, exposing a large matrix of

reactive surface. The presence of water molecules within the network prevents flocculation

and precipitation. The water used in hydration phase should have a low mineral content.

Dissolved metal cations present in slurry water preferentially replace sodium ions on the

clay surface and detrimentally affect the hydration, viscosity, and binding capacity of the

bentonite (American Colloid Co.). Typically, the bentonite-to-water ratio for slurries is 5-

6% (w/v). Heating non-agglomerated bentonite allows the platelets to fully separate and

slurry resembles a gel. Bentonite additions, especially those exceeding 48g/hL (4 lbs/1,000

gal), may strip wine flavor, body, and in the case of young red wines, significant color.

Further, it may impart an earthy character to the wine. Some winemakers choose to ferment

settled juice in contact with benttonite to aid protein stability and to eliminate or reduce the

38

amount of bentonite needed to stabilize the wine. The procedure for the fermentation of

white juice in contact with bentonite used in USA is as follows:

1. Settle juice to remove non-soluble solids by refrigeration and/ or fining agents (A

high solids level could foul the bentonite utilized during fermentation and reduce

overall efficiency). Add the desired quantity of bentonite in-line while racking into

the fermentor.

2. Add yeast nutrient and any needed sugar and/or acid where allowed.

3. Add yeast inoculum to juice surface.

2.22.3 Synthetic Polymers

Materials such as polyglycine, polyamide (nylon), and polyvinyl-polypyrrolidone (PVPP)

are synthetic products with available carbonyl oxygen atoms at the surface that act as

adsorption sites. Nylon and PVPP are both insoluble white powders that have been used in

wine. The more efficient adsorption of PVPP has led to its preferential use (Caputi and

Peterson, 1965; Rossi and Sigleton, 1966). The agents are generally added in a batch

treatment and commonly settled or filtered out of the wine and discarded after one use. The

catechins are involved in the chemical browning of white wines and are thought to be

particularly bitter in nature above their sensory threshold level of 20 mg/L (Singleton and

Noble, 1976).

2.22.4 The Colloids

The colloids that are used for wine clarification are classified into two groups:

2.22.4.1 Natural Polysaccharides

The polysaccharides, agar and acasia (gum arabic), both have protective colloid properties

and can partially neutralize surface charges on other naturally dispersed colloids, thereby

allowing them to either dissolve or to coagulate. The polysaccharides of this kind are

especially useful in neutralizing the charge of other haze components of what are generally

referred to as protective colloids, one polar or charged entity is adsorbed to the outer

surface of another, causing the overall complex to repel like species and causing a

suspension to occur. In wines and juices natural polysaccharide contents are in the range

200 to 1000 mg/L (Usseglio-Tomasset, 1976; Villettaz, 1988; Feuillat, 1987;

Wucherpfernnig and Dietrich, 1989).

39

2.22.4.2 Ferrocyanide Preparations

The application of ferrocyanide salts for the removal of transition metal cations from wine

has been widely practiced in Europe for more than 50 years. Sometimes referred to as

Moslinger fining, after its inventor, the form used is potassium ferrocyanide, K4Fe(CN)6.

The ferrocyanide salts of most metals are blue (hence the term blue-fining), sparingly

soluble, and when used properly, the residue should be less than 0.02 mg/L (Wurdig and

Woller, 1989).

2.22.5 Alternative Methods of Metal Depletion

The removal of metals could be achieved by the use of immobilized forms of ferrocyanide

or alternatively, other chelating materials. There are presently a number of chelating resins

available commercially, but several of them have limited chelating functions at pH values

less than 4. Alternative polymeric materials based on 1-vinylpyrrolidone and 1-

vinylimidazole have also been suggested for the removal of metals from wine (Fussnegger

et al., 1992). The polymer containing a ratio of the pyrrolidone to imidazole of 9:1 gave the

best results, although a significance pH rise occurred at treatment levels that were required

to lower the copper content to below 0.2 mg/L, the level generally considered to be

acceptable.

2.22.6 Activated Carbon

The activated carbons have high and broad affinities particularly for benzenoid and non-

polar substances. They are used to remove color pigments and a wide range of phenolics

but are rather nonselective in their adsorption (Singleton, 1964). They do not adsorb

substances such as sugars and amino acids which are highly water soluble. They are not

generally used and offer no benefits to most wines. They adsorb a range of other trace

constituents including some vitamins and this could have secondary effects on

microbiological stability. The carbons are usually sold as either decolorizing or

deodorizing forms. The treatment levels are determined by a dose-response trial on the

wine in question with the specific carbon.

2.22.7 Silica Suspension

Silica suspensions, or sols, are most often used in clarification and settling applications,

particularly with apple and fruit wines. In such cases, the silica sol preparations which vary

40

with the loading and wine concerned. The treatment limit in the United States is 2.4 g/L of

a 30% by weight silica solution.

2.22.8 Copper Sulfate

Copper sulfate is used to remove H2S and thiols from wines and the level of addition

should be less than 0.5 mg/L. The residue of copper should be less than 0.5 mg/L in the

United States and 0.2 mg/L in the most countries. Copper sulfate can be used in

conjugation with a sulfite/ascorbate addition to remove disulfites from wines.

2.23 Components of wine

Being produced from complex material, wine contains innumerable components. The chief

component of wine is ethyl alcohol but many other components specially present on the

mash of the fruit or may form during fermentation. Further more, other components are

formed during ageing or are extracted from the wood.

2.23.1 Ethanol

Ethanol (C2H5OH) is completely miscible with water and is an excellent solvent for

odorous materials such as essential oils, esters, tannins, various acids and certain other

organic compounds. It has a slight sweet taste and moderates the taste of acids. A de-

alcoholized wine is much more tart than the same wine with its alcohol. The odor

threshold, according to Berg et al. (1955B) is 0.004 to 0.0052 gm. per 100 ml. Most of the

alcohol in wines result from fermentation, a little may result from hydrolysis of glycosides

during prolonged ageing. The alcohol content varies from 5% up to 21% (v/v). Natural

wines contain 8.5 to 16% ethanol by volume. The legal limits for alcohol vary markedly

for different countries and are usually related to the tax structure (Amerine et al., 1967).

2.23.2 Methanol

It is generally agreed that methanol, CH3OH, is not generally produced by alcoholic

fermentation, but is primarily derived from hydrolysis of naturally occurring pectin in

fruits by pectolytic enzymes. Methanol is shown to be higher in wines from macerated

grapes than from non-macerated fruit. There is no change in the methanol content during

the fermentation of a white must. Methanol increases in red must during fermentation. The

methanol content is not related to the pectin content and that fermentation does not change

41

the methanol content. The amount of methanol in wines ranges from traces to 0.635 gm.

per liter, average about 0.1 (Amerine, 1954). The sensory importance of methanol has not

been studied.

2.23.3 Higher alcohols (Fusel oils)

The higher alcohols constitute a part of flavor in wine. The higher alcohols account for the

major portion of the products of yeast metabolism. At very low concentrations the higher

alcohols may play a desirable role in sensory quality. Higher concentrations lower the

quality of the wine (Wagener and Wagener, 1968). But the complexity of the substrates

rarely permits clear-cut conclusions (Guymon and Heitz, 1952). The higher alcohols

always present are 1-propanol, 1-butanol, 2-butanol, 2-methyl-1-propanol, 2-methyl-1-

butanol, 3-methyl-1-butanol, 1-pentanol, and 1-hexanol. Higher alcohols which are not

present or only in traces amounts include 2-propanol, 2-methyl-2-propanol, 3-methyl-2-

butanol, 2-pentanol, 3-pentanol, and 2-hexanol (Usseglio-Tomasset, 1964). The oxidative

conditions during fermentation favor higher alcohol production. The presence of pomace,

as in red wine production, aerates the wine and thus leads to greater amounts of higher

alcohols (Guymon et al., 1961). The amount of higher alcohols produced is less when

ammonium phosphate is added prior to fermentation.

2.23.4 Carbonyl compounds

Carbonyl compounds are always present as part of the aroma of wine (Rapp et al., 1973;

Schreier et. al. 1976; Webb, 1967). These are mainly aldehydes which are formed during

fermentation as intermediates in the formation of alcohol from sugars and amino acids; but

some ketones also occur (Lehtonen and Suomalainen, 1977). Acetaldehyde is a normal by-

product of alcoholic fermentation. Aldehyde retention is much greater when SO2 is added

before the fermentation and especially high when SO2 is added during fermentation. The

primary source of aldehydes is from enzymatic processes, i.e., in the presence of yeast.

Non-enzymatic production of aldehyde in white wines is very small, especially in the

absence of iron (Kielhofer and Wurdig, 1960B). The amounts found in newly-fermented

wines are below about 75 mg per liter, it has little sensory importance, especially so since

most of our wines have SO2 added which fixes most of the aldehyde. However, it has a

pronounced odor and a threshold is of only 1.3 to 1.5 mg. per liter in water (Berg et al.,

1955A). But the thresholds are of 100 to 125 p.p.m. The diketone, diacetyl, which is a well

42

known flavorant in many foods, occurs in wine where it is produced by yeasts during the

alcoholic fermentation. The diacetyl produced during the fermentation is immediately

metabolized by the yeasts (Suomalainen and Ronkinen, 1968).Its content in fermenting

mash, therefore, is zero. It rises slightly in the wine after the yeast is removed. Sherry has

usually high concentrations of aldehydes, yet there seems to be no correlation between

their concentration and the quality of the wine (Amerine et al., 1980).

2.23.5 Esters

Wine, as well as other fermented beverages, is rich in esters. Esters are very important for

the aroma of the wine because of their intense aroma at relatively small concentrations. A

total of more than 50 various esters have been identified in wines. For beverages with

relatively low ethanol concentrations, such as wine, enzymatic reactions are responsible for

the formation of esters, while strictly chemical reactions do not proceed (Nordstrom,

1964). Yeasts determine the composition and the amounts of the esters formation. The

effect of esters on wine quality is difficult to quantify. There is a tendency toward higher

quality for wines with higher ester concnetrations (Wagener and Wagener, 1968). Only

ethyl acetate seems to be important-below about 200 mg. per liter it may be a desirable

odor but above this it appears to give a spoiled character to the wine. Both neutral and acid

esters are found in wine. The total esters in various wines vary between about 200 and 400

mg. per liter (as ethyl acetate).

2.23.6 Acids

The acids includes both volatile and fixed. Besides acetic acid and lactic acid, which are

normal by-products of alcoholic fermentation, formic, butyric, propionic and traces of

other fatty acids are present. Acetic acid is not only a by-product of alcoholic fermentation

but during the course of fermentation an appreciable amount may be utilized by the yeast.

Wine always contains some volatile free acids, mainly aliphatic acids. They occur in higher

concentrations in spoiled wines because of the activity of film forming yeasts, and

particularly of acetic acid bacteria and lactic acid bacteria. Volatile acidity refers to the

volatility with steam of the fatty acids. The volatile acidity thus includes the fatty acids in

the series starting acetic but excludes lactic, succinic, carbonic and sulfurous acids. The

volatile acids are produced mainly during the initial stage of fermentation, and more is

formed in presence of oxygen than in absence (Amerine et al., 1967). The determination of

43

volatile acidity as an indication of spoilage has become a part of the legal requirements for

wine standardization. The amounts of acetic acid produced during alcoholic fermentation

are small-usually less than 0.03 gm. per 100 ml. Bacterial action before, during, and after

fermentation may lead to much higher quantities of alcohol or occasionally to bacterial

attack on fixed acids include tartaric, malic, and citric acids. These acids of the must are

found in resulting wines, but decreased amounts. They are important constituents of wine

not only for their acid taste but also because they protect the wine from spoilage, maintain

the color, and are themselves sometimes attacked by microorganisms (Amerine et al.,

1967). Both pH and titratable acidity are important in determining sensory response to

sourness.

2.23.7 Glycerol

Glycerol is a by-product of alcoholic fermentation. Glycerol production is favored by

lower temperatures, higher tartaric acid content, and by addition of SO2. Glycerol is of

considerable sensory importance because of its sweet taste and its oiliness. A dry table and

red table wines contain 0.3 gm. and 0.8 gm. per 100 ml of glycerol respectively (Hienreiner

et al., 1955A).

2.23.8 Minerals

The inorganic constituents of wines are of considerable biochemical, technological and

physiological importance. Many are needed in alcoholic fermentation. Some are significant

in human nutrition-usually desirable so but in a few cases from toxic point of view so that

legal maxima are prescribed. Wines contain anions (bromide, chloride, fluoride,

phosphates, silicate, sulfate etc.) and cations (aluminium, arsenic, cadmium, calcium,

copper, iron, magnesium, manganese, sodium, potassium, rubidium, etc.). The anion and

cation balance of wine is made in winemaking (Amerine et al., 1967).

2.23.9 Pectins and gums

Finished wines have 0.3 to0.5 gm. per 100 ml of pectins and gums. The gums of wines are

generally arabans, anhydrates of arabinose and galactans. To remove pectins, pectolytic

enzymes are frequently used-either before or after fermentation. They do aid filtration and

reduce the pectin content. They also raise the galacturonic acid content and slightly

increase the methanol content.

44

2.23.10 Water and sugar

Their amounts are variable. Dry wine contains less than 0.12% sugar and sweet wine

contains up to 7% sugar.

2.24 Yield

The yield of alcohol is of great importance to the wine maker. According to the Gay-

Lussac equation theoretical yields of 51.1 per cent alcohol and 48.9 per cent carbon dioxide

are possible. It is obvious that this is biologically unobtainable and in practice will depend

on a variety of factors-amount of by-products, amount of sugar used by yeasts, sugars used

by other micro-organisms, alcohol lost by evaporation or entrainment (which in turn

partially depends on the temperature and the rate of fermentation), presence of air, stirring

or other movement of fermenting mass, and other factors. The yield is therefore not a fixed

quantity but will vary depending on the variables mentioned above. The best practical

yardstick for yield is therefore empirical studies made under carefully controlled conditions

(Amerine et al., 1967).

Kirk (1967) has found that roughly each percentage of sugar fermented yield 0.55% of

alcohol by volume. Under special condition of simulation, 16-18% alcohol can be reached,

but normally in commercial operation, 13-15% is the maximum (Johnson and Peterson,

1974).

2.25 Wine defects and spoilage

Wine defects include turbidity, cloudiness, precipitation and loss of color, coloration due to

metals such as iron, tin and copper and their salts. Microorganisms, both aerobic and

facultative, cause wine spoilage by imparting cloudiness, bitter characteristics, ropiness,

undesirable flavors high volatile acidity and low alcohol content. Wild yeasts, molds and

bacteria of the genera Acetobactor, Lactobacillus, Leuconostoc and Pediococcus cause

spoilage if wine (Sivasankar, 2005). Leuconostoc mesenteroids causes sliminess or

ropiness of wine as well as it increases the viscosity of wine (Parry, 1973).

Wine defects may be caused by micro-organisms or non microbial causes. Defects

include those due to metals or their salts, enzymes and agents employed in clearing the

wine. Iron, for example, may produce a sediment known variously as grey, black blue or

ferric casse and in white wine may be responsible for white precipitate of iron phosphate

termed white casse. Tin and copper and their salts have been blamed for cloudiness. White

45

wines may be turned brown and red wines may have their color precipitate by peroxidases,

an oxidizing enzyme of certain molds. Gelatin, used in clarifying wines, may cause

cloudiness (Frazier, 1967).

2.26 Wine and its health benefits

No other drinks, except water and milk, have earned such universal acceptance and esteem

throughout the ages as has wine.

Most of the world’s wines have been consumed with food or as food itself. Wines

reputedly aided in maintaining health not only because of their own nutritive value, but

also because they replaced inadequate wine or otherwise unsatisfactory water supplies,

(Peterson, 1971).

Medical studies have revealed a lot of health benefits of drinking wine.

We have probably heard about the 'French Paradox'. This finding concerns wine and health

and shows that the French have a lower incidence of coronary disease as we find in the

United States, despite the rich, high fat foods found in French cuisine. Many experts

attribute this to the regular consumption of wine in the French diet. Some wine and health

medical studies have shown that an occasional glass of wine can reduce the risk of heart

disease or stroke. Moderate wine consumption can reduce the risk of death by

cardiovascular disease, stroke, and cancer. Studies are still being conducted on the red wine

health benefits. Wine contains flavanoids, anti-oxidants that help to prevent free-radicals

from damaging cells. One in particular helps to prevent hardening of the arteries. Wine also

contains a substance called reservatrol. This substance has been shown to boost the

immune system, block cancer, and protect against cardiovascular diseases. These

substances are found in all wines, but there are more in red wine than in white wine, that is

why we are speaking particularly about red wine health benefits. With red wines, the

grapes are pressed, and the juice sits for a while with the grape skins and stems still

present. Because of this, the grape juice has a chance to leach more of the flavanoids and

resveratrol from the grape skins. So, red wines will have a higher concentration of anti-

oxidants than white. (Source:http.//www.metalimagination.com/historyofwine.html).

White wine contains more riboflavin than red wine (Morgan et al., 1987). While studying

the vitamin content in fermenting musts, Castor found that riboflavin increased,

panthothenate usually decreased, vitamin B6 slightly decreased, and biotin was largely lost.

Lucia (1954) reported that when wines are taken along with a good and balanced diet, their

46

content of thiamine, riboflavin, pantothenate, niacin and vitamin B6 contribute to total

nutrition. Wine has an anti-inflammatory and anti-oxidant substance called resveratrol,

which is effective against lung disease (Srilakshmi, 2007).

2.27 Brief Introduction of Ginger wine

Ginger Wine is an alcoholic beverage made from a fermented blend of ground ginger

(Zingiber officinale Rosco.) and raisins. The word drink is primarily a verb, meaning to

ingest liquids, see Drinking. ... Ginger is usually used to flavor a wine. It also contributes to

color of wine. Powder ginger is not used in wine making. It has many health benefits.

Ginger wine can be consumed by blending with whisky, brandy or rum. The first

documented appearance of Ginger Wine occurred with the foundation of 'The Finsbury

Distilling Company' based in the City of London in 1740.

(Source:http.//www.statemaster.com/encyclopedia/Ginger-wine).

Part III

Materials and methods

3.1 Raw Materials

The raw materials viz. raisin, ginger and sugar were procured from local market of Dharan.

Baker’s yeast (Saccharomyces cerevisiae) manufactured by Vitabe-Belgium bvba

Langerbruggekaai 37, 9000 Gent-Belgium was collected from Gorkha Department Store,

Itahari and true wine yeast (Saccharomyces ellipsoideus) was obtained from Makalu wine

industries (P.) Ltd., Basantapur, Tehrathum.

3.2 Optimization of TSS and amount of ginger in the fermentation mash

3.2.1 Preparation of mash

Ginger was washed and removed its skin by a knife. Raisin was sorted and washed in tap

water. The ginger and soaked raisin were flaked by iron mortar and pestle. The sugar syrup

was made with warm water, and then stained by muslin cloth.

The different fermentation mashes having 10% raisin of the total mash and 4.5 pH were

prepared as follows:

Table 3.1 Composition of different mashes.

Mash Ginger (%) TSS (oBrix)

1 1 16

2 1 20

3 1 24

4 1.5 16

5 1.5 20

6 1.5 24

7 2 16

8 2 20

9 2 24

The TSS of these mashes was adjusted with addition of sugar syrup and pH was adjusted to

4.5 by using 5% sodium bi-carbonate and citric acid solutions with a pH meter (Henna,

48

Portugal). All these mashes were pasteurized by heating t0 70-72oC for 15 minutes and

brought to room temperature (30oC) by cooling in tap water.

3.2.2 Pitching and agitation

One set of the prepared mashes was inoculated with baker’s yeast and another one was

inoculated with wine yeast at the rate of 1g/ Liter, and was filled into clean and boiled

bottles up to 2/3rd of their capacity. The bottles were then plugged by cotton, agitated and

kept for fermentation at room temperature.

3.2.3 Fermentation

The progress of fermentation was monitored by measuring the drop of TSS. After 5 days of

pitching, the TSS of the fermented mashes was measured by hand refractometer every day

till constant TSS was obtained. After 11 days from pitching, the fermentation was assumed

to be completed when the TSS ceased to drop further.

(a) (b)

Fig 3.1 Fermentation of ginger wines.

3.2.4 Racking, pasteurization and bottling

The clear wines were siphoned off from the lees in still pot using a sterilized polythene

pipe, and covered with metal dish and pasteurized by heating to 70oC for 15 minutes and

cooled to room temperature (30oC) by cooling in tap water. Bentonite suspension was

49

added at the different concentrations (0.5, 1 and 1.5g/L) for the clarification of the ginger

wines. The cold wines were racked and filled in to the pre sterilized bottles and kept in

room until needed for further analyses.

Fig. 3.2 Flow sheet for the preparation of ginger wine

Screening

Weighing

Washing and soaking

Flaking

Ginger

Washing

Skin removal

Weighing

Flaking

Mixing

Adjustment of TSS and pH

Pasteurization at 70oC/15 minutes

Cooling to 30oC

Addition of starter culture @ 1 g/L

Fermentation at 28-30oC/11 days

Racking

Pasteurization at 70-72oC/15 minutes

Bottling

Storage at room temperature

Analyses

Raisin

Addition of bentonite suspension (5%)

Clarification

50

0.5 g/L 1 g/L 1.5 g/L 0 g/L

3.2.5 Quality analysis

The quality of the prepared wines were analyzed by chemical analysis (TSS and alcohol

content) and sensory analysis (smell, taste, mouth feel, color and overall acceptance) by

ANOVA.

3.3 Selection of the best yeast

The optimized fermentation mash (20oBrix TSS, ginger 1% m/v and 4.5pH) was inoculated

with true wine and baker’s yeasts separately at the same rate 1g per liter.

3.4 Clarification of ginger wine using Bentonite

Both fermented ginger wines were mixed with bentonite suspension at the rate of 0.5, 1.0

and 1.5 g/L separately, stirred for 10 minutes in magnetic stirrer and left for overnight to

separate clear layers. Next day, the upper clear layer was racked into the next bottle gently.

Fig. 3.3 Clarification of ginger wine using bentonite.

3.5 Analytical methods

The moisture content of ginger was determined by distillation method as per AOAC

(2005). Reducing sugar, esters, aldehydes, acidities (total, fixed and volatile) and alcohol

content were determined as per AOAC (2005). TSS was measured by hand refractometer

and pH was measured by potable digital pH meter (Henna, Portugal). Turbidity of the

clarified wines was measured by turbidometer (Henna, Portugal).

51

3.6 Quality analysis of prepared wines

3.6.1 Sensory evaluation

Prepared ginger wines fermented by Baker’s and true wine yeasts were subjected to

sensory evaluation for consumer’s acceptability. Sensory attributes (such as smell, taste,

mouth feel, color and overall acceptance) were evaluated using a 9 point Hedonic scale

rating test ranging from dislike extremely (1) to like extremely (9) as described by

Ranganna (2001) by the help of semi-trained 15 panelists selected from those who were

familiar with alcoholic beverage. Samples were served in clean transparent labeled glasses.

Questionnaires and water for mouth rinsing between each tasting were provided. Sensory

evaluation sheet is given in Appendix III.

3.6.2 Statistical analysis

The determination was conducted in triplicates. The data were analyzed by using Genstat

programming (GenStat Discovery Edition 3, 2008) at 5% level of significance. The means

were compared using LSD method and t-test.

Part IV

Results and discussion

Raisin, sugar and ginger, available at local market of Dharan, were used for the preparation

of fermentation mashes. The ginger wines were prepared from the mashes having 10%

raisin, sugar concentrations (16o, 20o and 24 oBrix), ginger amounts (1, 1.5 and 2% m/v)

and pH of 4.5 by using baker’s yeast at room temperature. The pH was adjusted to 4.5 by

sodium bicarbonate or citric acid solutions. Similarly, TSS was adjusted to by using sugar

syrup. In the next experiment, the mashes having 10% raisin, 20oBrix TSS, 1% ginger and

4.5pH were pitched with true wine yeast and baker’s yeast separately. The fermentation

was allowed to proceed for 11 days at room temperature (28-30oC) until constant TSS was

obtained. After completion of the fermentation, the wines were analyzed for chemical

properties and subjected to sensory evaluation.

4.1 Effect of TSS and ginger amount on the chemical and sensory quality of ginger

wine

4.1.1 Effect on chemical characteristics

0

2

4

6

8

10

12

14

10 12 14 16 18 20 22 24 26

Initial TSS of the mash

Res

idua

l TSS

of g

inge

r w

ine

1% (m/v) ginger1.5% (m/v) ginger2% (m/v) ginger

Fig. 4.1 Effect of mash TSS on the TSS of the ginger wines.

53

0

2

4

6

8

10

12

14

0 0.5 1 1.5 2 2.5

Ginger amount, % (m/v)

Res

idua

l TSS

of t

he g

inge

r w

ine

16 Brix TSS20 Brix TSS24 Brix TSS

Fig. 4.2 Effect of ginger amount on the TSS of the ginger wines.

The final TSS of the ginger wines made from the mashes having initial TSS of 16o, 20o and

24 oBrix were found to be 6o, 7o and 11.7oBrix respectively. The ANOVA result showed

that the variation of initial TSS had a significant effect on the final TSS of the ginger

wines. The LSD test showed that the TSS of ginger wines made from the mashes having

initial TSS of 16o and 20oBrix were not significantly different (p<0.05). The TSS of the

ginger wine prepared from the mash having TSS of 24oBrix was significantly different

higher than those of the ginger wines prepared from the mashes of 16o and 20oBrix.

However, the variation of ginger amount had no significant effect on the final TSS of the

ginger wines (Appendix IV). Dhakal (2007) also reported that the same result in palm

wines.

o

o

54

4

5

6

7

8

9

10

11

10 12 14 16 18 20 22 24 26

Initial TSS of the mash

Alc

ohol

con

tent

of t

he g

inge

r w

ine,

%(v

/v) 1% (m/v) ginger

1.5% (m/v) ginger2% (m/v) ginger

Fig. 4.3 Effect of initial TSS on the alcohol content of the ginger wines.

4

5

6

7

8

9

10

11

0 0.5 1 1.5 2 2.5

Ginger amount, % (m/v)

Aco

hol c

onte

nt o

f the

gin

ger

win

e, %

(v/v

)

16 Brix TSS20 Brix TSS24 Brix TSS

Fig. 4.4 Effect of ginger amount on the alcohol content of the ginger wines.

The alcohol contents of the ginger wines made from the mashes having initial TSS of 16o,

20o and 24oBrix were found to be 6.87, 8.63 and 9.46 % (v/v) respectively. The ANOVA

result concluded that the variation of initial TSS had significant effect but ginger amount

had no significant effect on the alcohol content of the ginger wines (Appendix IV). The

LSD test showed that the ginger wines made from the mashes having TSS of 16o and

o

o

o

55

20oBrix and 16o and 24oBrix were significantly different but the ginger wines prepared

from the mashes of 20o and 24oBrix TSS were not significantly different in alcohol content.

Similarly, the alcohol content of the ginger wines made from the mashes having different

ginger amounts (1, 1.5 and 2 % m/v) were not significantly different.

4.1.2 Effect on sensory quality

0123456789

Smell Taste Color Mouth feel OverallSensory attributes

Mea

n sc

ore

16 Brix20 Brix24 Brix

4.5 Effect of initial TSS on the sensory properties of ginger wines.

012345678

Smell Taste Color Mouthfeel

Overall

Sensory attributes

Mea

n sc

ore

1% (m/v) ginger1.5% (m/v) ginger2% (m/v) ginger

4.6 Effect of ginger amount on the sensory properties of the ginger wines.

The average scores for smell of ginger wines made from the mashes having 16o, 20o and

24o Brix were found to be 6.41, 6.81 and 6.11 respectively. The ANOVA report indicated

o

o

o

56

that the variation of TSS of the mashes had significant effect (p<0.05) (Appendix V). The

smell of ginger wines made from the mashes of 16o and 20oBrix, 20o and 24oBrix, and 16o

and 24oBrix was significantly different each other.

The average scores for taste of ginger wines made from the mashes having 16o, 20o and

24o Brix were found to be 6.26, 6.41 and 5.78 respectively. The ginger wines made from

the mashes of 16o and 20oBrix were not different in taste. But the taste of the wines made

the mashes of 16o and 24oBrix and 20o and 24oBrix was significantly different.

The average scores for color of ginger wines made from the mashes having 16, 20 and 24

Brix were found to be 6.46, 7.18 and 5.96 respectively. The ginger wines prepared from the

mashes having TSS of 16o and 20oBrix were similar in color but the wines made from the

mashes having TSS of 16o and 24oBrix and 20o and 24oBrix were significantly differ in

color.

The average scores for mouth feel of ginger wines made from the mashes having 16o, 20o

and 24oBrix were found to be 5.87, 6.48 and 5.71 respectively. The ginger wines made

from the mashes containing TSS of 20o and 24oBrix and 16o and 24oBrix had same mouth

feel but the wines prepared from the mashes of 16o and 20oBrix had significantly different

mouth feel.

The average scores for overall acceptance of ginger wines made from the mashes having

16o, 20o and 24oBrix were found to be 6.27, 6.48 and 5.24 respectively. The overall

acceptance of the ginger wines prepared from the mashes with 16o and 20oBrix TSS was

same. But the wines made from the mashes with TSS of 16o and 24oBrix, TSS of 20o and

24oBrix were significantly different in overall acceptance.

The average scores for smell, taste, color, mouth feel and overall acceptance of ginger

wines made from the mashes having 1, 1.5 and 2g/L were found to be 6.56, 5.75, 6.13,

5.27and 6.27, 6.48, 6.54, 6.54, 6.48 and 6.38 and 6.54, 5.74, 6.14, 6.34 and 5.97

respectively.

The ANOVA report showed that the variation of ginger amount in the mashes had no

significant effect on sensory characteristics (smell, taste, color, mouth feel and overall

acceptance) of the ginger wines (p>0.05). However, the LSD test showed that there was

not significant different difference in smell of the ginger wines containing different ginger

amounts while the taste of the ginger wines having the ginger amounts 1 and 1.5% m/v was

significantly different. Similarly, the wines containing 1.5 and 2% m/v were also different

in taste. But the ginger wines having ginger amounts 1 and 2% m/v were not significantly

57

different in taste. There was not significant different difference in color of the ginger wines

containing different ginger amounts. The ginger wines made from the mashes having

ginger 1 and 1.5 % (m/v) and 1.5 and 2 % (m/v) were significantly different in mouth feel.

But the ginger wines made from the mashes containing 1 and 2% (m/v) were similar in

mouth feel. The LSD test indicated that the overall acceptance of the wines made from the

mashes containing 1 and 1.5% m/v ginger, 1 and 2% m/v ginger was not significantly

different but the wines made from the mash having 1.5 and 2% m/v ginger were

significantly different.

By the chemical and sensory analyses, the ginger wine made from the mash having

20oBrix TSS, 1% ginger (m/v) and 4.5pH was found to be the best among all prepared

ginger wines.

4.2 Effect of yeast culture on chemical and sensory properties

The ginger wines prepared from pre optimized composition of mash (20oBrix, 1% m/v

ginger and 4.5pH) by fermenting by true wine and baker’s yeasts were analyzed for

reducing sugar, acidities (total, fixed and volatile), esters, total aldehydes, alcohol content

and results are shown in Table 4.1. (Appendix VII)

Table 4.1 Chemical composition of the ginger wines.

Parameters Values*

Wine A Wine B

pH 4.1a (0.06) 4.2b (0.08)

TSS (oBrix) 5.80a (0.20) 6.20a (0.15)

Alcohol Content (%v/v) 7.80a (0.15) 8.71b (0.16)

Total Acidity (as g lactic acid/L alcohol) 7.56a (0.10) 7.92a (0.17)

Fixed Acidity (as g lactic acid/L alcohol) 5.4a (0.20) 5.88a (0.24)

Volatile Acidity (as g lactic acid/L alcohol) 2.16a (0.05) 2.04a (0.10)

Reducing sugar (g/L) 5a (0.2) 6b (0.21)

Esters (as mg ethyl acetate/L alcohol) 45.5a (3.23) 40.23a (4.36)

Total aldehydes (as acetaldehyde), mg/L alcohol 30.61a (4.0) 40.23a (4.36)

Note: * Values are the means of three determinations. Figures in the parenthesis are the

standard deviation. Means having same superscripts in a row are not significantly different

(p>0.05).

58

Wine A=fermented by true wine yeast.

Wine B=fermented by baker’s yeast.

4.2.1 pH

The average pH of the ginger wines fermented by true wine and baker’s yeasts were found

to be 4.1 and 4.2 respectively. t-test result indicated that the pH of ginger wines fermented

by true wine and baker’s yeasts were significantly different (p<0.05). Wine prepared by

true wine yeast had significantly higher pH than that of baker’s yeast. Dhakal (2007) found

that palm wine made from high pH had the high final pH while the TSS of mash had no

effect on the pH of the wines. He prepared palm wine having pH 3.23 from the mashes

having 20oBrix TSS and 4.5 pH. The prepared palm wine had low pH. The palm sap is

sensitive to spoilage microorganisms. It could be acidified before the initiation of desirable

fermentation by yeast culture.

4.2.2 TSS

The average TSS of ginger wines fermented by true wine and baker’s yeasts were found to

be 5.8 and 6.2 respectively. Statistical analysis indicated that the ginger wines were not

significantly different in TSS (p>0.05) (Appendix VII). Dhakal (2007) found that the

variation in TSS and pH had a significant effect on the final TSS of palm wine. The palm

wine made from mash having initial TSS of 20oBrix and pH of 4.5 had final TSS of 6.8 oBrix. According to Gautam (1992), apple wine prepared from the mash having TSS of

20oBrix and pH of 4.5 by using S. ellipsodeus and murcha had the final TSS of 5oBrix. He

reported that the wine with high initial TSS had high final TSS. These values of TSS were

similar to that of other wines. The TSS of the prepared ginger wines lies within the

reported range of TSS.

4.2.3 Alcohol content

The alcohol contents were found to be 7.8 and 8.71% (v/v) in the ginger wines fermented

by true wine and baker’s yeasts respectively. t-test indicated that ginger wines fermented

by true wine and baker’s yeasts were significantly different in alcohol content (Appendix

VIII). The alcohol in wines is formed as a result of utilization of carbohydrates present in

the mash for the growth and metabolic activities of yeast during fermentation (Patel, 1999).

In a suitable environment and substrate, the amount of alcohol produced depends upon the

59

amount of sugar present and the efficiency of the yeast in converting the sugar to alcohol.

The alcohol content varies from 5% up to 21% (v/v). Natural wines contain 8.5 to 16%

ethanol by volume (Boulton et al., 1997). The legal limits for alcohol vary markedly for

different countries and are usually related to the tax structure (Amerine et al., 1967).

Dhakal (1988) prepared banana and ginger wines containing 8.39% and 7.06% (v/v)

alcohol. Shakya (2002) reported 11% (v/v) alcohol in bael wine. Gubhaju (2006) prepared

Rhododendron wine having 11.03% (v/v) alcohol. According to Gvaladez (1936), the

alcohol yield varies from 47.87-48.12% of TSS of fermentation mash. The alcohol contents

of the ginger wines were within the range of other reported wines. The result showed that

baker’s yeast could produce more alcohol than that of true wine yeast. It could be due to

higher fermentation temperature (28 to 30oC) than that optimum fermentation temperature

required wine yeast (below 20oC). Baker’s yeast could tolerate higher temperature and

produce more alcohol than wine yeast.

4.2.4 Total acidity

The average total acidity of the ginger wines fermented by true wine and baker’s yeasts

were found to be 7.56 and 7.92 g/L (as lactic acid) respectively. t- test indicated that ginger

wines fermented by baker’s and true wine yeasts were not significantly different (p> 0.05)

in total acidity (Appendix VIII). It was found that baker’s yeast results higher total acidity

than that of ginger wine fermented by true wine yeast. Total acidities are important

constituents of wine not only for their acid taste but also because they protect the wine

from spoilage, maintain the color, and are themselves sometimes attacked by

microorganisms (Amerine et al., 1967). Dhakal (2007) prepared palm wines having

acidities 4.05 g/L as lactic acid from the mashes having 20oBrix TSS and 4.5 pH. Gubhaju

(2006) also prepared wines having similar results. According to Reed, (1987) the higher

amount of total acid of naturally fermenting wine may be due to the presence of significant

numbers of Kloeckera species. The calculated values of total acidities were higher than

other reported values. It could be caused by the growth of the spoilage micro-organisms in

the ginger wines. The growth of lactic acid bacteria could produce lactic acid and increased

total acidity.

4.2.5 Fixed acidity

The average fixed acidity of the ginger wines fermented by true wine and baker’s yeasts

were calculated to be 5.4 and 5.88 g/L (as lactic acid) respectively, but the values of the

60

ginger wines were not significantly different (Appendix VIII). Dhakal (2007) prepared

palm wine having acidities 2.96 g/L as lactic acid from the mashes having TSS of 20oBrix

and 4.5 pH. The calculated values of fixed acidities were higher than other reported values.

The growth of lactic acid bacteria could produce lactic acid and increased fixed acidity in

the ginger wines.

4.2.6 Volatile acidity

It was found that the amount of volatile acidities of the wines fermented by true wine and

baker’s yeasts were found to be 2.16 and 2.04 g/L (as lactic acid) respectively. Statistical

analysis result showed that the values were not significantly different (Appendix VIII). The

volatile acids are produced mainly during the initial stage of fermentation, and more is

formed in the presence of oxygen than in absence (Amerine et al., 1967). The

determination of volatile acidity as an indication of spoilage has become a part of the legal

requirements for wine standardization. The amounts of acetic acid produced during

alcoholic fermentation are small-usually less than 0.03 gm. per 100 ml. The volatile acidity

of a sound, newly fermented dry table wine may range from 0.2 to 0.4 g/L (Ribereau-

Gayon, 1961). Increase beyond this level, however, may signal microbial involvement and

potential spoilage. According to Egon et al., (1981) the volatile acidity of wines such as

Port, Sherry, Claret, Burgundy, Hock and Campagne were 0.05-0.10, 0.05-0.23, 0.09-0.15,

0.20-0.35, 0.05-0.15 and 0.03-0.20% (m/v) respectively. The calculated values of volatile

acidity were out of the range of all types of wines. It could be due to the activity of film

forming yeasts, and particularly of acetic acid bacteria and lactic acid in the ginger wines.

2.4.7 Reducing sugar

The average reducing sugar in ginger wines fermented by wine and baker’s yeast were

found to be 5 and 6 g/L as dextrose respectively. t-test indicated that ginger wines

fermented by baker’s and true wine yeasts were significantly different in reducing sugar

content in the ginger wines at p<0.05 (Appendix VIII). The amount of reducing sugar is

variable. Dry wine contains less than 0.12% sugar and sweet wine contains up to 7% sugar.

The amount of reducing sugar depends on the conditions for fermentation. The

consumption of sugar by yeast depends on the strain of the yeast. The results obtained

showed that wine yeast utilized more sugar than that by baker’s yeast.

61

4.2.8 Esters

The average values of esters in ginger wines fermented by true wine and baker’s yeasts

were 45.5 and 40.23 mg/L as acetaldehyde respectively. The values of esters were not

significantly different in ginger wines fermented by true wine and baker’s yeast (Appendix

VIII). Esters are an important part of odor of wines. Only ethyl acetate seems to be

important-below about 200 mg. per liter for a desirable odor but above this it appears to

give a spoiled character to the wine. Both neutral and acid esters are found in wine. The

total esters in various wines vary between about 200 and 400 mg. as ethyl acetate per liter

(Amerine et al., 1967). These values are lower than that of various wines. Esters are

formed mainly during ageing by the oxidation of higher alcohols. These lower values of

esters in the ginger wines could be due to lack of ageing as the ginger wines were not aged.

4.2.9 Total aldehydes

The average total aldehydes as acetaldehyde in ginger wines fermented by true wine and

baker’s yeasts were found to be 30.61 and 23.16 mg/L as acetaldehyde respectively. t-test

result indicated that the wines fermented by true wine and baker’s yeasts were not

significantly different (p>0.05) (Appendix VIII).Acetaldehyde is a normal by-product of

alcoholic fermentation. The amount of aldehydes ranges from 200 to 500 mg/L as

acetaldehyde in various wines (Amerine et al., 1967). The amounts found in newly-

fermented wines are below about 75 mg per liter. These values are within the range.

4.3 Sensory Evaluation

The sensory evaluation of ginger wines was done by semi trained panelists including

teachers, research students, and staff of CCT, Dharan. The parameters selected for sensory

evaluation were smell, taste, color, mouth feel and overall acceptance. The data was

analyzed by t-test at p=0.05 (Appendix IX).

62

0123456789

10

Smell Taste Mouth feel Color OverallacceptanceSensory attributes

Mea

n Sc

ores Fermented by wine yeast"

Fermented by baker's yeast"

Fig. 4.7 Effect of yeast type on the sensory properties of ginger wines fermented by true

wine and baker’s yeasts.

4.3.1 Smell

The average score for smell were found to be 8.1 and 6.9 for ginger wines fermented by

true wine and baker’s yeasts respectively. Statistical analysis showed that the smell of

ginger wine made by fermenting with true wine yeast was superior to that made by

fermenting with baker’s yeast (Appendix IX). Baker’s yeast could tolerate higher

temperature and consumed sugar faster. The fermenting yeast settled down at the bottom.

The ginger wine fermented by baker’s yeast came in contact with yeast cells and gained

yeasty flavor. So, ginger wine made by using true wine yeast was superior to that made by

using baker’s yeast at the provided conditions.

4.3.2 Taste

The average taste scores were found to be 7.2 and 7.5 respectively. The taste is combined

effect of oBrix and acidity. The t-test on the data on taste showed that there was no

significance difference between ginger wines fermented by true wine and baker’s yeasts.

These ginger wines did not differ each other at p<0.05 (Appendix IX).

4.3.3 Mouth feel

The average scores of ginger wines fermented by true wine and baker’s yeasts were found

to be 7.8 and 6.8 respectively. t-test of the data on mouth feel showed that there was not

significant difference between these ginger wines at p<0.05 (Appendix IX). The mouth feel

63

score of ginger wine fermented by true wine yeast was higher than that of ginger wine

fermented by baker’s yeast (Table 4.1). It could be due to higher sugar and alcohol content

in the ginger wine fermented by baker’s yeast.

4.3.4 Color

The average scores were found to be 7.8 and 6.8 for ginger wines fermented by true wine

and baker’s yeasts respectively. The t-test on the data on color showed that there was

significant difference between ginger wines fermented by true wine and baker’s yeasts at

p<0.05 (Appendix IX). The color score of ginger wine A higher than that of ginger wine B

(Table 4.1).

4.3.5 Overall acceptance

The average scores for overall acceptance were found to be 7.6 and 7.2 for ginger wines

fermented by true wine and baker’s yeasts respectively. The t-test showed that there was

significance difference between ginger wines at p<0.05 (Appendix IX). The overall score

of ginger wine fermented by true wine yeast is higher than that of ginger wine fermented

by baker’s yeast (Appendix VI).

4.4 Effect of bentonite on the clarification of ginger wine.

Bentonite was used for the clarification of the ginger wines at the rates of 0.5, 1 and 1.5

g/L. The average values of turbidities were found to be 16.52, 17.01, 17.81 and 526 FTU

for the ginger wines clarified by bentonite at the rate of 0, 0.5, 1.0 and 1.5 g/L respectively.

The ANOVA result showed that there was significant different on the clarity of ginger

wines by the use of bentonite in different concentrations (0.5, 1 and 1.5 g/L) at p<0.05. But

the ginger wines clarified by bentonite at the rate of 0.5, 1 and 1.5 g/L were not

significantly different each other. The turbidity of the ginger wine, which was clarified by

bentonite at the rate of 0.5g/L, was found to be the least (Fig. 4.8).

64

0

100

200

300

400

500

600

0 0.5 1 1.5

Bentonite concentration (g/L)

Turb

idity

(FTU

)

Fig. 4.8 Effect of bentonite on the clarification of ginger wine.

The result showed that the use of bentonite at the rate of 0.5 g/L was effective for the

clarification of the ginger wines. According to Boulton et al., 1997, the recommended

amount of bentonite for white table wine is 0.12 to 0.72 g/L (Table 2.4). In the experiment,

the effective amount of bentonite was found to be 0.5g/L for the prepared ginger wines.

The amount of bentonite for the clarification of wines depends on the pH. The amount of

bentonite for the clarification of the ginger wine was within the prescribed amount.

Part V

Conclusion and recommendation

On the basis of the results and discussion, the following conclusions were drawn:

5.1 Conclusions

1. The ginger wine could be prepared from the mash having 10% raisin, 20oBrix TSS, 1%

ginger (m/v) and 4.5pH by using baker’s yeast as comparable quality of the ginger

wine fermented by true wine yeast in the provided conditions.

2. Bentonite was found to be most effective at the rate of 0.5g/L for the clarification of

ginger wine.

5.2 Recommendations

1. The clarifying effects of different fining agents other than bentonite can be studied in

ginger wine.

2. Study on anti-microbial effects of ginger can be done.

3. Study on the physicochemical changes during its ageing can be carried out.

4. Study on the flavor profiles of the ginger wines can be carried out.

Part VI

Summary

The term “wine’’ used alone generally refers to the fermented product made from grape

juice and the wines made from fermented mashes of other fruits are commonly identified

by the specific fruit names such as pineapple wine, pumpkin wine etc. In the same way,

wine made by using spice to flavor and to add some nutritional values, the wine is also

named by the name of the spice used such as ginger wine.

In this study, the ginger was bought from local market of Dharan which was cultivated in

Bhojpur district, baker’s yeast was bought from Gorkha Department Store, Itahari-1, which

was manufactured in Belgium and true wine yeast was collected from Makalu Wine

Industries (P.) Ltd., Basantapur, Tehrathum. First, mashes having different sugar

concentrations (16o, 20o and 24oBrix) and ginger amounts (1, 1.5 and 2% m/v) were

fermented by baker’s yeast at the rate 1 g/L at room temperature. After completion of

fermentation, final TSS and alcohol content were determined. The ginger wines thus

prepared were analyzed for both chemical and sensory properties. The data obtained was

analyzed and interpreted statistically by ANOVA at p<0.05. The initial TSS of the mash

had a significant effect but ginger amount had no significant effect on the chemical and

sensory properties. The ginger wine made from the mash having TSS of 20oBrix, 4.5pH

and ginger amount 1% m/v was found to be superior to other wines.

The mashes having TSS of 20oBrix, 4.5pH and ginger amount 1% m/v were made,

inoculated with true wine and baker’s yeasts at the rate of 1gm. per liter separately and left

for fermentation. After completion of fermentation, they were clarified by using bentonite

suspension (5% m/v) at the rates of 0.5, 1 and 1.5 g/L. The use of bentonite had a

significant effect on the clarification of the ginger wines. The bentonite at the rate of 0.5

g/L was found to the most effective for the clarification of the ginger wines. The ginger

wines were analyzed for chemical properties (pH, TSS, alcohol content, total acidity, fixed

acidity, volatile acidity, reducing sugar, esters and total aldehydes). The pH, TSS, and

alcohol content of the ginger wines A and B were 4.1 pH, 5.8oBrix, 7.8% (v/v) and 4.2 pH,

6.2oBrix, 8.71 respectively. The total, fixed and volatile acidities (as g lactic acid) of the

ginger wines fermented by true wine and baker’s yeasts were 7.56, 5.4, 2.16 and 7.92 g,

67

5.88, 2.04 g/L respectively. The reducing sugar of the ginger wines were 5 and 6 g/L

respectively.

Similarly, the esters and total aldehydes of the ginger wines fermented by true wine and

baker’s yeasts were 45.5, 30.61 and 40.23, 23.16 mg/L alcohol respectively.

There was significant difference in the pH, alcohol content and reducing sugar between

them by t-test at p<0.05. TSS, total acidity, fixed acidity, volatile acidity, esters and total

aldehydes were not significantly different by t-test.

The prepared ginger wines were subjected for sensory evaluation (smell, taste, mouth

feel and overall acceptance). These ginger wines were not significantly different in taste

and mouth feel but they were significantly different in smell, color and overall acceptance

by t-test at p<0.05.

The ginger wine could be prepared from the mash having 10% raisin, 20oBrix TSS, 1%

ginger (m/v) and 4.5pH by using baker’s yeast as comparable quality of the ginger wine

fermented by true wine yeast in the provided conditions. Bentonite was found to be most

effective at the rate of 0.5g/L for the clarification of ginger wine.

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Appendices

Appendix I

Table A.1 TSS reduction during fermentation of mashes.

Days from pitchingTSS

Mash A Mash B

5 days 8 9.5

6 days 7.5 8.5

7 days 7 7.5

8 days 6.5 7

9 days 6 6.5

10 days 5.8 6.2

11 days 5.8 6.2

Appendix II

Table A.2 Data obtained from analysis of samples of wines.

Parameters Values*

Wine A Wine B

pH 4.1a (0.06) 4.2b (0.076)

TSS (oBrix) 5.80a (0.2) 6.20a (0.153)

Alcohol Content (%v/v) 7.80a (0.150) 8.71b (0.160)

Total Acidity (as gm lactic acid/L alcohol) 7.56a (0.104) 7.92a (0.166)

Fixed Acidity (as gm lactic acid/L alcohol) 5.4a (0.200) 5.88a (0.239)

Volatile Acidity (as gm lactic acid/L alcohol) 2.16a (0.051) 2.04a (0.104)

Reducing sugar (g/L) 5a (0.2) 6b (0.208)

Esters (as mg ethyl acetate/L alcohol) 45.5a (3.229) 40.23a (4.364)

Total aldehydes (as acetaldehyde), mg/L alcohol 30.61a (3.992) 23.16a (1.967)

Note: * Values are the means of three determinations.

Wine A=fermented by true wine yeast

Wine B=fermented by baker’s yeast

75

Appendix III

Table A.3 Specimen cards for sensory evaluation by hedonic rating

Hedonic Rating Test

Name:

Date:

Product: Ginger Wine

Please taste the samples (wines fermented by baker’s yeast and wine yeast separately) and

taste how much you like or dislike. Use the appropriate scale to show your attitude by

giving the point that best describe your feeling about the sample.

Give points as follows:

Like extremely 9

Like very much 8

Like moderately 7

Like slightly 6

Neither Like nor dislike 5

Dislike slightly 4

Dislike moderately 3

Dislike very much 2

Dislike extremely 1

Parameters Scores of samples

A B

Smell

Taste

Color

Mouth feel

Overall acceptance

Comment if any:

76

Appendix IV

Table A.4 ANOVA table for the chemical properties (TSS and alcohol content) of the

ginger wines fermented by baker’s yeast.

Variate: TSS

Source of variation d.f. s.s m.s. v.r. F pr.

Initial TSS 2 127.527 63.7639 255 <0.001

Ginger amount, % m/v 2 0.3611 0.1866 0.72 0.512

Ginger amount, % m/v× Initial TSS 4 6.4722 1.6181 6.47 0.010

Residual 9 2.2500 0.2500

Total 27 136.6111

Variate: Alcohol content

Source of variation d.f. s.s m.s. v.r. F pr.

Initial TSS 2 21.623 10.81159 128.11 <0.001

Ginger amount, % m/v 2 0.01034 0.00517 0.06 0.941

Ginger amount, % m/v× Initial TSS 4 1.04858 0.26215 3.11 0.073

Residual 9 0.75954 0.08439

Total 17 23.44164

77

Appendix V

Table A.5 ANOVA table for sensory characteristics of ginger wines fermented by baker’s

yeast.

Variate: Smell

Source of variation d.f. s.s m.s. v.r. F pr.

Ginger amount, % m/v 2 0.296 0.148 0.14 0.871

Initial TSS 2 6.741 3.370 3.14 0.049

Ginger amount, % m/v× Initial TSS 4 5.630 3.14 1.31 0.274

Residual 72 77.333 1.074

Total 80 90.000

Variate: Taste

Source of variation d.f. s.s m.s. v.r. F pr.

Ginger amount, % m/v 2 4.519 2.259 1.30 0.278

Initial TSS 2 5.852 2.926 1.69 0.192

Ginger amount, % m/v× Initial TSS 4 14.963 3.741 2.16 0.083

Residual 72 124.889 1.735

Total 80 150.222

Variate: Mouth feel

Source of variation d.f. s.s m.s. v.r. F pr.

Ginger amount, % m/v 2 8.617 4.369 3.04 0.054

Initial TSS 2 1.506 0.753 0.53 0.590

Ginger amount, % m/v× Initial TSS 4 5.679 1.420 1.00 0.412

Residual 72 102.000 1.417

Total 80 117.802

78

Variate: Color

Source of variation d.f. s.s m.s. v.r. F pr.

Ginger amount, % m/v 2 9.5840 5.2920 3.46 0.066

Initial TSS 2 1.54420 0.7210 0.65 0.67

Ginger amount, % m/v× Initial TSS 4 5.25640 1.5410 1.34 0.617

Residual 72 102.230 1.523

Total 80 2.85900

Variate: Overall acceptance

Source of variation d.f. s.s m.s. v.r. F pr.

Ginger amount, % m/v 2 2.691 1.346 1.01 0.368

Initial TSS 2 7.802 3.901 2.94 0.059

Ginger amount, % m/v× Initial TSS 4 2.864 0.716 0.54 0.707

Residual 72 95.556 1.327

Total 80 108.914

Appendix VI

Table A.6 Sensory evaluation scores of the wine samples.

Parameters

Panelists

1 2 3 4 5 6 7 8 9 10

A B A B A B A B A B A B A B A B A B A B

Smell 9 8 8 6 8 7 8 7 9 8 8 6 8 7 7 6 8 7 8 7

Taste 8 8 7 8 7 8 7 8 8 8 7 6 7 8 7 8 7 8 7 6

Mouth Feel 8 7 7 8 7 6 8 8 8 7 7 7 7 6 7 6 8 8 8 7

Color 8 8 8 6 8 7 8 7 8 7 7 6 8 7 7 6 8 7 7 8

Overall acceptance 8 7 7 8 8 7 8 8 8 8 7 6 8 7 7 6 8 7 8 7

Note: In Table A= Wine fermented by baker’s yeast

B= Wine fermented by wine Yeast

79

Appendix VII

Table A.7 Chemical composition of the ginger wines fermented by true wine and baker’s

yeasts.

Parameters Values*

Wine A Wine B

pH 4.1a (0.06) 4.2b (0.08)

TSS (oBrix) 5.80a (0.20) 6.20a (0.15)

Alcohol Content (%v/v) 7.80a (0.15) 8.71b (0.16)

Total Acidity (as g lactic acid/L alcohol) 7.56a (0.10) 7.92a (0.17)

Fixed Acidity (as g lactic acid/L alcohol) 5.4a (0.20) 5.88a (0.24)

Volatile Acidity (as g lactic acid/L alcohol) 2.16a (0.05) 2.04a (0.10)

Reducing sugar (g/L) 5a (0.2) 6b (0.21)

Esters (as mg ethyl acetate/L alcohol) 45.5a (3.23) 40.23a (4.36)

Total aldehydes (as acetaldehyde), mg/L alcohol 30.61a (4.0) 40.23a (4.36)

Appendix VIII

Table A.8 t-test table for chemical properties of the ginger wines.

Parameters Mean Variance

df tcal. ttab. A B A B

pH 4.1333 4.3166 0.0033 0.0058 4 3.316 2.7764

TSS 5.8 6.1667 0.04 0.0233 4 2.524 2.7764

Alcohol 7.7933 8.7067 0.0226 0.0256 4 7.201 2.7764

Total acidity 6.2667 6.5967 0.0108 0.0277 3 2.910 3.1824

Fixed acidity 4.54331 4.8867 0.0401 0.0443 4 2.045 2.7764

Volatile acidity 1.80667 1.7333 0.0026 0.0108 3 1.094 3.1824

Reducing sugar 5.1 6.0333 0.04 0.0433 4 5.6 2.7764

Esters 45.5367 40.23 10.428 19.047 4 1.693 2.7764

Total aldehydes 30.7667 23.163 15.936 3.8682 3 2.959 3.1824

80

Appendix IX

Table A.9 t-test table for sensory properties of the ginger wines.

Parameters Mean Variance

tcal. ttab. A B A B

Smell 8.1 6.9 0.3222 0.5444 17 4.076 2.110

Taste 7.2 7.6 0.1778 0.4889 15 1.549 2.131

Mouth feel 7.5 7 0.2778 0.6667 15 1.627 2.131

Color 7.7 6.9 0.2333 0.5444 16 2.868 2.112

Overall acceptance 7.7 7.1 0.2333 0.5444 16 2.151 2.112

Appendix X

Table A.10 ANOVA table for turbidities of ginger wines.

Source of variation d.f. s.s m.s. v.r. F pr.

Replicate 2 0.05840 0.02920 0.46 0.663

Amount of bentonite 2 2.54420 1.27210 19.85 0.008

Residual 4 0.25640 0.06410

Total 8 2.85900

81

Appendix XI

Table A.11 Effect of bentonite on the clarification of ginger wine.

Bentonite concentration Turbidity (FTU)

0.0 g/L 526

0.5 g/L 16.52

1.0 g/L 17.01

1.5 g/L 17.81

Appendix XII

Table A.12 Average chemical analysis of prize-winning high quality wines.

Component

(g per 100 ml)

Dry

White

Dry Red Sweet White Sweet Red Sparkling

Alcohol by volume, (%) 2.45 12.61 18.38 19.30 13.22

Alcohol 9.88 10 14.58 10.48

Glycerol 0.7019 0.6355 0.3025 0.5089 0.4177

Ash 0.196 0.247 0.203 0.311 0.153

Total acids 0.586 0.649 0.412 0.502 0.658

Volatile acids 0.101 0.128 0.092 0.122 0.082

Reducing sugars 0.134 0.146 11.30 10.20 3.409

Protein 0.162 0.150 0.162 0.232 0.214

Tannins 0.039 0.236 0.036 0.096 0.035

Specific gravity 0.9917 0.9947 1.0298 1.0276 1.0045

Source: Amerine et al., 1972

82

Appendix XIII

Table A.13 Major wine producing countries of the world-1996.

Countries

Wine

production

(million L)

Wine

exports

(million L)

Wine

consumption

(million L)

Total

grape’000

Ton

Area of

vines’000

Hectare

France 5965 1229 3479 7701 917

Italy 5877 1511.5 3562 9459 922

Spain 3267 672.9 1475 4846 1224

USA 1864 163.8 2046 4935 311

Argentina 1268 125.4 1355 2040 211

S. Africa 1000 99.6 406 1440 106

Portugal 953 200 580 1270 259

Germany 830 300.8 1866 1297 105

Romania 766 46.7 725 1427 256

Australia 678 147.1 329 1086 81

Others 4785 1241.6 6499 23180 3350

World

total

27253 5738.4 22322 58681 7742

(Source: Anon, 1996)

83

Appendix XIV

Table A.14 Composition of some wines.

Parameters Port Sherry Claret Burgundy Champagne

Specific gravity 0.995-

1.050

0.992-

1.015

0.990-

1.001

0.995-

1.001

1.040-1.055

Alcohol (gm/100ml) 13.5-20.0 13.5-20.5 7.5-12.5 7.5-12.5 10.0-14.0

% Total solid 3.3-13.0 2.0-9.6 2.0-3.5 2.0-3.5 9.5-18.0

% Free volatile acid (as

acetic acid)

0.05-0.10 0.15-0.23 0.09-0.15 0.2-0.35 0.03-0.20

% Fixed acid (as acetic

acid)

0.35-0.55 0.25-0.50 0.30-0.50 0.3-0.60 0.30-0.45

% Ash 0.25-0.35 0.35-0.55 0.2-0.3 0.2-0.4 0.25-0.45

% Sugars 2.5-12.0 2.0-7.0 0.0-0.7 0.03-0.55 8.5-16.0

(Source: Pearson, 1981)