SUKRASNO SITI KUSMARDIYANI

SCOPE OF MEASUREMENT

Biological Physical Chemical Physicochemical

BIOLOGICAL PARAMETERS

TO BE MEASURED

Crude Drug

Chemical Constituents

Biological Activity

Quality

Assay

Biological Parameters

Toxic

Cytotoxic

Increase membrane

permeability

Brine shrimp lethality test

Fish index

Hemolytic test

Biological Activity

EXTRACTS / FRACTIONS / ISOLATES

2 days

1 day

LD50

10 nauplii( á ~0.5 mm)

Evaporate + ASW 5 mL

Artificial sea water (ASW) 38 gram sea salt/L

Brine shrimp eggs

Count the number of dead nauplii, use Program Finney

BRINE SHRIMP LETHALITY TEST

A brine shrimp hatchery made from a plastic soap dish and placed in the light. The shrimp should hatch within 24-48h. Significant if LD50 <30 ug/mL. KUSMARDIYANI

BIOASSAY- Explain -

Adults have three eyes and 11 pairs of legs and can grow to about 15 millimetres in size.

Indeks ikan adalah bilangan atau angka yang menunjukkan pada pengenceranberapa larutan suatu zat membunuh 3 dari 5 ekor ikan yang panjangnya 2 – 4 cmdalam waktu satu jam.

Species:

Leuciscus rutilusCarracius vulgarisTillapia mossambica (mujair) Ikan seribu

Tanda-tanda keracunan saponin, diamati setelah disimpan selama 24 jam dalam lemari pendingin:

Insang menjadi merah, pembuluh darah insang membengkak. (Mati karena sebab lain tidak menyebabkan insang merah)

Apandi, A., 1959, Penentuan kekuatan beberapa bahan tumbuhan dengan cara biologi, Suara Farmasi, 23 – 28.

Translate and discuss.

© SK

Red blood cells without (left and middle) and with (right) hemolysis. Note that the hemolyzed sample is transparent,

because there are no cells to scatter light.

HEMOLYSIS

-Wikipedia-

PHYSICAL PARAMETERS

TO BE MEASURED

PHYSICAL PROPERTIES

Crude drug

Extract

Organoleptic

Microscopic

Chemical Component

Powder

MorphologyColourOdourTaste Sound

Physical property of the major component

Density Refractive index Boiling point Optical Rotation Solubility/miscibility

Physical properties related to liquid state

Swelling capacity swelling index Decrease surface tension foaming index Pungency Scoville index Bitter bitterness index Sweet sweetness index Solubility solvent extractive Ignitability ash residue

Physical properties of the major component

CHEMICAL PARAMETERS

TO BE MEASURED

Crude Drug

Chemical component

ReactivityChemical group

CHEMICAL PROPERTIES

Chemical Groups

Alkaloid Flavonoid Tannin Quinone Steroid/Triterpenoid ……. …….

Nucleic acid – Plant identification

• RFLP (Restriction fragment length polymorphism)– DNA– Restrictive endonuclease– Gel Electrophoresis– Finger printing (DNA fragment profile)

• RAPD (Random amplified polymorphism of DNA)– DNA– PCR– Gel electrophoresis– Much more simple DNA fragment profile

PHYSICOCHEMICAL PARAMETERS

TO BE MEASURED

• Spectrophotometry/Spectrometry

• Chromatography

PHYSICOCHEMICAL PROPERTIES

• Spectrophotometry–UV-Vis– IR

• Spectrometry– Mass Spectrometry– NMR• Proton NMR• 13C-NMR

PHYSICOCHEMICAL PROPERTIES

--- get more recent information ---

UV-Vis Spectrophotometry

– Qualitative measurement– Quantitative measurement

• Chlorophyl• Carotenoid• Piperin• Curcuminoid

--- get more recent information ---

IR Spectrophotometry

• Not much use in analytical pharmacognosy• Suitable for volatile oils• IR profile can be used as one of identity

parameter.

--- get more recent information ---

Mass Spectrometry• Not much use in analytical pharmacognosy.

• Its combination with gas chromatography: very useful tool in volatile oil analysis.

• In addition to finger printing, mass spectrum of each component can be obtained and identified base on data base present in system.

--- get more recent information ---

NMR Spectrometry

• Not much use in analytical pharmacognosy.

• Very important instrument for structure elucidation of pure compound.

--- get more recent information ---

CHROMATOGRAPHY

• Paper chromatography• Thin layer chromatography• Column chromatography• HPLC• GC

--- get more recent information ---

Paper Chromatography

• Suitable for polar compound, such as: carbohydrates, phenolics, flavonoids.

• Limited variation of solvent can be used.

--- get more recent information ---

Thin Layer Chromatography• Very widely used in analytical pharmacognosy.• Finger printing of crude drug.• Determination of marker.• Chromatographic profiling.• Can be eluted vertically or circularly.• Various composition of solvent

can be used.

--- get more recent information ---

THIN LAYER CHROMATOGRAPHY OF CINCHONA BARKS

Developing system: chloroform - diethylamine (9:1) Spray reagent: (a) 10% H2SO4 in ethanol UV365nm

(b) 10% H2SO4, then iodoplatinate reagent visible light

1 = Cinchona calisaya extract 2 = Cinchona succirubra extract TC = a mixture of cinchona alkaloids

T1 = quinine T2 = sinchonidine T3 = quinidine T4 = sinchonine

(a) (b) Wagner, TLC Atlas..

--- Explain ---

RUTAE HERBA p144

BELADONNAE, HYOSCYAMI, STRAMONII FOLIUM, p49

ATLAS OF PLANT DRUG ANALYSIS

Wagner, TLC Atlas..

Column Chromatography• Suitable for sample preparation.• Mostly use to fractionate chemical

constituents.• Example:

– Analysis of pesticide residue.– Sample clearance

prior to HPLC analysis.

--- get more recent information ---

HPLC• High performance

liquid chromatography.• Very widely used in analytical

pharmacognosy.• Can be used for qualitative or

quantitative analysis.

• Detection system varies.• Coupled with Diode Array Detector

provide more information on identity of the peak.

--- get more recent information ---

HPLC hyphenated

• Combined with MS (LC-MS) provide better performance of the device.

• More versatile.

• Higher sensitivity for quantitative analysis.

• Better information on identity of each peak by the availability of the mass spectrum of each peak.

--- get more recent information ---

Gas Chromatography GC is very suitable for analysis of

volatile oil.

It is also used for the analysis of pesticide residue.

Example:Analysis of Peppermint Oil

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SCOPE OF MEASUREMENT

Biological Physical Chemical Physicochemical

Find one example of the monographs, read carefully, and discuss the scope of measurement.

Farmakope Indonesia Edisi IVp225. -- an example --

Farmakope Indonesia Edisi IV

p226.

Find: specific and non–specific parameters for this example of crude drug.

Discuss.

Find: specific and non–specific parameters for this example of crude drug. Discuss.

Find: specific and non–specific parameters for this example of crude drug. Discuss.

Find: specific and non–specific parameters for this example of crude drug. Discuss.

I. Pharmacopoeial summaries for quality assurance purposes.. Botanical features. Identity tests. Purity requirements. Chemical assays . Major chemical constituents

II. Review of scientific research. Clinical applications of plant material. Detailed pharmacological information :

. contraindications

. warnings

. precautions

. adverse reactions

. dosage

Bulbus Allii CepaeBulbus Allii SativiAloeAloe Vera GelRadix AstragaliFructus BruceaeRadix BupleuriHerba CentellaeFlos ChamomillaeCortex CinnamomiRhizoma CoptidisRhizoma Curcumae LongaeRadix EchinaceaeHerba Echinaceae PurpureaeHerba Ephedrae

Folium GinkgoRadix GinsengRadix GlycyrrhizaeRadix PaeoniaeSemen PlantaginisRadix PlatycodiRadix RauwolfiaeRhizoma RheiFolium SennaeFructus SennaeHerba ThymiRadix ValerianaeRhizoma Zingiberis

--------------------- VOLUME 1

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Fructus Ammi MajorisFructus Ammi VisnagaeFructus AnethiAetheroleum AnisiFructus AnisiSemen ArmenicaeFlos ArnicaeFolium AzadirachtiOleum AzadirachtiFlos CarthamiStigma CrociFructus FoeniculiRadix Gentianae LuteaeRadix Gentianae ScabraeGummi GugguliRadix Harpagophyti

--------------------- VOLUME 3

Rhizoma HydrastisRadix IpecacuanhaeAetheroleum LavandulaeFlos LavandulaeStrobilus LupuliGummi MyrrhaHerba Passifl oraeTesta PlantiginisRadix RehmanniaeFructus SchisandraeRadix ScutellariaeRadix cum Herba TaraxaciSemen Trigonellae FoenugraeciCortex UncariaeFructus Zizyphi

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