Antioxidative and Antibacterial coating for fish steaks
1
Anti-oxidative and Antibacterial coating for fish steaks using composite pullulan and polyphenols of pomegranate Mozhiarasu S, Satish Kushwaha, Dr. Manjeet Aggarwaal National Institute of Food Technology Entrepreneuship and Management Abstract In order to reduce the spoilage in foods, various technologies can be used hand in hand, one such technology is the edible coating, carbohydrate based coating on food can help to reduce the oxygen diffusion and if the coating contains antimicrobials, it also helps to protect against microbial growth. Pomegranate peel having large set of polyphenols which are reported to act both anti-oxidative and antimicrobial agents. In this work the pomegranate polyphenols were extracted with water, purified using Amberlite XAD 16 column obtained the yield of 70.36±12.56 mg gallic acid eq /g of peel and 52.82±11.83 mg gallic acid eq /g of peel during extraction and purification re- spectively. The anti-oxidative and antimicrobial properties were analysed. The DPPH IC 50 value of polyphenol was found to be 16.15 µg/mL and MIC of 5 test bacterium was determined. The effect of different formulation of polyphenols and pullulan coat- ing solution on microbial growth and TBARS values were evaluated. Thus 9.5% pul- lulan, 0.5% Polyphenol with 0.5% carboxy methyl cellulose improved coating stability and the increased the shelflife of the fish steak from 3 days to 8 days at 4 o C in terms of microbial quality. Coating with pullulan-Polyphenols has also reduced the TBARS value approximately 4 fold and improved the fish steaks from oxidative stress. Introduction The safety and quality of the food depends on the nutrition, hygiene, and organoleptic characteristic of the food [3]. But the characteristic may deteriorate during storage and commercialization. The shelf-life of the fish depends on two factors, the first one is the microbiological load, in which fishes are highly prone to microbial spoilage because of its nutritious quality, water activity (0.95) and neutral pH 6.5-7 [6], apart from that, autooxidation of lipids is another factor which converts the nutritious fatty acids into harmful toxic components like malondialdehyde, so in order to prevent from spoilage, various technologies can be used hand in hand, one such is edible films and coatings, coating material will consist of biopolymers like carbohydrates, proteins, or lipids and resins and composite material made of those. Carbohydrate coating can prevent the diffusion of the oxygen and prevent oxidation, thus an antibacterial and antioxidative components along with polysaccharide coating helps to achieve the goal of enhancing shelf-life. Pomegranate are rich in phytochemicals and its peel found to have compo- nents which are antibacterial and anti-oxidative potential. Using of pomegranate peel utilizes the fruit waste and added value to both food (fish steak) and waste (peel). Materials & Methods ● Extraction of Polyphenols from pomegranate peel: Pomegranates (Bhaguva variety from local market at Narela) were washed with potable water and peels were removed. The peels blended with water in ratio 1: 10 (peel:water) (w/v) transfered to conical flask and it was kept at 50 o C for 15 minutes for maximum extraction with occasional stirring. Then it was filtered through muslin cloth and centrifuged at 3000g for 15 minutes to remove insoluble. The extract was stored in brown bottle. ● Purification of polyphenols: The pomegranate polyphenols were purified by using Amberlite XAD 16 resin [7] ● Total Phenolic Content: It was measured using Folin Ciocalteu method ● Characterization of polyphenols in HPLC-UV: HPLC System: Waters Inc USA Model 2707; Column: C18 Spherisorb ODS 250 column 4.6mm (5µ); Column Temperature : 25oC; Mobile phase A- 1.3g potassium di hydrogen phosphate solution in water ; Mobile phase B – Acetonitrile 100%; Standards used: Gallic acid, Ellagic Acid, Punicalgin. ● Antioxidative Assay: DPPH scavenging activity was performed as described in Sweetie Kannat, 2010 ● Antibacterial activity: Antibacterial well diffusion assay and Minimum inhibitory concentration(1-10mg/mL) was performed as described in Al-Zoreky, 2009. ● Preparation and Coating of Fish: The fishes were procured in local market and brought to lab with ice pack, it was wased with potable water, beheaded, degut- ted and made into steaks. The fishes were placed in coating formulation in the ratio (1:2) in a container and kept inside the vacuum chamber for the vacuum im- pregnation at 100mmHg at room temperature for 15 minutes (Duan, Cherian, & Zhao, 2010). The coated fish samples were removed and air dried for 30 min- utes and kept inside the LDPE zipped pouches for the storage at 4 o C in refriger- ator. The samples were taken on 3, 6, 8, 10th day and all the parameters were analyzed until all the sample reached the maximum permissible microbial count 10 7 CFU/g or 7 log units. Results and Discussions Approximately 2 fold increase of antioxidative activity of the polyphenols after purifi- cation. Purified Powder of Pomegranate Polyphenol Presence of ellagic acid and punicalgin. ● Anti Bacterial activity represent the polyphenols are more active against gram positive bacteria than gram negative bacteria except Pseudomonas aeruginosa. While all the tested bacteria can be inhibited at 2mg/mL concentration but Sa- monella typhimurium was quite resistant, inhibited only at 4 mg/mL. ● Scavenging property for free radicals of pomegranate polyphenols is greater than BHT IC 50 value: 16.15 µg/mL (BHT- 21.2 µg/mL L (Kannat et al., 2010)) COATING EXPERIMENT RESULTS Effect on Microbial Load: Polyphenols reduced the bacteral count and enhanced the shelf life, while pullu- lan-polyphenol coating enhanced shelflife from 3 to 6 days and addition of CMC in- to pullulan and polyphenols increased the retention and stabilized the coating from washout during drip loss. Basic-Total Volatile Nitrogen has increased more in control samples while reduction upto 2.5 fold in sample containing Polyphenol. TBARS determination results shown, decreasing in TBARS value in coated sample even without polyphenols shows pres- ence of pullulan reduced oxygen diffusion. There was 4 fold reduction in the CMC- Pullulan-Polyphenol coated sample. An edible coating has been developed using pullulan and cmc (polysaccharide) and polyphenol from pomegranate peel. ● The coating on to fish steaks increased the shelf life from 3 days to 8 days at 4 o C. ● The present project involving the use of polyphenols from pomegranate peel helps in waste utilization from peel ● Besides increasing the shelf-life of the product, the coating helps in providing the functionality to the product. ● The cost economics of the product can be further brought down by blending a polymer from other sources (possibly from fruit waste). References 1. N. S. Al-Zoreky, “Antimicrobial activity of pomegranate (Punica granatum L.) fruit peels.,” Int. J. Food Microbiol., vol. 134, no. 3, pp. 244–8, Sep. 2009. 2. S. Kanatt, R. Chander, and A. Sharma, “Antioxidant and antimicrobial activity of pomegranate peel extract improves the shelf life of chicken products,” Int. J. food …, 2010. 3. F. Debeaufort, J. a Quezada-Gallo, and A. Voilley, “Edible films and coatings: tomorrow’s packagings: a review.,” Crit. Rev. Food Sci. Nutr., vol. 38, no. 4, pp. 299–313, May 1998. 4. O. Lie, Improving Farmed Fish Quality and Safety. Elsevier, 2008. 5. J. Duan, G. Cherian, and Y. Zhao, “Quality enhancement in fresh and frozen ling- cod (Ophiodon elongates) fillets by employment of fish oil incorporated chitosan coatings,” Food Chem., vol. 119, no. 2, pp. 524–532, Mar. 2010. 6. M. R. Adams and M. O. Moss, Food Microbiology. Royal Society of Chemistry, 2008. 7. N. Seeram, R. Lee, M. Hardy, and D. Heber, “Rapid large scale purification of el- lagitannins from pomegranate husk, a by-product of the commercial juice indus- try,” Sep. Purif. …, 2005.
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Transcript of Antioxidative and Antibacterial coating for fish steaks
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Anti-oxidative and Antibacterial coating for fish steaks using composite pullulan and polyphenols ofpomegranate
Mozhiarasu S, Satish Kushwaha, Dr. Manjeet AggarwaalNational Institute of Food Technology Entrepreneuship and Management
AbstractIn order to reduce the spoilage in foods, various technologies can be used hand inhand, one such technology is the edible coating, carbohydrate based coating on foodcan help to reduce the oxygen diffusion and if the coating contains antimicrobials, italso helps to protect against microbial growth. Pomegranate peel having large set ofpolyphenols which are reported to act both anti-oxidative and antimicrobial agents.In this work the pomegranate polyphenols were extracted with water, purified usingAmberlite XAD 16 column obtained the yield of 70.3612.56 mg gallic acid eq /g ofpeel and 52.8211.83 mg gallic acid eq /g of peel during extraction and purification re-spectively. The anti-oxidative and antimicrobial properties were analysed. The DPPHIC 50 value of polyphenol was found to be 16.15 g/mL and MIC of 5 test bacteriumwas determined. The effect of different formulation of polyphenols and pullulan coat-ing solution on microbial growth and TBARS values were evaluated. Thus 9.5% pul-lulan, 0.5% Polyphenol with 0.5% carboxy methyl cellulose improved coating stabilityand the increased the shelflife of the fish steak from 3 days to 8 days at 4oC in termsof microbial quality. Coating with pullulan-Polyphenols has also reduced the TBARSvalue approximately 4 fold and improved the fish steaks from oxidative stress.
IntroductionThe safety and quality of the food depends on the nutrition, hygiene, and organolepticcharacteristic of the food [3]. But the characteristic may deteriorate during storage andcommercialization. The shelf-life of the fish depends on two factors, the first one is themicrobiological load, in which fishes are highly prone to microbial spoilage becauseof its nutritious quality, water activity (0.95) and neutral pH 6.5-7 [6], apart from that,autooxidation of lipids is another factor which converts the nutritious fatty acids intoharmful toxic components like malondialdehyde, so in order to prevent from spoilage,various technologies can be used hand in hand, one such is edible films and coatings,coating material will consist of biopolymers like carbohydrates, proteins, or lipids andresins and composite material made of those. Carbohydrate coating can prevent thediffusion of the oxygen and prevent oxidation, thus an antibacterial and antioxidativecomponents along with polysaccharide coating helps to achieve the goal of enhancingshelf-life. Pomegranate are rich in phytochemicals and its peel found to have compo-nents which are antibacterial and anti-oxidative potential. Using of pomegranate peelutilizes the fruit waste and added value to both food (fish steak) and waste (peel).
Materials & Methods Extraction of Polyphenols from pomegranate peel: Pomegranates (Bhaguva
variety from local market at Narela) were washed with potable water and peelswere removed. The peels blended with water in ratio 1: 10 (peel:water) (w/v)transfered to conical flask and it was kept at 50oC for 15 minutes for maximumextraction with occasional stirring. Then it was filtered through muslin cloth andcentrifuged at 3000g for 15 minutes to remove insoluble. The extract was storedin brown bottle.
Purification of polyphenols: The pomegranate polyphenols were purified byusing Amberlite XAD 16 resin [7]
Total Phenolic Content: It was measured using Folin Ciocalteu method Characterization of polyphenols in HPLC-UV: HPLC System: Waters Inc USA
Model 2707; Column: C18 Spherisorb ODS 250 column 4.6mm (5); ColumnTemperature : 25oC; Mobile phase A- 1.3g potassium di hydrogen phosphatesolution in water ; Mobile phase B Acetonitrile 100%; Standards used: Gallicacid, Ellagic Acid, Punicalgin.
Antioxidative Assay: DPPH scavenging activity was performed as described inSweetie Kannat, 2010
Antibacterial activity: Antibacterial well diffusion assay and Minimum inhibitoryconcentration(1-10mg/mL) was performed as described in Al-Zoreky, 2009.
Preparation and Coating of Fish: The fishes were procured in local market andbrought to lab with ice pack, it was wased with potable water, beheaded, degut-ted and made into steaks. The fishes were placed in coating formulation in theratio (1:2) in a container and kept inside the vacuum chamber for the vacuum im-pregnation at 100mmHg at room temperature for 15 minutes (Duan, Cherian, &Zhao, 2010). The coated fish samples were removed and air dried for 30 min-utes and kept inside the LDPE zipped pouches for the storage at 4oC in refriger-
ator. The samples were taken on 3, 6, 8, 10th day and all the parameters wereanalyzed until all the sample reached the maximum permissible microbial count107 CFU/g or 7 log units.
Results and Discussions
Approximately 2 fold increase of antioxidative activity of the polyphenols after purifi-cation.
Purified Powder of Pomegranate Polyphenol
Presence of ellagic acid and punicalgin.
Anti Bacterial activity represent the polyphenols are more active against grampositive bacteria than gram negative bacteria except Pseudomonas aeruginosa.While all the tested bacteria can be inhibited at 2mg/mL concentration but Sa-monella typhimurium was quite resistant, inhibited only at 4 mg/mL.
Scavenging property for free radicals of pomegranate polyphenols is greaterthan BHT IC 50 value: 16.15 g/mL (BHT- 21.2 g/mL L (Kannat et al., 2010))
COATING EXPERIMENT RESULTS
Effect on Microbial Load:
Polyphenols reduced the bacteral count and enhanced the shelf life, while pullu-lan-polyphenol coating enhanced shelflife from 3 to 6 days and addition of CMC in-to pullulan and polyphenols increased the retention and stabilized the coating fromwashout during drip loss.
Basic-Total Volatile Nitrogen has increased more in control samples while reductionupto 2.5 fold in sample containing Polyphenol. TBARS determination results shown,decreasing in TBARS value in coated sample even without polyphenols shows pres-ence of pullulan reduced oxygen diffusion. There was 4 fold reduction in the CMC-Pullulan-Polyphenol coated sample.
An edible coating has been developed using pullulan and cmc (polysaccharide) andpolyphenol from pomegranate peel.
The coating on to fish steaks increased the shelf life from 3 days to 8 days at4oC.
The present project involving the use of polyphenols from pomegranate peelhelps in waste utilization from peel
Besides increasing the shelf-life of the product, the coating helps in providing thefunctionality to the product.
The cost economics of the product can be further brought down by blending apolymer from other sources (possibly from fruit waste).
References1. N. S. Al-Zoreky, Antimicrobial activity of pomegranate (Punica granatum L.) fruit
peels., Int. J. Food Microbiol., vol. 134, no. 3, pp. 2448, Sep. 2009.2. S. Kanatt, R. Chander, and A. Sharma, Antioxidant and antimicrobial activity of
pomegranate peel extract improves the shelf life of chicken products, Int. J. food, 2010.
3. F. Debeaufort, J. a Quezada-Gallo, and A. Voilley, Edible films and coatings:tomorrows packagings: a review., Crit. Rev. Food Sci. Nutr., vol. 38, no. 4, pp.299313, May 1998.
4. O. Lie, Improving Farmed Fish Quality and Safety. Elsevier, 2008.5. J. Duan, G. Cherian, and Y. Zhao, Quality enhancement in fresh and frozen ling-
cod (Ophiodon elongates) fillets by employment of fish oil incorporated chitosancoatings, Food Chem., vol. 119, no. 2, pp. 524532, Mar. 2010.
6. M. R. Adams and M. O. Moss, Food Microbiology. Royal Society of Chemistry,2008.
7. N. Seeram, R. Lee, M. Hardy, and D. Heber, Rapid large scale purification of el-lagitannins from pomegranate husk, a by-product of the commercial juice indus-try, Sep. Purif. , 2005.
