sirindhorn international institute of technology thammasat university growths, antibacterial and...
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Sirindhorn International Institute of TechnologyThammasat University
GROWTHS, ANTIBACTERIAL AND ANTIFUNGAL ACTIVITIES OF KEFIR GROWN
IN DIFFERENT CULTURE MEDIASathika Jariyatamkitti, Somrutai Kitchalao, Pattarawan Wuttisan,
Rachnarin Nitisoravut*, Issara Poljungreedand and Siwarutt Boonyarattanakalin
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OUTLINES
• Background• Statement of Problem•Objectives•Methodology• Results and Discussion• Conclusion
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WHAT IS KEFIR?
Kefir Fermented drink Natural prebiotic: rich in
enzymes and beneficial live bacteria or microbial
Benefits Nutrition Additive product Therapeutic effects http://ecofriendlyfreckles.blogspot.com/2010/01/eco-friendly-freckles-is-now-
selling.html
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WHAT IS KEFIR GRAINS?
Diameter: 3 to 20 mm
Grain matrix
Polysaccharide
Protein (dry weight)
Cellular debris and unknown components
Microorganisms: A complex symbiotic relationship
Mostly lactic acid bacteria and yeasts
http://en.wikipedia.org/wiki/File:Kefirpilze.jpg
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WHAT ARE KEFIRANS?
Exopolysaccharides (EPS)Mainly produced from bacteriaProvide…
Specific rheology and texture of fermented milk products
Biological Properties- Antimicrobial and
antifungal- Antitumor
o Fernando Lopitz-Otsoa, A. R., Natalia Elguezabal and Javier Garaizar, Rev Lbaroam Micol, (2006) 23, 67.
o Hsi-Chia Chen, S.-Y. W., Ming-Ju Chen, Food Microbiology, (2008) 25, 492.
- Anti-inflammatory- Antimutagenic and
antioxidant
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STATEMENT OF PROBLEM
Can milk kefir grown in media other than cow milk Would kefir grown in other media possess health benefits Alternative culturing liquid… Tropical country Plenty of coconut trees Relatively Cheap
Growth of kefir grains grown in three different culture mediaMicroorganism profiles of kefir and kefir grainsCompare antibacterial and antifungal activities of milk, coconut milk and coconut water kefir
Coconut milkCoconut water
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OBJECTIVES
To see the effects of coconut milk and coconut water, as a culturing medium for kefir, on kefir culture growth
To classify microorganism profiles in kefir and kefir grains
To examine antibacterial and antifungal activities of kefir products
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METHODOLOGY
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ISOLATION AND CULTIVATION OF LACTIC ACID BACTERIA, ACETIC ACID BACTERIA AND YEASTS
Suspend grains in phosphate buffer and homogenize using mortar
Perform serial dilutions for enumerations
Isolate bacteria using MRS agar, DECA agar, and YM agar and incubate
Count and observe the colonies
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ANTIBACTERIAL AND ANTIFUNGAL TESTING
Inoculum Preparation
Inoculation of Test Plates
Interpreting Results
Application of Disc to Inoculated Plates
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ANTIBACTERIAL AND ANTIFUNGAL TESTING
Inoculum Preparation
Inoculation of Test Plates
Interpreting Results
Application of Disc to Inoculated Plates
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INOCULUM PREPARATION
Select isolated colonies and transfer them into a broth tube
Incubate the broth culture
Compare the turbidity with 0.5 McFarland Standard
http://www.bio-world.com/productinfo/3_43_287_688/10171/Potato-Dextrose-Broth.html
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ANTIBACTERIAL AND ANTIFUNGAL TESTING
Inoculum Preparation
Inoculation of Test Plates
Interpreting Results
Application of Disc to Inoculated Plates
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INOCULATION OF TEST PLATES
Streak the swab over the entire agar surface
http://www.ehow.com/info_8681698_purpose-streaking-agar-plate.htmlhttp://www.flickr.com/photos/medmicro/page10/
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ANTIBACTERIAL AND ANTIFUNGAL TESTING
Inoculum Preparation
Inoculation of Test Plates
Interpreting Results
Application of Disc to Inoculated Plates
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APPLICATION OF DISC TO INOCULATED PLATES
Place on inoculated plates
Incubate plates
http://butterflyandbiotech.blogspot.com/2012/06/pepper-antimicrobial-effect.html
Soak ø5 mm no.1 filter paper by fermented solutions and positive
control solutions
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ANTIBACTERIAL AND ANTIFUNGAL TESTING
Inoculum Preparation
Inoculation of Test Plates
Interpreting Results
Application of Disc to Inoculated Plates
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INOCULATION OF TEST PLATES
Measure the diameter of clear zones
Sliding calipers
A ruler
http://ftp.cdc.gov/pub/infectious_diseases/artesting/images_photos/disk%20diffusion/
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RESULTS AND DISCUSSION
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RESULTS AND DISCUSSIONCoconut milk and coconut water can be used as an alternative culturing medium for kefir culture
Growth data for kefir grains grown in three different culture media (50 mL) at 25°C
4.7g/100g Sugar level
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RESULTS AND DISCUSSIONCoconut milk and coconut water can be used as an alternative culturing medium for kefir culture
Growth data for kefir grains grown in three different culture media (50 mL) at 25°C
2.8g/100g Sugar level
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RESULTS AND DISCUSSIONCoconut milk and coconut water can be used as an alternative culturing medium for kefir culture
Growth data for kefir grains grown in three different culture media (50 mL) at 25°C
2.6g/100g Sugar level
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RESULTS AND DISCUSSION
Species of bacteria and yeasts in kefir and kefir grains can be found
Selective plates GECA MRS YMShape of microorganisms
Bacillus Coccus Yeast Bacillus Coccus Yeast Bacillus Coccus Yeast
Milk kefir + + - + + - + + -
Milk kefir grain + + + + - - + - +
Coconut milk kefir + - + + - - + - +Coconut milk kefir grain + - + - - - + - +
Coconut water kefir + - + + - - + - +Coconut water kefir grain + - + + - - + - +
*+ indicates the presence of the microbial where – indicates the absence of the microbials
Table 1*: Microscopic examination of kefir and kefir grains in three different culturing media
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RESULTS AND DISCUSSION
Species of bacteria and yeasts in kefir grains can be found
Selective plates GECA MRS YMShape of microorganisms
Bacillus Coccus Yeast Bacillus Coccus Yeast Bacillus Coccus Yeast
Milk kefir + + - + + - + + -
Milk kefir grain + + + + - - + - +
Coconut milk kefir + - + + - - + - +Coconut milk kefir grain + - + - - + - +
Coconut water kefir + - + + - - + - +Coconut water kefir grain + - + + - - + - +
*+ indicates the presence of the microbial where – indicates the absence of the microbials
Table 1*: Microscopic examination of kefir and kefir grains in three different culturing media
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RESULTS AND DISCUSSIONKefir grown in coconut milk and coconut water have antibacterial and antifungal activities
Strains
Antibiotic Agents Kefir Products
Flucozole Ampicillin Chloram-phenicol Penicillin Milk Coconut
milk Coconut water
E. Coli (-) 0.0 15.5 25.0 0.0 8.9 11.5 11.0S. Typhi (-) 0.0 27.0 35.5 8.5 11.5 15.5 11.0S. Aureus (+) 8.0 29.5 25.5 33.5 12.5 12.0 10.0B. Subtilis (+) 10.5 51.5 34.5 53.5 11.5 13.0 14.5C. albican 18.5 0.0 20.5 0.0 11.0 0.0 0.0
** The results represent the mean of clear zone diameters (mm). The non fermented media (milk, coconut milk and coconut water) show no inhibition zone
Table 2**: Antibacterial and antifungal activities using disk diffusion method of 24 h kefir fermentation in various culturing media
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RESULTS AND DISCUSSIONKefir grown in coconut milk and coconut water have antibacterial and antifungal activities
Strains
Antibiotic Agents Kefir Products
Flucozole Ampicillin Chloram-phenicol Penicillin Milk Coconut
milk Coconut water
E. Coli (-) 0.0 15.5 25.0 0.0 8.9 11.5 11.0S. Typhi (-) 0.0 27.0 35.5 8.5 11.5 15.5 11.0S. Aureus (+) 8.0 29.5 25.5 33.5 12.5 12.0 10.0B. Subtilis (+) 10.5 51.5 34.5 53.5 11.5 13.0 14.5C. albican 18.5 0.0 20.5 0.0 11.0 0.0 0.0
** The results represent the mean of clear zone diameters (mm). The non fermented media (milk, coconut milk and coconut water) show no inhibition zone
Table 2**: Antibacterial and antifungal activities using disk diffusion method of 24 h kefir fermentation in various culturing media
Sirindhorn International Institute of TechnologyThammasat University
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RESULTS AND DISCUSSIONKefir grown in coconut milk and coconut water have antibacterial and antifungal activities
Strains
Antibiotic Agents Kefir Products
Flucozole Ampicillin Chloram-phenicol Penicillin Milk Coconut
milk Coconut water
E. Coli (-) 0.0 15.5 25.0 0.0 8.9 11.5 11.0S. Typhi (-) 0.0 27.0 35.5 8.5 11.5 15.5 11.0S. Aureus (+) 8.0 29.5 25.5 33.5 12.5 12.0 10.0B. Subtilis (+) 10.5 51.5 34.5 53.5 11.5 13.0 14.5C. albican 18.5 0.0 20.5 0.0 11.0 0.0 0.0
** The results represent the mean of clear zone diameters (mm). The non fermented media (milk, coconut milk and coconut water) show no inhibition zone
Table 2**: Antibacterial and antifungal activities using disk diffusion method of 24 h kefir fermentation in various culturing media
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CONCLUSION
Coconut milk and coconut water can be used as the alternative to cow milk
Bacillus and Yeasts All kefir and grainsCoccus Milk kefir and grains
Antibacterial activities- Coconut milk kefir- Coconut water kefirAntifungal activities- Only milk kefir
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REFERENCESSemih Otes , a. O. C., Pakistan Journal of Nutrition, (2003) 2, 54.Fernando Lopitz-Otsoa, A. R., Natalia Elguezabal and Javier Garaizar, Rev Lbaroam Micol, (2006) 23, 67.Hsi-Chia Chen, S.-Y. W., Ming-Ju Chen, Food Microbiology, (2008) 25, 492.Zhou Jianzhonga, L. X., Jiang Hanhub, Dong Mingshengb, Food Microbiology, (2009) 26, 770.Chujian Chen, McGill University Montreal, (2005), p 222.Prof.Dr.Lalitha Kameswaran, Manual on Antimicrobial Susceptibility Testing, In (2004).http://www.neogen.com/Acumedia/pdf/ProdInfo/7101_PI.pdf (Retrieved September 26,2012)Kamila Leite Rodrigues, L. R. G. C., Jose Carlos Tavares Carvalho, Joao Evangelista and Jose Mauricil Schneedorf, International Journal of Antimicrobial Agents, (2005) 25, 404.Michel Andre Wattiaux, Milk Composition and Nutritional Value, In Dairy Essentials - Lactation and Milking.Muhammad Majeed, P. D., Lakshmi Prakash, Ph.D., CococinTM: The Nourishment FactorTM Freeze-Dried Coconut Water Solids, In (2007).http://www.fineli.fi/food.php?foodid=29014&lan g=en (Retrieved September 15, 2012)
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QUESTIONS?