Orange Serum Agar

Description

Orange Serum Agar is not a differential agar but a culture medium in which the orange extract provides a favourable acidic environment in which aciduric microorganisms can be recovered, including those damaged by food processing. Tryptone provides the main source of carbon and nitrogen, providing optimal growth conditions. Yeast extract supplies Group B complex vitamins that stimulate growth and the phosphate provides an osmotic buffer for cell survival. Dextrose is a supplementary source of carbon and the agar is a solidifying agent.

Technique

The International Fruchtsaft-Union (IFU) recommends the use of Orange Serum Agar in several standardised methods, using the plate count method:

1. Prepare serial 10-fold dilutions of the sample using a suitable diluent such as Buffered Peptone Water.
2. Distribute aliquots of 1 mL of the diluted sample in sterile Petri dishes.
3. Add 20 mL of molten sterile medium cooled to 45 °C, gently swirl the dish to mix the sample and medium properly.
4. Allow it to solidify and incubate at 30 ± 1 °C for 48 hours before enumeration. If there is no growth, extend the incubation to 5 days, reading daily before giving a negative result.

Generally the colonies of yeasts and moulds are distinguished by their morphology, but those of aciduric bacteria need to be Gram stained and examined microscopically to be appropriately categorised.

Physical/chemical control

Microbiological control

Melt medium - prepare plates - spiral spreading: practical range 100 ± 20 CFU; min. 50 CFU (productivity).

Microbiological control according to ISO 11133:2014/Amd 1:2018.

Microaerophilic incubation at 30 ± 1 °C for 72 ± 3 h.

Sterility control

Incubation 48 hours at 30-35 °C and 48 hours at 20-25 °C: NO GROWTH.

Check at 7 days after incubation in same conditions.

• HAYS, G.L. (1951) The isolation, cultivation and identification of organisms which have caused spoilage in frozen concentrated orange juice. Proc. Fla. State Hortic. Soc. 54:135-137.
• HAYS, G.L. & D.W. REISTER (1952) The control of 'off-odour' spoilage in frozen concentrate orange juice. Food Technol. 6:386-389.
• HATCHER, W.S., M.E. PARISH, J.L. WEIHE, D.F. SPLITTSTOESSER & B.B. WOODWARD (2001) Fruit Beverages, in Compendium of Methods for the Microbiological Examination of Foods. 4th ed., F.P. Downes & K. Ito, editors. APHA Inc., Washington D.C., USA.
• IFU Method No. 2 (1996) Total Count of Potential Spoilaging Microorganisms of Fruits and Related Products. International Federation of Fruit Juice Producers. Microbiological Methods (2004). Schweizerischer Obstverband. Postfach CH-6302 Zug.
• IFU Method No. 6 (1996) Mesophilic & Thermoduric-Thermophilic Bacteria: Spores Count. D-II Mesophilic Anaerobic Sporeforming Bacteria: Spores Count. International Federation of Fruit Juice Producers. Microbiological Methods (2004). Schweizerischer Obstverband. Postfach CH-6302 Zug.
• IFU Method No. 7 (1998) 'Sterility' Testing of 'Aseptic Filled Products', 'Commercial Sterile Products' and 'Preserved Products'. International Federation of Fruit Juice Producers. Microbiological Methods (2004). Schweizerischer Obstverband. Postfach CH-6302 Zug.
• IFU Method No. 10 (1998) Microbiological Examination of Potential Spoilaging Microorganisms of Low Acid and High pH Vegetable Products. International Federation of Fruit Juice Producers. Microbiological Methods (2004). Schweizerischer Obstverband. Postfach CH-6302 Zug.
• ISO 11133:2014/Amd 1:2018. Microbiology of food, animal feed and water. Preparation, production, storage and performance testing of culture media.
• MURDOCK, D.I., J.F. FOLINAZZO & V.S. TROY (1952) Evaluation of plating media for citrus concentrates. Food Technol. 6:181-185.
• MURDOCK, D.I. & C.H. BROKAW (1958) Sanitary control in processing citrus concentrates. I. Some specific sources of microbial contamination from fruit bins to extractors. Food Technol. 12:573-576.
• STEVENS, J.W. (1954) Preparation of dehydrated agar media containing orange juice serum. Food Technol. 8:88-91.