SOP: Standard Plate Count in Fruit Juice Concentrates
GIFSQ — Global Institute for Food Safety & Quality
Principle
The plate-count method estimates the number of viable microorganisms in the sample that can grow in a specified medium; the number of visible colonies equals the number of microorganisms, expressed as viable organisms per millilitre of single-strength juice. When the number exceeds 300 per ml, the sample is serially diluted. In the citrus-processing industry, particularly for frozen concentrates, plate counts are made at various stages of manufacture at frequent intervals: a high initial count at one stage points to the source of contamination, and a high count in the finished product indicates improperly cleaned equipment or product abuse between the evaporator and packaging. In concentrates of 45 °Brix or higher, lactic-acid organisms are of no concern; in high-density concentrates yeasts are the only organisms of importance to the processor.
Apparatus required
- Sterile pipettes
- Sterile Petri dishes
- Dilution bottles with 99 ml of sterile distilled water
- Beer-can opener (for opening cans aseptically)
Medium
Orange serum agar (preparation as given in the book’s microbiology section, page 692 — the preparation recipe is not repeated here; follow the printed book for the medium formulation).
Procedure
Prepare serial dilutions of the concentrate so that the number of colonies per plate ranges from 30 to 300
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Mix the contents by shaking; open the can under aseptic conditions — flame the can and the opener to sterilize, then open one end of the can
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Withdraw 1 ml with a sterile pipette and deliver it to the dilution bottle containing 99 ml of sterile distilled water; close the bottle and shake — this gives a 1:100 dilution
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Pipette 1.0 ml to one Petri dish and 0.1 ml to another, representing the 1:100 and 1:1,000 dilutions respectively
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To prepare the 1:10,000 dilution, transfer 1.0 ml of the 1:100 dilution to 99 ml of sterile water in a bottle; pipette 1 ml of this to a sterile Petri dish
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Melt the medium, cool to 45 °C, pour 20 ml into each plate, mix by a rotating motion of the plate, and let the agar harden
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Incubate the plates at 38 °C for 48 hours; run a control using 1.0 ml of water in place of the sample, as a check on the sterility of the dilution water, medium and glassware
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Count the number of colonies after 48 hours of incubation
Calculation
Number of viable organisms per ml of sample = (Number of colonies on the Petri dish) × (Dilution factor)
Interpretation
A maximum count of 1,00,000 organisms per 100 g or ml of the concentrate is considered normal; some manufacturers set a limit of 25,000–50,000 in their packs. The count decreases after storage at −18 °C for 4 to 5 months.
Precautions
- Prepare the orange serum agar exactly as in the book’s microbiology section (page 692) — the medium formulation is part of the method.
- Open the can under aseptic conditions and flame the can and opener before sampling.
- Always run the water control plate; contaminated dilution water or medium invalidates the count.
References
Ranganna, S., Handbook of Analysis and Quality Control for Fruit and Vegetable Products, 2nd edition.