SOP: Determination of Diacetyl Value

GIFSQ — Global Institute for Food Safety & Quality

Principle

Heavy bacterial growth while citrus juice is being concentrated can spoil the flavour of the finished product, through diacetyl (2,3-butanedione) made by acid-tolerant bacteria such as Leuconostoc and Lactobacillus. Measuring diacetyl gives a rapid index of sanitation during processing. The test is a modified Voges-Proskauer reaction: the distillate is treated with α-naphthol and alkaline creatine, and the colour developed is measured at 530 nm.

Apparatus required

• All-glass distillation apparatus
• Colorimeter tubes
• Colorimeter (reading at 530 nm)
• Measuring cylinder

Reagents

a. α-Naphthol solution: dissolve 5 g in 99% isopropanol.

b. KOH–creatine solution: dissolve 40 g of KOH in 60 ml of water, cool, add 3 g of creatine, and stir until dissolved; make up the volume to 100 ml. The reagent is stable for 3 days at 4.4 °C (40 °F) and 21 days at −17.8 °C (0 °F).

Procedure

Distil 300 ml of single-strength juice (or concentrate reconstituted to single-strength juice at 12 ± 0.2 °Brix) in the all-glass distillation apparatus; collect the first 25 ml of the distillate

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Let the distillate slide down the side of a measuring cylinder to prevent dispersion of oil; filter the distillate through Whatman No. 1 filter paper if needed to eliminate interference by cloud or oil

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Develop the colour in two stoppered test tubes:

• Blank: 10 ml distilled water + 5 ml α-naphthol solution + 2 ml KOH–creatine solution
• Sample: 10 ml distillate + 5 ml α-naphthol solution + 2 ml KOH–creatine solution

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Stopper the tubes, mix, and transfer immediately to colorimeter tubes

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Within 60 seconds of the addition of the KOH–creatine solution, measure the colour in the colorimeter at 530 nm, using the reagent blank to set the instrument to 100% transmittance

Standard curve

Prepare standard solutions containing 0.5, 1, 2, 3 and 5 ppm of diacetyl in distilled water; develop the colour from 10 ml of each standard exactly as for the sample.

Interpretation

The basis for interpreting diacetyl values for manufacturing frozen orange-juice concentrate (Murdock) is:

• 0.040–0.350 ppm diacetyl in the distillate from 12 °Brix orange juice: normal microbial activity
• 0.375–0.450 ppm: safe working range; area of caution — if subsequent tests show higher values than the preceding ones, the need for corrective action is indicated
• 0.475–0.575 ppm: danger zone — trace the source of trouble and flush the equipment with chlorinated water or detergent solution
• 0.600 ppm and higher: the product would spoil if the operation is continued

Precautions

• Read the colour within 60 seconds of adding the KOH–creatine solution — the colour changes with time.
• Do not let the reagent exceed its stated stability period.

References

Ranganna, S., Handbook of Analysis and Quality Control for Fruit and Vegetable Products, 2nd edition.

Hill, E. C., Wenzel, F. W. and Barreto, A., Food Technol. 8, 168 (1954) — as cited in the book.

Murdock, D. I., Food Technol. 21, 643 (1967) — as cited in the book.

Murdock, D. I., Food Technol. 22, 90 (1968) — as cited in the book.