Page 100 - Analytical method for food addtives

58 Analytical methods for food additives

chromatograph. Start temperature program when solvent peak emerges. Measure
peak heights and calculate peak height ratios of benzoic acid/phenylacetic acid.
Use average of duplicate ratios. Peak height ratios for duplicate injections should
differ ≤5 %.

Preparation of standard curves
Transfer 1.00 mL standard solutions to five 8 mL test tubes with Teflon-lined
screw caps. Add 0.20 mL silylating agent to each tube, cap, and let stand 15 min in
oven or H O bath at 60 ºC. Inject duplicate 1 µL portions of standard solutions into
2
gas chromatograph. Use same conditions as for test portion solution. Measure peak
heights and calculate peak height ratios of benzoic acid/phenylacetic acid. Peak
height ratios for duplicate injections should differ ≤5 %. Plot weight ratios (x)
versus average peak height ratios (y) for each preservative. Calculate slope and
intercept of standard curve by method of least squares.

Calculation
y – a W′
Preservative, mg/kg ––––– × –– × 1000 [7.1]
b W

where:
b = slope of standard curve
a = intercept
y = average peak height ratio of preservative/internal standard
W = weight of test portion in g
W′ = weight of internal standard in mg.

AOAC liquid chromatographic method for benzoic acid in orange juice 8

Preparation of test samples for HPLC
Place 10.0 mL orange juice sample into 50 mL centrifuge tube and centrifuge 5 min
at 1500 g. Using 10 mL syringe, precondition C18 cartridge by passing 2 mL
methanol through cartridge, followed by 5 mL H O. Pipette 1.0 mL portion of test
2
sample supernate into syringe and through conditioned cartridge. Slowly wash
cartridge (let eluate drip by slowly pushing plunger) with 3.0 mL 2 % acetonitrile
in hexane and discard eluate. Push syringe plunger 3x, blowing air through
cartridge to eliminate excess hexane in cartridge. Add 3.0 mL methanol to syringe.
Slowly elute cartridge with 3 mL methanol and collect eluate in 5 mL graduated
centrifuge tube. Adjust eluate final volume to 3.0 mL with methanol. Pass eluate
through 0.45 µm filter into vial.

Recovery test
Determine recovery of benzoic acid in juices by dividing spiked juice sample into
2 equal portions. Filter 1 portion through 0.45 µm filter, as control sample, while
passing other portion through C18 cartridge and 0.45 µm filter. Calculate recoveries