Gas Chromatography Analysis of Benzoic Acid and Sorbic Acid in Food

1 Scope of application

Joint testing of apple juice, almond paste and fish sauce (0.5-2 g/kg level), representative of carbohydrate-rich slurries, fat and carbohydrate-rich foods, and protein-rich foods.

2 Principle summary

Benzoic acid and sorbic acid were extracted with *** and then partitioned in aqueous NaOH and CH 2 Cl 2 . The acid is separated from the food and converted to trimethylsilyl (TMS) ester, which is determined by GC. *** and hexanoic acid were used as internal standards for benzoic acid and sorbic acid, respectively.

3 instruments and reagents

3.1 Instrument

(a) Lutron GC-9860 Gas Chromatograph - using a linear temperature-programming furnace, flame ionization detector and recorder. The following conditions can be employed: a column, 1.8 m x 2 mm (inside diameter) filled with a spiral glass tube coated with a 3% OV-l 100-120 mesh Varaport 30 carrier. Operating temperature: furnace temperature 80-120 ° C, 8 ° C / min; injection chamber 200 ° C, detector 280 ° C. The nitrogen carrier gas flow rate was 20 ml/min. The approximate retention times of caproic acid, sorbic acid, benzoic acid and *** were 2.5, 4, 5 and 6 min, respectively.

(b) Centrifuge - with 30 ml and 200 ml centrifuge bottles.

(c) Mechanical oscillators - Bülher SM-B or similar products.

3.2 Reagents

Analytical reagents were used in all analyses.

(a) Internal standard solution - 250 mg *** and 250 mg hexanoic acid were dissolved in 100 ml of a 3% KOH aqueous solution.

(b) Silylating agent - N-methyl-N-trimethylsilane-trifluoroacetic acid amine (MSTFA).

(c) Standard solution - a mixed standard solution of benzoic acid, sorbic acid, *** and hexanoic acid was prepared with CHCl3 at the following concentrations:

1) 200, 200, 750 and 750 μg / ml

2) 400, 400, 750 and 750 μg / ml

3) 600, 600, 750 and 750 μg/ml

4) 800, 800, 750 and 750 μg/ml

5) 1000, 1000, 750 and 750 μg / ml

4 test process

4.1. Preparation of samples

Mix the sample in a mechanical mixer. If it is difficult to mix because the sample is very thick, any other technique can be used to ensure uniformity of the sample.

4.2. Extraction

(a) General method - Accurately weigh 5.0 g of the mixed sample and place it in a 30 ml centrifuge tube with a Teflon-lined nut. Add 3.00 ml of internal standard solution, 1.5 ml of H2SO4 (1+5), 5 g of sand and 15 ml of ***. Tighten the nut and do not leak. Mechanically shake for 5 min, then centrifuge at 1500 x g for 10 min. The layer was transferred to a 250 ml separatory funnel with a pipette, and each time 15 ml *** was used, and the extraction was repeated twice.

The combined *** phase was extracted twice with 15 ml of 0.5 N NaOH and 10 ml of saturated NaCl solution each time. The aqueous layer was collected, placed in a 250 ml sep. funnel, and 2 portions of methyl orange was added, and the solution was acidified to pH=1 with HCl (1+1). Continuous extraction was carried out with 75, 50 and 50 ml of CH 2 Cl 2 , respectively. If emulsification occurs, 10 ml of a saturated solution of NaCl is added. The CH 2 Cl 2 extract was decanted through a filter containing 15 g of anhydrous Na 2 SO 4 into a 250 ml round bottom flask, and the CH 2 Cl 2 solution was evaporated to dryness at 40 ° C using a rotary evaporator.

(b) Cheese and paste-like viscous food - Accurately weigh 5.0 g of the homogeneous sample into a 200 ml centrifuge bottle. 15 ml of water was added and stirred with a glass rod until the sample was suspended in the aqueous phase. Add 3.00 ml of internal standard solution, 1.5 ml of H2S04 (1+5) and 25 ml of ***. Carefully plug the centrifuge bottle and check for leaks. Mechanically shake for 5 min, then centrifuge at 2000 x g for 10 min. The layer of *** was pipetted into a 250 ml separatory funnel. The extraction was repeated twice with 25 ml *** each time. Press (a) to continue the operation from "Extracting the merged *** phase".

4.3. Derivatization gas chromatography

10.0 ml of CHCl3 was added to a 250 ml round bottom flask containing the residue. Plug tightly and shake by hand for 2 min. Pipette 1.00 ml of CHCl3 solution into a test tube of 8 ml of Teflon-lined cap and add 0.20 ml of silylation reagent. Cover it and place it in a 60 ° C oven or on a water bath for 15 min. The 1 μl sample solution was repeatedly injected into the gas chromatograph, and when the solvent peak appeared, the temperature was started to be programmed. The peak height was measured and the peak height ratio of benzoic acid/*** and sorbic acid/hexanoic acid was calculated. The peak height ratio of the repeated measurements was averaged. The difference in peak height ratio for repeated injections should be ≤ 5%.

4.4. Preparation of standard curve

Transfer 1.00 ml of the standard solution to 5 tubes of 8 ml Teflon-lined caps. 0.20 ml of silanization reagent was added to each tube. Cover it and place it in a 60 ° C oven or on a water bath for 15 min. Repeat the injection of 1 μl of the standard solution into the chromatograph using the same conditions as the sample solution. The peak height was measured and the peak ratio of benzoic acid/*** and sorbic acid/hexanoic acid was calculated separately. The peak-to-height ratio difference for repeated injections should be ≤5%. The average peak height ratio (y) is plotted against the weight ratio (x) of each preservative, and the slope and intercept of the standard curve are calculated by the least squares method.

4.5. Calculation

Preservative, mg/kg=[(ya)/b]×(W'×W)×l000

Where b = the slope of the standard curve; a = intercept; y = (preservative / internal standard) the average of the peak height ratio; W = the amount of sample in g; W' = within the unit of mg Scalar.