DETECTION AND ANALYSIS

Analytic methods used or tested for qualitative and quantitative detection of sorbates in foods include acidimetry, bromometry, colorimetry, enzymatic, spectrophotometry, polarography, and chromatography. The most widely used methods, however, have been colorimetric (Schmidt, 1960, 1962) and spectrophotometric (Melnick and Luckman, 1954a), although chromatographic methods have gained acceptance in recent years.

Detection methods require quantitative extraction and separation of sorbic acid from the food material without food ingredient interference (DeLuca et al., 1995; Mandrou et al., 1999; Montano et al., 1995; Sofos, 2000). Extraction methods include acid-steam distillation, selective gas diffu-sion, and solvent extension using ethyl or petroleum ether, dichloromethane, and isooctane. In some foods, filtration, dialysis, or direct analysis has been used (Sofos, 1989).

Extraction by steam distillation has been used extensively, but it is time consuming and the compounds present in the food or generated by decomposition of lipid materials may interfere with colorimetric or spectrophotometric detection of sorbic acid (Harrington et al., 1962; Sofos, 1989).

Several modifications have been proposed as useful in avoiding interference and in improving the accuracy of the steam distillation procedure (Luckmann and Melnick, 1955; Sofos, 1989), whereas combinations of various treatments have also been used to improve extraction and reduce interfer-ence (Sofos, 1989). Such combinations have involved extraction under acid conditions from the steam distillate and reextraction with sodium hydroxide; dialysis and solvent extraction; successive extractions with ether, sodium hydroxide, and methylene chloride; and double distillation and ether extraction (Tjan and Konter, 1972; Noda et al., 1973; Larsson, 1983; Puttermans et al., 1983; Sofos, 1989).

The colorimetric detection of sorbic acid at an absorbance of 532 nm is an official method for quantitative determination in foods and beverages (AOAC Int. method 975.31), cheese (AOAC Int.

methods 971.15 and 975.22), and wine (AOAC Int. method 975.10) by the color reaction with α-thiobarbituric acid (Roy et al., 1976; AOAC Int., 2000). The method is simple and usually very specific (Lück, 1980). The spectrophotometric (ultraviolet/UV, absorption) procedure has also been used in many foods, including fruit products, bakery items, wine, cheese, and sausage products (Maxstadt and Karasz, 1972; Wilamowski, 1974; Stafford, 1976; Holley and Millard, 1980). After development by Melnick and Luckman (1954a), the method was modified by Alderton and Lewis (1958); it involves measurement of absorbance at 260 nm (250 to 290 nm). Special extraction and purification steps have been proposed in the literature to reduce interference problems (Sofos,

1989). The method is an official first action procedure for ground beef (AOAC Int. method 980.17) and final action procedure for wines (AOAC Int. method 974.08) and dairy products (AOAC Int.

method 974.10) (AOAC Int., 2000).

The literature contains numerous reports of chromatographic methods used or evaluated for determination of sorbate in food products (Sofos, 1989). They include gas chromatography, high-performance liquid chromatography (HPLC), thin-layer chromatography, paper chromatography, and micellar electrokinetic capillary chromatography (MECC) and ion chromatography. A gas chromatographic method is a first action procedure (AOAC Int. method 983.16) for sorbic and benzoic acids in foods (AOAC Int., 2000). Solid extraction has been used as a cleanup procedure for the determination of sorbic acid by liquid chromatography in fruit products (Mandrou et al., 1999). Others, however, have reported that cleanup procedures did not improve determination by liquid chromatography (Benassi and Cecchi, 1998). Microdialysis was used to extract sorbic acid and benzoic acid from food to be separated and detected by HPLC (Mannino and Cosio, 1996).

In addition, HPLC was found to be adequate in obtaining accurate stability data for sorbic acid in creams (de Villiers and Bergh, 2000). A rapid method for the identification and quantitation of sorbic and benzoic acids in beverages and foods by MECC has been reported (Pant and Trenerry, 1995). Ion chromatography has been shown to be in good agreement with HPLC in determinations of sorbic acid in food and pharmaceutical preparations (Chen and Wang, 2001). In addition, ion chromatography was effective in simultaneously determining artificial sweeteners, preservatives, caffeine, theobromine, and theophylline and as such may be a beneficial alternative to conventional HPLC.

A polarographic method using differential-pulse voltammetry with the use of a hanging mercury drop electrode (HMDE) was developed for the determination of sorbic acid in fruit juices and drinks. The procedure is simple and specific and is not subject to interference from many substances commonly found in soft drinks such as ethanol and benzoic acid. Moreover, it can be used for the analysis of intensely colored juice samples. Thus, the scope of application of the proposed method for the determination of sorbic acid has been extended to cover juice and drink samples that cannot be analyzed by the AOAC procedure (Fung and Luk, 1990). The use of reversed-phase high-performance liquid chromatography (RP-HPLC) was found to be a simple, rapid method with high reproducibility for determining sorbic acid and other additives in fruit juices (Zou et al., 2001).

An enzymatic method used to determine sorbic acid based on the spectrophotometric measure-ment of sorbyl CoA at 300 nm has been developed (Hofer and Jenewein, 2000). The method is based on sorbic acid being converted to sorbyl CoA with acyl CoA synthetase in the presence of coenzyme A and adenosine-5’-triphosphate. The reaction is quantified by irreversibly hydrolyzing pyrophosphate to phosphate in the presence of inorganic pyrophosphatase, where the absorbance is measured at 300 nm and is specific for sorbyl CoA, which is proportional to sorbic acid levels in the sample. The method was found to be fast, precise, and reliable and is thus well suited for routine determinations, especially for high sample throughputs (Hofer and Jenewein, 2000).

Capillary electrophoresis was also applied for detection in citrus juices, wine, and other sub-strates (Cancalon, 1999; Mercier et al., 1998; Castineira et al., 2000; Dobiasova et al., 2002).

Inhibition of microorganisms has also been evaluated as a procedure for the qualitative detection of sorbate (Ellerman, 1977). It is obvious, however, that in addition to sorbate, such inhibition may be caused by a number of other inhibitors.

REGULATORY STATUS

Several forms of sorbate are allowed for use in a variety of foods throughout the world (Sofos, 1989). This major use may expand in the future, considering their extensive testing, low toxicity, and advantages over other preservatives. In the United States, sorbic acid and potassium sorbate are GRAS, and this status has been reaffirmed (U.S. Department of Agriculture, 1978) by a select committee of the U.S. Food and Drug Administration. In addition, sorbates in the United States

Sorbic Acid and Sorbates 75 are sanctioned in more than 80 food products designated by standards of identity. According to the Code of Federal Regulations, when a food preservative is used in a food product, its common name (e.g., potassium sorbate or sorbic acid) should be listed on the product label and its function should be indicated by an explanatory description (e.g., “to maintain freshness,” “to extend shelf life,” or

“as a preservative”). The use of sorbates may be requested for any food product that allows preservatives.

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