Andrew Damant

Ochratoxin A in dried vine fruit: method development and survey

A method is described for the determination of concentrations of the mycotoxin ochratoxin A in dried vine fruits (currants, raisins and sultanas) using acidic methanolic extraction, immunoaffinity chromatography clean-up and HPLC determination. The limit of detection was estimated as 0.2 microgram/kg, and recoveries of 63-77% were achieved at 5 micrograms/kg. HPLC-mass spectrometric confirmation of the identity of ochratoxin was obtained. Ochratoxin A and aflatoxins were determined in 60 samples of retail dried vine fruits purchased in the United Kingdom. Ochratoxin A was found in excess of 0.2 microgram/kg in 19 of 20 currant, 17 of 20 sultana and 17 of 20 raisin samples examined, an overall incidence of 88%. The maximum level found was 53.6 micrograms/kg. No aflatoxin was found in any sample analysed, using a method with a detection limit of 0.2 microgram/kg for each of aflatoxin B1, B2, G1 and G2.

Composition and role of tapetal lipid bodies in the biogenesis of the pollen coat of Brassica napus

Abstract. The composition of the two major lipidic organelles of the tapetum of Brassica napus L. has been determined. Elaioplasts contained numerous small (0.2–0.6 μm) lipid bodies that were largely made up of sterol esters and triacylglycerols, with monogalactosyldiacylglycerol as the major polar lipid. This is the first report in any species of the presence of non-cytosolic, sterol ester-rich, lipid bodies. The elaioplast lipid bodies also contained 34- and 36-kDa proteins which were shown by N-terminal sequencing to be homologous to fibrillin and other plastid lipid-associated proteins. Tapetosomes contained mainly polyunsaturated triacylglycerols and associated phospholipids plus a diverse class of oleosin-like proteins. The pollen coat, which is derived from tapetosomes and elaioplasts, was largely made up of sterol esters and the C-terminal domains of the oleosin-like proteins, but contained virtually no galactolipids, triacylglycerols or plastid lipid-associated proteins. The sterol compositions of the elaioplast and pollen coat were almost identical, consisting of stigmasterol > campestdienol > campesterol > sitosterol ≫ cholesterol, which is consistent with the majority of the pollen coat lipids being derived from elaioplasts. These data demonstrate that there is substantial remodelling of both the lipid and protein components of elaioplasts and tapetosomes following their release into the anther locule from lysed tapetal cells, and that components of both organelles contribute to the formation of the lipidic coating of mature pollen grains.

Thermal stability of polyethylene terephthalate (PET): Oligomer distribution and formation of volatiles

Two ovenable PET (polyethylene terephthalate) samples were investigated under severe heating conditions and oligomers and volatile substances were analysed as potential migrants into foods. The samples were tested for migration into water, 3% acetic acid and 15% ethanol solution for 1 hour at 95°C. Overall migration and the specific migration of terephthalic acid, ethylene glycol and diethylene glycol were all very low. The plastics were heated at 150°C, 260°C and 270°C, for 5 minutes 30 minutes and 60 minutes. Oligomer analysis by LC/MS (liquid chromatography-MS) showed that the concentration of the second series alicyclic oligomers increased up to 15-fold on heating whereas the major oligomer fraction, the cyclic trimer, tetramer, pentamer and hexamer showed only minor concentration changes with heating. Volatiles evolved by the samples were trapped on a Tenax trap and identified by GC/MS (gas chromatography-MS). They were few in number and low in concentration and none merited migration tests. It is concluded that even when tested up to melting point, PET plastics of this type have good temperature stability and are well suited for high-temperature food contact applications. Copyright

A comparison of the Kjeldahl and Dumas methods for the determination of protein in foods, using data from a proficiency testing scheme.

Both the Kjeldahl and the Dumas methods for the determination of protein in foodstuffs are currently in use, but the empirical nitrogen factors used to convert the determined nitrogen content to protein content are based on the Kjeldahl method alone. Non-equivalence between the two methods could therefore result in some laboratories reporting an incorrect protein content. We report here a study using data accumulated over several years in the results of a proficiency testing scheme. On average the Dumas method provided results that were relatively higher by about 1.4% than the Kjeldahl method, but the difference between the methods depended on the type of foodstuff. The methodology of looking for bias between analytical methods is critically discussed.

Analytical Methods for Food Additives

Sunset Yellow Azorubine (Carmoisine) Copper complexes of chlorophylls and chlorophyllins Caramel class III Annatto extracts Sorbic acid and its salts Benzoic acid Sulphites Nitrites Fumaric acid and its salts Gallates BHA L-tartaric acid and its salts Adipic acid and its salts Propylene glycol (propan-1,2-diol) Karaya gum Polysorbates Ammonium phosphatides Sucrose acetate isobutyrate Mono/diacetyl tartaric acid esters of mono/diglycerides of fatty acids Polyglycerol esters of polycondensed fatty acids of caster oil Stearoyl lactylates Stearyl tartrate Sorbitan esters Aluminium Saccharin.

Determination of dimetridazole, ronidazole and their common metabolite in poultry muscle and eggs by high performance liquid chromatography with UV detection and confirmatory analysis by atmospheric pressure chemical ionisation mass spectrometry†‡

A method was developed for the determination of the nitroimidazole compounds dimetridazole (DMZ) and ronidazole (RNZ) and their common metabolite, 2-hydroxymethyl-1-methyl-5-nitroimidazole (2-OH-M). Extracts obtained from a clean-up process using strong cation exchange (SCX) solid phase extraction (SPE) can be analysed either by high performance liquid chromatography with UV detection (HPLC-UV) or by high performance liquid chromatography with atmospheric pressure chemical ionisation mass spectrometry (HPLC-APCI-MS) as a confirmatory method. Up to 20 samples can be extracted in approximately 4 h. The HPLC-UV analysis had a limit of detection of 0.5 microgram kg-1. Validation in chicken muscle fortified at a concentration of 5 micrograms kg-1 gave recoveries of 75% DMZ, 77% RNZ and 81% 2-OH-M with RSDs of 16.4, 11.3 and 14.0%, respectively (n = 17). Validation in egg fortified at the same concentration gave recoveries of 77% DMZ, 80% RNZ and 80% 2-OH-M, with RSDs of 14.9, 22.0 and 18.2%, respectively (n = 18). The limit of detection of the HPLC-APCI-MS method was 0.1 microgram kg-1 for DMZ and RNZ and 0.5 microgram kg-1 for 2-OH-M. This method gave mean recoveries in fortified egg samples of 65% DMZ, 87% RNZ and 75% 2-OH-M with RSDs of 22, 11 and 14%, respectively (n = 10). The ratios of the peak areas of the molecular ion and a fragment ion were monitored as added confirmation of the presence of the analyte. Both the HPLC-UV screening procedure and the HPLC-APCI-MS confirmatory method have subsequently been used for the analysis of several hundred samples as part of UK surveillance programmes.

Qualitative liquid chromatographic-atmospheric-pressure chemical-ionisation mass spectrometric analysis of polyethylene terephthalate oligomers

Abstract The oligomeric fraction of polyethylene terephthalate (PET) has been studied as it has the potential to migrate to foods and beverages packaged in virgin and recovered PET plastics. We have applied positive ion atmospheric-pressure chemical ionisation (APCI) to extracts of food-grade PET resin and beverage bottles. A reversed-phase HPLC system was connected directly to a VG Platform mass spectrometer. An acetonitrile-water-acetic acid gradient elution was performed. Low APCI probe temperatures (as appropriate for a polyethylene glycol calibrant) produced no significant ions from a PET cyclic trimer standard. A high probe temperature of 500–600°C gave a strong protonated molecular ion. Characteristic spectra of the cyclic oligomers from the trimer to the heptamer were obtained. A second homologous series of substances 44 mass units higher than each PET oligomers eluted prior to each PET oligomer. The mass spectra indicated these to be oligomers with one monoethylene glycol unit replaced by a diethylene glycol unit. To our knowledge this is the first time that cyclic oligomers above the tetramer have been confirmed by LC-MS. The technique was considerably more sensitive than published thermospray methods and gave food spectra with sub-microgram quantities injected. This work demonstrates the advantages of APCI over thermospray as an MS technique for substances of this type.

The duplicate method of uncertainty estimation: are eight targets enough?

This paper presents methods for calculating confidence intervals for estimates of sampling uncertainty (s(samp)) and analytical uncertainty (s(anal)) using the chi-squared distribution. These uncertainty estimates are derived from application of the duplicate method, which recommends a minimum of eight duplicate samples. The methods are applied to two case studies--moisture in butter and nitrate in lettuce. Use of the recommended minimum of eight duplicate samples is justified for both case studies as the confidence intervals calculated using greater than eight duplicates did not show any appreciable reduction in width. It is considered that eight duplicates provide estimates of uncertainty that are both acceptably accurate and cost effective.

Reactions of epoxide monomers in food simulants used to test plastics for migration

The reactions of four epoxides used as monomers for food contact plastics were studied in the food simulants distilled water, 15% aqueous ethanol, 3% aqueous acetic acid and olive oil. Loss of the parent substance and formation of products was monitored to establish the transformation products to be expected in each simulant following migration testing of plastics. Each epoxide was stable in olive oil but suffered extensive loss in the three aqueous simulants. Reaction half-lives were from < 1 to 10 h in aqueous acetic acid, 25-63 h in distilled water, and 33-87 h in aqueous ethanol simulant. Hydrolysis to the diol was the main reaction pathway. Epoxide ring opening in aqueous ethanol simulant gave the diol and also the diol monoethyl ether. It is concluded that, for aqueous simulants and by implication for most foods, testing plastics against specific migration limits for epoxides is not likely to give reliable results due to their reactivity. The present EC mode of control for these reactive monomers, via compositional limits in food contact plastics, is more practical since the hydrolysis products are less toxic than the parent epoxide.

Single-laboratory validation of a GC/MS method for the determination of 27 polycyclic aromatic hydrocarbons (PAHs) in oils and fats

A protocol for the measurement of 27 polycyclic aromatic hydrocarbons (PAHs) in vegetable oils by GC/MS has undergone single-laboratory validation. PAHs were measured in three oils (olive pomace, sunflower and coconut oil). Five samples of each oil (one unfortified, and four fortified at concentrations between 2 and 50 µg kg−1) were analysed in replicate (four times in separate runs). Two samples (one unfortified and one fortified at 2 µg kg−1) of five oils (virgin olive oil, grapeseed oil, toasted sesame oil, olive margarine and palm oil) were also analysed. The validation included an assessment of measurement bias from the results of 120 measurements of a certified reference material (coconut oil BCR CRM458 certified for six PAHs). The method is capable of reliably detecting 26 out of 27 PAHs, at concentration <2 µg kg−1 which is the European Union maximum limit for benzo[a]pyrene, in vegetable oils, olive pomace oil, sunflower oil and coconut oil. Quantitative results were obtained that are fit for purpose for concentrations from <2 to 50 µg kg−1 for 24 out of 27 PAHs in olive pomace oil, sunflower oil and coconut oil. The reliable detection of 2 µg kg−1 of PAHs in five additional oils (virgin olive oil, grapeseed oil, toasted sesame oil, olive margarine and palm oil) has been demonstrated. The method failed to produce fit-for-purpose results for the measurement of dibenzo[a,h]pyrene, anthanthrene and cyclopenta[c,d]pyrene. The reason for the failure was the large variation in results. The likely cause was the lack of availability of 13C isotope internal standards for these PAHs at the time of the study. The protocol has been shown to be fit-for-purpose and is suitable for formal validation by inter-laboratory collaborative study.