A reliable analytical method for the simultaneous determination of dl-α-tocopherol acetate and dl-α-tocopherol in foods was established by HPLC using post-column photochemical reaction with UV and fluorescence detection. For low-fat food such as fruit juice and vegetable sauce, the tocopherols were extracted with methanol containing 0.1% ascorbic acid and the extract solution was injected into the HPLC. For fatty foods such as butter and margarine, the tocopherols were extracted with a mixed solvent of acetonitrile-2-propanol (9 : 1) containing ascorbic acid. The extract was cleaned up using a Sep Pak plus C18 cartridge and the eluent from the cartridge was injected into the HPLC. The peaks corresponding to tocopherols on the chromatogram were confirmed by comparing their UV spectra with those of the standard mixture at lamp-on and lamp-off of the photochemical reactor. The recoveries of tocopherols from low-fat foods (orange juice and barbecue sauce) fortified at levels of 10 and 100 μg/kg each were 88.3 to 105.8% (RSD 0.5 to 6.0% ) and those from the fatty foods (peanut butter and margarine) fortified at 100 μg/kg each were 57.1 to 88.3% (RSD 3.0 to 6.4%). The determination limits corresponded to 10 μg/kg of the tocopherols in the low-fat foods and 20 μg/kg in the fatty foods.
In previous papers, we showed that Ginkgo biloba extract (GBE) induced hepatic cytochrome P450 (CYP) activity, in particular pentoxyresorufin O-dealkylase (PROD; corresponding to CYP2B type) in rats, and that GBE influenced the efficacy of co-administered drugs. In this study, to clarify the nature of the induction, we examined the effects of GBE samples from different sources and some major constituents of GBE on rat hepatic CYP in vitro and in vivo. In the study in vitro, eight GBE samples dose-dependently inhibited PROD activity in microsomes prepared from GBE-treated rats, and the inhibitory ratio correlated well with the content of proanthocyanidin in the GBE samples. Moreover, among six GBE constituents examined, proanthocyanidin markedly inhibited the PROD activity. However, administration of two GBE extracts with different proanthocyanidin contents to rats induced hepatic CYP activity, including PROD, to similar extents, and proanthocyanidin alone did not induce PROD activity. Furthermore, GBE samples extracted with both acetone-water and ethanol-water showed similar induction of CYPs in rats in vivo. These results suggest that most GBE samples available in Japan induce CYPs in rats regardless of the preparation method of the GBE, and that proanthocyanidin is not responsible for the induction. Further studies will be necessary to identify the constituent(s) of GBE involved in the induction of CYPs in vivo.
Headspace GC using the standard addition method has been developed for the simultaneous determination of organic solvents in natural flavorings. The procedure can be outlined as follows: an aliquot of the sample is transferred to a 10 mL vial. To each vial, a DMSO solution containing solvents at different concentrations is added as the standard solution. The vials are kept at 50°C (for automatic injection) or 40°C (for hand-operated injection) for 40 minutes. One mL of the vapor phase in each vial is injected into a gas chromatograph equipped with an Aquatic-2 column (0.25 mm i. d.×60 m). To evaluate this method, we conducted a performance study in collaboration with 10 laboratories, using ginger oleoresin. We analyzed 6 solvents (methanol, 2-propanol, acetone, dichloromethane, hexane, and 1,1,2-trichloroethene) for which the maximum residue limits are established in Japan's Specifications and Standards for Food Additives. Methanol and acetone existed in the ginger oleoresin, so only the other that four kinds of solvents were added to it. Eight of the laboratories used automatic injection, while the remaining two used hand-operated injection. Statistical analyses were conducted on the data obtained from the 8 laboratories. Repeatability standard deviations (RSDr) ranged from 4.3 to 11.4%, and reproducibility standard deviations (RSDR) ranged from 8.4 to 19.0%.
A simple method for analysis of trichothecenes [Type A: diacetoxyscirpenol, neosolaniol, HT-2 toxin, and T-2 toxin, Type B: deoxynivalenol, nivalenol, fusarenon-X, 3-acetyldeoxynivalenol, 15-acetyldeoxynivarenol] in barley tea and beer using liquid chromatography tandem mass spectrometry (LC/MS/MS) was developed. Trichothecenes were extracted with ethyl acetate-methanol (19 : 1). The solvent was evaporated to dryness and the residue was dissolved in water-methanol (3 : 1) for injection into the LC/MS/MS. The LC separation was performed with an octadecylated silica column at a flow-rate of 0.2 mL/min, using a mobile phase consisting of water, methanol and acetonitrile. MS/MS was used in multiple reaction monitoring, employing electrospray ionization (ESI-MRM). The recoveries of trichothecenes from drinks at 1 μg/L (Type A) and 10 μg/L (Type B) were 52.5-115.2% (barley tea) and 68.1-127.5% (beer). Five barley tea and ten beer samples were analyzed by this method. Trichothecenes were not detected in them. This method may have applications in quality assurance.
Inter-laboratory evaluation studies were conducted for the ELISA methods for allergic substances (buckwheat). Extracts of snack, bun and udon spiked with buckwheat standard protein at a level of 5-20 ng/mL as sample solutions were analyzed in replicate at 10 laboratories. Coefficients of variation (CVs) of the ELISA methods using a Buckwheat Protein ELISA Kit (Buckwheat kit) and a FASTKITTM Buckwheat ELISA kit (Buckwheat ELISA kit) were mostly below 10%. Mean recoveries of the buckwheat standard protein from the food extracts were over 40% in the two ELISA methods. Repeatability relative standard deviations of buckwheat standard protein in three food extracts were in the ranges of 6.8-78.5% and 5.0-33.9% for the Buckwheat kit and the Buckwheat ELISA kit, respectively. Reproducibility relative standard deviations of buckwheat standard protein in three food extracts were 11.9-69.5% and 16.5-34.1% for the Buckwheat kit and the Buckwheat ELISA kit, respectively. The detection limits of both ELISA methods were 1 ng/mL in sample solutions. These results suggest that the notified ELISA methods are reliable and reproducible for the inspection of buckwheat protein levels in extracts of snack, bun and udon.
An analytical method for the determination of 32 kinds of pesticide residues in onions, Welsh onions and mushrooms using gas chromatograph with an atomic emission detector (GC-AED) was developed. The pesticides were extracted with acetone-n-hexane (2 : 3) mixture. The crude extract was partitioned between 5% sodium chloride and ethyl acetate-n-hexane (1 : 4) mixture. The extract was passed through a Florisil® mini-column for cleanup with 10 mL of acetone-n-hexane (1 : 9) mixture. Although the sensitivity of GC-AED was inferior to that of GC-ECD, GC-AED has a superior element-selectivity. Therefore pesticide residues in foods could be analyzed more exactly by using GC-AED. Thirty-two pesticides including chlorine in onion, Welsh onion and shiitake mushroom were detected without interference. Recoveries of these pesticides from samples determined by GC-AED were 64-114%, except for a few pesticides.
Inter-laboratory evaluation studies were conducted for ELISA methods for allergic substances (peanuts). Extracts of biscuit, sauce, chocolate and butter spiked with peanut standard protein at a level of 5-20 ng/mL as sample solutions were analyzed in replicate in 10 laboratories. Coefficients of variation (CVs) of the ELISA methods using a Peanut Protein ELISA Kit (Peanut kit) and a FASTKITTM Peanut ELISA kit (Peanut ELISA kit) were mostly below 10%. Mean recoveries of the peanut standard protein from the food extracts were over 40% in the two ELISA methods. Repeatability relative standard deviations of peanut standard protein in four food extracts were in the ranges of 15.2-49.7% and 3.0-28.3% for the Peanut kit and the Peanut ELISA kit, respectively. Reproducibility relative standard deviations of peanut standard protein in four food extracts were 23.5-44.4%, 9.6-28.4% for the Peanut kit and the Peanut ELISA kit, respectively. The detection limits of both ELISA methods were 2-2.5 ng/mL in sample solutions. These results suggested that the notified ELISA methods are reliable and reproducible for the inspection of peanut protein levels in extracts of biscuit, sauce, chocolate and butter.
The modified Rankine colorimetric method for measuring sulfite added to food as a food additive has a low determination limit and is little influenced by interfering substances from foods. However, it can give erroneous results for foods containing Liliaceae Allium. So, four different methods, alkaline titration, a colorimetric method, ion chromatography and qualitative analysis with potassium iodate-starch paper, were examined. It was found that the sodium azide used in the colorimetric method forms sulfur dioxide during bubbling and heating. The proposed colorimetric method can be applied to food containing sulfur compounds, if sodium azide is omitted and 1% triethanolamine solution is used as an absorbent instead of 0.1 mol/L sodium hydroxide solution.
Daily intakes of 12 phenols which are possible endocrine disruptors were estimated in hospital meals from 2000 to 2001. 4-Nonylphenol (4-NP (mix)) and bisphenol A (BPA) were detected at levels of 5.0 to 19.4 ng/g and 0.2 to 1.1 ng/g, respectively. 4-tert-Butylphenol, 4-n-pentylphenol, 4-tert-pentylphenol, 4-n-hexylphenol, 4-n-heptylphenol and 4-tert-octylphenol were detected at levels of 0.1 to 2.4 μg/g. The daily intakes of 4-NP (mix) and BPA were 5.8 μg/day and 0.42 μg/day, respectively. The daily intakes of other phenols were less than 1 μg/day.