Peyote identification on the basis of differences in morphology, mescaline content, and trnL/trnF sequence between Lophophora williamsii and L. diffusa.

Genus Lophophora (Cactaceae) has two species: Lophophora williamsii Coulter, which is called peyote, and L. diffusa Bravo. Although it was reported that L. williamsii contained mescaline and L. diffusa did not, we found L. williamsii specimens that did not contain mescaline. This finding indicated that the two species could not be differentiated in terms of mescaline content. Moreover, the relationship between mescaline content and morphology of the two species is also unknown. In this study, we attempted to clarify the difference in morphology, mescaline content, and DNA alignment of the chloroplast trnL/trnF region between L. williamsii and L. diffusa. As a result, L. williamsii specimens were classified into two groups. Group 1 had small protuberances on the epidermis, contained mescaline, and the analyzed region on the trnL/trnF sequence was 881 base pairs (bp) long in all except one (877 bp). Group 2 had large protuberances on the epidermis, did not contain mescaline, and the analyzed region was 893 bp long. On the other hand, L. diffusa had medium-sized protuberances on the epidermis, did not contain mescaline, and the analyzed region was 903 bp long. Also investigated was the potential application of the PCR-restriction fragment length polymorphism (RFLP) method as a means of identification based on the trnL/trnF sequence. By applying the PCR-RFLP method, the two species could be distinguished and L. williamsii specimens could be differentiated into group 1 and group 2.

Differentiation between sulfoaildenafil and its analogs.

An analog of aildenafil, which is a potent and highly selective inhibitor of phosphodiesterase 5, was found in a dietary supplement marketed for enhancement of sexual function. The compound was isolated by silica gel column chromatography, and its structure was identified by means of 13C-NMR spectrometry, 1H-NMR spectrometry, high-resolution MS, and X-ray structure determination. The compound was identified to be sulfoaildenafil (other names: thioaildenafil, dimethyl sildenafil thione, and thiomethisosildenafil). Sulfoaildenafil is very similar to the compound thiohomosildenafil. As it is difficult to distinguish between them by LC-photodiode array detector analysis, ultra-performance LC (UPLC)/MS, ion trap LC/MS/MS (LC/IT-MS/MS), and GC/MS were performed. The mass spectra of thiohomosildenafil by UPLC/MS and LC/IT-MS/MS showed mass fragments of m/z 58, 72, and 355, and the mass spectrum by GC/MS showed mass fragments of m/z 56, 72, and 420. Some of these fragments had low intensities, but they were useful for distinguishing between the two compounds. The relationship between aildenafil (other names: dimethylsildenafil and methisosildenafil) and homosildenafil is similar to that between sulfoaildenafil and thiohomosildenafil. Therefore, these compounds were also examined.

Liquid chromatographic analysis of cinchona alkaloids in beverages.

A method for the determination of Cinchona extract (whose main components are the alkaloids cinchonine, cinchonidine, quinidine, and quinine) in beverages by liquid chromatography was developed. A beverage with an alcohol content of more than 10% was loaded onto an OASIS HLB solid-phase extraction cartridge, after it was adjusted to pH 10 with 28% ammonium hydroxide. Other beverages were centrifuged at 4000 rpm for 5 min, and the supernatant was loaded onto the cartridge. The cartridge was washed with water followed by 15% methanol, and the Cinchona alkaloids were eluted with methanol. The Cinchona alkaloids in the eluate were chromatographed on an L-column ODS (4.6 mm id x 150 mm) with methanol and 20 mmol/L potassium dihydrogen phosphate (3 + 7) as the mobile phase. Cinchona alkaloids were monitored with an ultraviolet (UV) detector at 230 nm, and with a fluorescence detector at 405 nm for cinchonine and cinchonidine and 450 nm for quinidine and quinine (excitation at 235 nm). The calibration curves for Cinchona alkaloids with the UV detector showed good linearity in the range of 2-400 microg/mL. The detection limit of each Cinchona alkaloid, taken to be the concentration at which the absorption spectrum could be identified, was 2 microg/mL. The recovery of Cinchona alkaloids added at a level of 100 microg/g to various kinds of beverages was 87.6-96.5%, and the coefficients of variation were less than 3.3%. A number of beverage samples, some labeled to contain bitter substances, were analyzed by the proposed method. Quinine was detected in 2 samples of carbonated beverage.

[Confirmation of non-permitted dyes detected in Akasu (red vinegar) by LC/MS].

Suitable liquid chromatography/mass spetrometry (LC/MS) conditions were examined for Amaranth, Red 2G (R2G), Azo Rubine (Azo), Fast Red E (FRE) and Brilliant Blue FCF, which were detected in Akasu, a red vinegar made in Hong Kong from sake lees, on both thin layer chromatography (TLC) and photodiode array high-performance liquid chromatography (PDA-HPLC). Molecular-related ions for each dye were detected with excellent sensitivity by LC/MS using electro-spray ionization with negative ion mode, capillary voltage 3.00 kV, cone voltage 50 V and desolvation temperature 400 degrees C. LC/MS analysis of refined Akasu under these conditions enabled us to obtain clear mass spectra of R2G, Azo and FRE, which were present at trace levels in the Akasu. The results were consistent with those from TLC and PDA-HPLC. These experiments suggested that LC/MS analysis is applicable for confirmation of dyes in food.

[Simulation of migration from a multi-layer laminated film intended for retort foods].

Migration from multi-layer laminated film pouches intended for retort foods was investigated through HPLC analysis with a fluorescence detector, and measurements of residue on evaporation, consumption of potassium permanganate and total organic carbon. HPLC analysis revealed that the levels of migrants in oil and the water which were heated in the pouches (121 degrees C, 30 min) were ten times of those in the corresponding official simulants under the official conditions; n-heptane (25 degrees C, 60 min), and water (95 degrees C, 30 min). Bisphenol A diglycidyl ether and related compounds were found in the oil and the water heated in the pouches, as well as in the simulants. These compounds were thought to have been present in the adhesive between the laminated films, and migrated through the food-contact film of the package. Consumption of potassium permanganate and residue on evaporation of the heated water were ten times of those of the water simulant, while the total organic carbon level of the heated water was several-hold greater than that of the water simulant. In addition, migrant levels per surface area of the pouch were one-fourth of the concentrations per content volume of the pouch. Since compliance with the legal limits is evaluated based on the concentration per surface area, real migration into foods would be underestimated by a factor of another four.

[Structural analysis of unknown dyes detected in hajikami (ginger pickled in vinegar)].

Hajikami (ginger pickled in vinegar) is often used as a relish for grilled fish, and it often contains coloring agents. We detected Rose Bengal (R105) and two unknown dyes in some Hajikami by thin layer chromatography. In this study, we tried to characterize these unknown dyes by HPLC with photodiode array detection (PDA-HPLC), liquid chromatography/mass spectrometry (LC/MS) and nuclear magnetic resonance (NMR). PDA-HPLC analysis showed that the spectra of the unknown dyes resembled that of R105, suggesting the molecular structures of these two dyes are similar to that of R105. Furthermore, LC/MS analysis suggested that the these dyes are R105 in which one or two iodines are replaced by hydrogen. Finally, the two dyes were determined by 1H-NMR and 13C-NMR to be 4,5,6,7-tetrachloro-3',6'-dihydroxy-2',7'-diiodo spiro[isobenzofuran- 1 (3H),9'-[9H]xanthen]-3-one disodium salt and 4,5,6,7-tetrachloro-3',6'-dihydroxy-2',7'-diiodo spiro[isobenzofuran-1(3H),9'[9H]xanthen]-3-one disodium salt.

Asbestos Fibers in Sake.

Asbestos filters have been widely used to clear beverages in many countries including Japan, and despite warnings from the authorities, some Japanese sake companies have continued using such filters until recently. Forty-seven Japanese sake samples were analyzed using a transmission electron microscope to detect contamination with asbestos fibers. The concentrations of fibers found in sake samples ranged from <7.8 × 103 fibers/L (the detection limit) to 1.96 × 108 fibers/L. Ten sake manufacturers provided information as to whether they had used asbestos in the filtration process and the remaining 37 did not (unknown group). Five out of the 10 manufacturers from which information was available had used asbestos (with asbestos group), and the other 5 had not (without asbestos group). The concentrations of fibers in the samples of the "with asbestos" group were more than 7.7 × 106 fibers/L while those in the "without asbestos" group did not exceed 2.8 × 105 fibers/L. The concentrations of fibers in the samples of the "unknown group" ranged from below the detection limits to 1.46 × 108 fibers/L. Some brands of sake showed extremely high concentrations of asbestos fibers, and therefore it may be necessary to investigate the fate of ingested asbestos in the body and its possible carcinogenicity.