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Biomedical subjects

B P Lau

Publications and source records attributed to B P Lau.

18 recordsLinked to original sources

Quantitative determination of ochratoxin A by liquid chromatography/electrospray tandem mass spectrometry.

Mass spectrometry of ochratoxin A (OTA) and B (OTB) under electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) was studied. ESI offers higher sensitivities and less fragmentation than APCI. A sensitive LC/MS/MS method for the determination of ochratoxin A (OTA) in human plasma samples was developed. The absolute minimum detection limit was around 10-20 pg per injection, corresponding to 0.5 ppb in an injection equivalent to 20-40microg of human plasma. Ochratoxin B (OTB) was used as an internal standard and its absence in real-life samples was carefully checked before samples were spiked with the internal standard. It was found that these two ochratoxins are susceptible to sodium adduct formation. Fragment ions from the [M + H](+) and [M + Na](+) ions of both OTA and OTB were monitored in the multiple reaction monitoring mode. Three quantitative approaches, standard addition method, internal standard method (using ochratoxin B as an internal standard) and external standard method, were compared in the analysis of human blood plasma. Results from the mass spectrometric method were comparable to those from a conventional LC/fluorescence method. The LC/MS/MS method was also applied to the analysis of contaminated coffee samples.

Atmospheric Pressure↗

Survey of Canadian human blood plasma for ochratoxin A.

Blood plasma samples collected from 144 healthy volunteers in 16 locations across Canada in 1994 were analysed for ochratoxin A (OTA). The method of analysis included cleanup by C18 solid phase extraction and immunoaffinity columns followed by liquid chromatography (LC) with fluorescence detection, which gave 86.5% (s.d. = 9.7) recovery (n = 31) of OTA (added at 2 ng/ml) with a detection limit of 0.15 ng/ml. The arithmetic mean concentration found in the plasma samples, corrected for volume of anticoagulant added, was 0.88 ng/ml with a standard deviation of 0.35 ng/ml and a range of 0.29-2.37 ng/ml. Confirmation of identity of OTA was by methyl ester formation for 65 samples and by LC-tandem mass spectrometry for 17 samples (some of which were included in pooled samples). Statistical analysis, by ANOVA of the log OTA plasma concentrations, showed a highly significant effect due to location in Canada (p = < 0.0001) but no effect due to age, sex or blood group of donors. The highest mean concentration was found in Winnipeg, significantly different (p = 0.05) by the Student-Newman-Keuls multiple range test from the lowest levels found in Toronto, Vancouver and Saint John.

Adult↗

Blood serum levels of PCBs and PCDFs in Yucheng women 14 years after exposure to a toxic rice oil.

In 1979, a mass poisoning of more than 2000 people occurred in central Taiwan due to consumption of rice-bran oil contaminated with PCBs and their heat-degraded byproducts. The incident was later referred to as Yucheng (oil disease). Serum samples from 56 women with the 1979 exposure were collected in February 1992 and analyzed for their contaminant content using sample enrichment and isotope dilution mass spectrometry. In most of the samples, levels of PCDFs and PCBs were detectable, and the median values of 2,3,4,7,8-PCDFs and 1,2,3,4,7,8-PCDFs were 1,030 and 2,220 ng/kg serum lipid, respectively. The median level of the total PCBs on a whole weight basis was 8,730 ng/kg. The PCB/PCDF concentrations in Yucheng women 14 years after the toxic exposure were still one to two orders of magnitude higher than controls. Concentrations of PCB levels in 1992 were positively correlated with the 1980-1981 measured PCB levels in these women and both PCBs and PCDFs were negatively correlated with the total duration when these women breast fed their children between 1979 and 1992. It is concluded that serum levels of congener-specific PCBs/PCDFs in exposed women are good indicators of previous exposure and may provide important information for more reliable estimation of dose-response relationship.

Benzofurans↗

Mutagenic heterocyclic aromatic amines (HAAs) in 'processed food flavour' samples.

Eight samples of 'processed food flavours' (PFFs), chosen from five different categories, were analysed for their mutagenic activity using the Ames Salmonella assay, and also for the presence of eight heterocyclic aromatic amines (HAAs), namely 2-amino-3-(trideuteromethyl)imidazo[4,5-f]quinoline (IQ), 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline (7,8-DiMeIQx), 2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline (4,7,8-TriMeIQx), 2-amino-I-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) using liquid chromatography and mass spectrometry (LC/MS). The isolation of HAAs was based on sequential liquid-liquid extraction procedures of samples at both acidic and basic pH values. The recoveries and the clean-up were monitored by introduction of quality control samples and by spiking with three tri-deuterated standards of HAAs. Although the results for the mutagenicity assay were comparable by testing less-purified and highly-purified extracts, the analysis for identification and quantification of HAAs by LC/MS required highly purified concentrates. Four samples had little or no mutagenic activity and these results were in agreement with their LC/MS results: they had no detectable levels (detection limits 1-3 ppb) of any of the HAAs monitored. The mutagenic activity of one sample was in complete agreement with the quantification of HAAs by LC/MS. Two samples produced strong mutagenic responses (3115 and 2664 revertants/g). In one sample, LC/MS analysis revealed the presence of 9.6 ppb IQ, whereas LC/MS of the other could not confirm the presence of any of the eight HAAs monitored. Two samples produced mild mutagenic activity (204 and 160 revertants/g), but relatively elevated concentrations of IQ (6.7 and 6.8 ng/g) by LC/MS. The extracts from all samples were tested for their modifying effects on mutagenicity of four HAAs. The discrepancy between the Ames test and the LC/MS analysis of some samples indicates several possibilities, such as the presence of some other HAAs, of their isomers or of other mutagens. In addition, the presence of mutagen modifiers (inhibitors or synergists) was observed in most samples. The results indicate that although chemical tests (e.g. LC/MS) can provide quantitative data for the HAAs monitored, the Ames mutagenicity test should also be conducted to determine the mutagenic activities of PFFs, in order to assess their health risk potential.

Amines↗

Heterocyclic aromatic amine content in pre-processed meat cuts produced in Canada.

In an ongoing survey, the presence of heterocyclic aromatic amines (HAAs) was determined in processed, ready-to-eat meat products sold as 'meat cuts'. HAAs are a group of recently recognized mutagenic/carcinogenic contaminants in foods that are produced during the heat processing of meat. 16 samples of meat cuts (e.g. turkey breast, salami, chicken loaf, cooked ham, all beef meat, pepperoni, etc.), randomly purchased from supermarkets and specialty food stores in the Ottawa area, were analysed for the presence of eight HAAs. The isolation of HAAs was based on sequential liquid-liquid extraction procedures of the samples at both acidic and basic pH values. The mutagenic activity of these samples was determined using the Ames/Salmonella microsome assay with the strain TA98 plus rat liver S-9 metabolic activation. The mutagenicity of these samples ranged from undetectable to slightly active. The highest mutagenic activity, 141 induced revertants/g, was found in a smoked turkey breast sample. 11 samples were not mutagenic, including two that indicated a tendency for inhibition of the spontaneous revertants. The remaining four samples exhibited very low mutagenic activity. For chemical analysis, the extracts were purified with two solid phase extraction cartridges. Quantitative analysis was performed by using liquid chromatography for separation and mass spectrometry for detection. With the exception of trace amounts (0.4 ng/g) of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in the sample with highest mutagenic activity, the chemical analysis did not detect the presence of any of the eight most frequently found HAAs in fried or broiled meat products. These data suggest that consumption of meat cuts does not present a serious health risk from HAA-type contaminants.

Amines↗

Comparison of UV absorption and electrospray mass spectrometry for the high-performance liquid chromatographic determination of domoic acid in shellfish and biological samples.

Domoic acid, a neurotoxic amino acid produced by the marine diatom Nitchia pungens multiseries, was determined in samples of anchovies, razor clams, mussels, crab, rat serum, urine and feces by HPLC with UV absorption and electrospray (ESI) mass spectrometric (MS) detection. Shellfish samples were extracted with methanol-water followed by clean-up of the extracts with solid-phase extraction cartridges (strong anion or strong cation exchange). An aliquot of the fraction containing the domoic acid was analysed by HPLC. HPLC column size, mobile phase composition and flow-rate were selected so that essentially the same conditions could be used for both HPLC-UV and HPLC-ESI-MS with selected ion monitoring (SIM) determinations. These included the use of acetonitrile-water-formic acid as the mobile phase, at a flow-rate of 0.2 ml/min (split 13:1 for HPLC-ESI-MS-SIM, 10 microliters/min to the mass spectrometer). The results indicated that extracts found positive by the HPLC-UV method could be readily confirmed directly by HPLC-ESI-MS-SIM without additional sample treatment down to levels of 0.1 micrograms/g of domoic acid. This study demonstrates the use of HPLC-ESI-MS-SIM for the routine confirmation of domoic acid in a wide variety of samples.

Animals↗

Gas chromatographic-mass spectrometric determination of ethyl carbamate in alcoholic beverages.

A sensitive and specific method based on gas chromatography-mass spectrometry for the quantitative determination of ethyl carbamate in table wines, fortified wines (such as ports and sherries), distilled spirits, brandies and liqueurs has been developed. Three characteristic ions from ethyl carbamate [m/z 89 (molecular ion), 74 and 62] were monitored in the selected-ion monitoring (SIM) mode. The lowest detection limit (based on the response on the m/z 62 ion) was estimated to be 0.5 ng/g (ppb). Additional confirmation techniques, including high-resolution SIM, and methane or isobutane chemical ionization are described.

Alcoholic Beverages↗

Identification and confirmation of arsenobetaine and arsenocholine in fish, lobster and shrimp by a combination of fast atom bombardment and tandem mass spectrometry.

Using fast atom bombardment (FAB) ionization combined with tandem mass spectrometry (MS/MS), the identity of arsenobetaine (AB) in extracts of sole, haddock, lobster and shrimp was unequivocally confirmed. Evidence was also presented for the presence of arsenocholine (AC) in the shrimp extract. Confirmation was obtained by comparing the collision-induced dissociation (CID) daughter ion and parent ion spectra of major ions in the FAB spectra from these extracts with those from the authentic standards.

Animals↗

Definitive nonsurgical therapy of epithelial malignancies of the anal canal. A report of 12 cases.

Twelve patients with epidermoid carcinoma of the anal region ranging in age from 40-89 years were treated with combined chemotherapy (CT) and radiotherapy (RT) in lieu of abdominal-perineal resection. Ten patients received no prior treatment and two patients were treated for local recurrence following limited surgical excision. Two courses of 5-FU infusion and mitomycin C were given 3-4 weeks apart simultaneously with whole pelvis RT to 3000-4140 rad. One patient received an additional tumor volume dose of 3094 rad by interstitial Iridium implant and one patient received an orthovoltage boost of 1000 rad to the anal ring. All patients completed treatment. Complete regression of the anal mass occurred in all patients. Biopsies of the anal region performed after completion of therapy revealed no evidence of residual cancer. Ten of the 12 patients are alive without evidence of disease 4-24 months (median, 14 months) after completion of treatment. Two patients have died seven months after treatment of unrelated causes and were tumor-free at autopsy. All patients developed proctitis, diarrhea, and moist perineal desquamation which resolved by four weeks posttreatment. Based on their experience and that of others recently reported, the authors conclude that the described CT-RT protocol offers a definitive alternative to surgery of epidermoid cancer of the anal region.

Adult↗

Analysis of Ginkgo biloba for the presence of ginkgotoxin and ginkgotoxin 5'-glucoside.

Hot water extracts of Ginkgo biloba seeds were analyzed for the presence of ginkgotoxin (4'-O-methylpyridoxine) by reversed-phase liquid chromatography (LC) using methanol-0.05M KH2PO4 (1 + 9, v/v) adjusted to pH 3 as mobile phase. Detection was by fluorescence (excitation 280 nm, emission 370 nm). A straight line calibration curve was obtained for the 10-100 ng injected. After addition of beta-glucosidase (37 degrees C/h), an earlier eluting peak disappeared and the ginkgotoxin peak increased. The identity of the ginkgotoxin was confirmed by LC/MS and LC/MS/MS. LC/MS/MS also confirmed the 5'-glucoside by comparison with the 3-glucoside. This is the first identification of a glucoside of ginkgotoxin in Ginkgo biloba. An unknown compound of MW 267 also observed in the Ginkgo biloba seed extract was shown not to be 3,5'-diacetylginkgotoxin by its different LC retention time. Extraction of ground Ginkgo biloba seeds with boiling water in a Soxhlet for 2 x 2 h yielded a total of 179 microg/g of free ginkgotoxin. The concentration in powder from Ginkgo biloba capsules was several times lower than this (17-64 microg/g) in 3 samples but higher in another (457 microg/g). Canned ginkgo seeds (white nuts) contained no detectable free ginkgotoxin but the glucoside was present. Different extraction times were studied: 0.5 h gave only 52 microg/g free ginkgotoxin in the ginkgo seeds. However, boiling an extract for 4 h showed about 15% loss of ginkgotoxin and its glucoside.

Chromatography, Liquid↗

Comparison of liquid chromatography/mass spectrometry, ELISA, and phosphatase assay for the determination of microcystins in blue-green algae products.

More than 100 samples of blue-green algae products (consisting of Aphanizomenon, Spirulina, and unidentified blue-green algae) in the form of pills, capsules, and powders were collected from retail outlets from across Canada. The samples were extracted with 75% methanol in water and centrifuged to remove solids. Aliquots of the extracts along with spiked blank sample extracts were sent to each participating laboratory and independently analyzed for microcystins by enzyme-linked immunosorbent assay (ELISA), protein phosphatase inhibition assay, and by liquid chromatography-tandem mass spectrometry (LC-MS/MS) after sample cleanup using C18 solid-phase extraction. The results obtained by ELISA and LC-MS/MS agreed very well over a concentration range of about 0.5-35 microg/g. The colorimetric phosphatase results generally agreed with the other 2 methods. While the 2 biochemical assays measured total microcystin content compared with a standard of microcystin LR, the LC-MS/MS method measured specific microcystins (LA, LR, RR, YR) using external standards of these for identification and quantitation. Microcystin LR was found in all positive samples by LC-MS/MS. Microcystin LA was the only other microcystin found in the samples analyzed. These 2 microcystins represent essentially all the microcystins that were present in the extracts. Otherwise, the LC-MS/MS results would have been significantly lower than the results of the biochemical assays had other unknown microcystins been present.

Chromatography, Liquid↗

Capillary column gas chromatographic determination of ethyl carbamate in alcoholic beverages with confirmation by gas chromatography/mass spectrometry.

A method is described for determining ethyl carbamate at low microgram/kg levels in several types of alcoholic beverages by capillary column gas chromatography with Hall electrolytic conductivity detection and confirmation by mass spectrometry. Samples are diluted to obtain a uniform concentration of ethanol (ca 10%) then saturated with NaCl and extracted with methylene chloride. Extracts are evaporated to a small volume and injected in ethyl acetate solution for chromatographic analysis. The method was evaluated by 5 laboratories, 4 employing the Hall detector and one using mass spectrometric detection. Overall between-laboratory mean percent recoveries were: wine, 85.3 +/- 21.0% coefficient of variation (CV) (spiking level 20-45 micrograms/kg); sherry, 83.8 +/- 16.1% CV (spiking level, 81-142 micrograms/kg); whiskey, 79.5 +/- 13.9% CV (spiking level 127-190 micrograms/kg); and brandy, 85.0 +/- 12.5% CV (spiking level 297-446 micrograms/kg). Mass spectrometric results agreed well with the Hall results for all commodities. Detection limits were about 5 micrograms/kg for the Hall detector and about 0.5 microgram/kg for mass spectrometric detection.

Alcoholic Beverages↗