Determination of the food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human hair by solid-phase extraction and gas chromatography-mass spectrometry
ABSTRACT A method for the determination of the food mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in human hair has been developed and validated. Hair samples (200 mg) were dissolved in NaOH (1 M) and PhIP
was isolated by successive solid-phase extraction on a polystyrene-divinylbenzene column and on a silica-based mixed-mode
column with C8 and-SO3
− functional groups. Quantification was performed by gas chromatography-electron-impact ionization high-resolution mass spectrometry
in selected-ion-monitoring mode. The method was validated for determination of PhIP in the concentration range 0.5–25 ng g−1 hair with [2H3]PhIP as internal standard. The limit of quantification was 0.26 ng g−1 hair. Within-day and between-day precision were in the ranges 1–27% and 2–15% relative standard deviation, respectively.
The hair sample used for method validation was found to contain 0.26 ng PhIP g−1 hair.
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ABSTRACT: : The purpose of this work was to correlate the amount of the food mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) determined in mouse fur with the number of intestinal tumours induced by PhIP, to further evaluate incorporation of PhIP into hair as a putative exposure biomarker for food mutagens. We have previously shown that PhIP increases intestinal tumourigenesis in C57BL/6J-Min/+ (Multiple Intestinal Neoplasia) mice. Fur was sampled from mice exposed according to various PhIP protocols, and the amount of PhIP in the fur was quantitated by high performance liquid chromatography – mass spectrometry (HPLC-MS). A quantitative incorporation of PhIP in the fur was demonstrated after a single subcutaneous injection of PhIP, and between one and eight PhIP exposures either via direct subcutaneous injections or through breast milk from PhIP-injected dams. However, after higher exposures, the amount of PhIP in the fur appeared to reach saturation. After low exposures to PhIP, the number of intestinal tumours correlated with the amount of PhIP in the fur of individual mice, whereas after higher exposures, the number of tumours was relatively higher than the amounts of incorporated PhIP in the fur. Other factors, e.g. amounts of reactive PhIP metabolites formed, are also determining the number of intestinal tumours. The demonstrated quantitative incorporation of PhIP into mice fur and the correlation with number of intestinal tumours at the lower exposures, indicate that determination of PhIP in human hair may be a suitable biomarker for exposure to dietary PhIP, which is found in human hair in 10−3 lower amounts than in these mice.Pharmacology & Toxicology 01/2003; 92(3):131-136.
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ABSTRACT: A simple and easy-to-use extraction procedure has been optimised, validated, and applied for extraction of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in urine and spiked plasma samples. PhIP is a carcinogenic and mutagenic heterocyclic aromatic amine that is formed during cooking of meat and fish. The novelty of the extraction procedure lies in using a short piece of narrow capillary-like microporous hollow-fibre (HF) membrane as extraction device. The HF membrane was filled with a few microlitres of acidic solution and the membrane pores were impregnated with an organic extraction solvent. Therefore, the technique was called hollow-fibre supported liquid membrane (HF-SLM) extraction. The HF extraction device was then supported by a syringe needle and directly immersed in urine (1.4 mL) or plasma (0.3 mL) previously made alkaline by adding 0.5 mol L(-1) NaOH solution to give a final volume of 1.6 mL. The operation of the HF-SLM extraction at the optimal conditions resulted in a PhIP extraction efficiency of 74% from both spiked urine and plasma, corresponding to enrichment factors of 126 and 27, respectively. For 90 min extraction time, limits of detection and quantification were, respectively, 8 and 25 pg mL(-1) for urine and 6 and 11 pg mL(-1) for plasma. Within-day repeatability (n = 6) and between-day reproducibility (n = 3) were, respectively, 5% and 13% for urine and 6% and 7% for plasma. Analysis of urine samples collected for 12 h after a volunteer had eaten 250 g well-done chicken showed the PhIP concentration was 124 +/- 21 pg mL(-1), calculated assuming an extraction efficiency of 74%.Analytical and Bioanalytical Chemistry 02/2008; 390(2):689-96. · 3.66 Impact Factor
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ABSTRACT: Various methods of exposure assessment, such as questionnaires, sometimes combined with pictures of cooked meat, have been employed in investigations on the relationship between heterocyclic amines (HA) and health effects. However, as the content of heterocyclic amines vary greatly with cooking conditions, it is difficult to obtain an accurate estimate of the exposure. To improve the exposure assessment, the use of biomarkers has been investigated. The metabolism of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is well characterised. In humans, the major part of the dose is excreted in urine within 24-48 h following a meal. A few percent is excreted as parent compounds, whereas the major part is metabolites. Urinary level of parent HA reflects only recent exposure. However, the pattern of excreted metabolites might indicate the capacity to activate or detoxify HAs. The excretion of glucuronide conjugates of N-hydroxy-PhIP and N-hydroxy-MeIQx could be a marker for the N-hydroxylation capacity and the dose of the proximate metabolites. Recently, we proposed 5-OH-PhIP as a marker for the ultimate reactive metabolite of PhIP, since it is formed from this compound as a by-product along with the formation of PhIP-DNA adducts. In a search for biomarkers reflecting exposure over some time, blood protein adducts with a longer lifespan have been investigated, and PhIP adducts of serum albumin and haemoglobin from meat-consuming humans were recently reported. Many compounds, like drugs, nicotine and narcotics, bind to melanin in hair and give information on exposure for longer time periods. In mice, PhIP is irreversibly incorporated in a dose-dependent manner into hair, and in humans exposed to an ordinary diet, it was found to vary from <50 to 5000 pg PhIP/g hair. The incorporation is also dependent on the content of eumelanin. The use of PhIP in hair as a biomarker of exposure is promising, but needs validation, using other methods of exposure assessment.Food and Chemical Toxicology 08/2002; 40(8):1131-7. · 3.01 Impact Factor