Chemical Markers for Short- and Long-Term Areca Nut Exposure.

Background: Areca nut (AN) chewing causes oral cancer. AN cessation programs are the most effective approach to reduce AN chewing induced cancers but require biomarkers to determine program compliance and success. Objectives: To explore chemical markers for short- and long-term AN exposure using non-invasively collected saliva, buccal cells (BCs), and scalp hair of chewers. Methods: Saliva was collected from a male chewer before and up to 2 days after AN chewing. Saliva was separated into supernatant and pellet (BCs) then analyzed by spectrophotometry and liquid chromatography (LC) with UV/VIS detection. Scalp hair was collected from four chewers and analyzed for areca alkaloids using direct analysis in real time-tandem mass spectrometry (DART-MSMS). Results: The red pigmented saliva after chewing showed no valuable signals when either the saliva supernatant or pellet (BCs) were analyzed by spectrophotometry. Saliva analysis by LC-UV/VIS showed diagnostically valuable signals at 488 nm up to 5 and 24 h post chewing in the supernatant and pellet, respectively. DART-MSMS analysis detected two of the four AN specific alkaloids (arecoline and arecaidine) in male but none in female hair. Conclusions/Importance: LC-UV/VIS analysis of the red pigments extracted from saliva and BCs after AN chewing showed distinct signals up to 24 h post chewing while DART-MSMS analysis in BCs and scalp hair showed selective signals of AN alkaloids for several weeks or months after AN exposure. Chemical hair treatment might prevent detection of areca alkaloids in hair. AN cessation trials and other programs now have essential tools for bioverification.

Composition of betel specific chemicals in saliva during betel chewing for the identification of biomarkers.

Betel nut chewing causes cancer in humans, including strong associations with head and neck cancer in Guam. In the search for biomarkers of betel chewing we sought to identify chemicals specific for the 3 most commonly consumed betel preparations in Guam: nut ('BN'), nut + Piper betle leaf ('BL'), and betel quid ('BQ') consisting of nut + lime + tobacco + Piper betle leaf. Chemicals were extracted from the chewing material and saliva of subjects chewing these betel preparations. Saliva analysis involved protein precipitation with acetonitrile, dilution with formic acid followed by LCMS analysis. Baseline and chewing saliva levels were compared using t-tests and differences between groups were compared by ANOVA; p < 0.05 indicated significance. Predominant compounds in chewing material were guvacine, arecoline, guvacoline, arecaidine, chavibetol, and nicotine. In chewing saliva we found significant increases from baseline for guvacine (BN, BQ), arecoline (all groups), guvacoline (BN), arecaidine (all groups), nicotine (BQ), and chavibetol (BL, BQ), and significant differences between all groups for total areca-specific alkaloids, total tobacco-specific alkaloids and chavibetol. From this pilot study, we propose the following chemical patterns as biomarkers: areca alkaloids for BN use, areca alkaloids and chavibetol for BL use, and areca alkaloids plus chavibetol and tobacco-specific alkaloids for BQ use.

Pilot study for the establishment of biomarkers for radiation damage after computed tomography in children.

Computed tomography (CT) is an imaging modality that exposes patients to ionizing radiation (IR). We review and report findings from our pilot study evaluating whether blood markers are altered in 17 children undergoing medically indicated CT scans. Blood was drawn before ('pre-CT') and 1 hour after ('post-CT' CT scans. Plasma carotenoids, tocopherols, Q10, ascorbic acid (AA) and uric acid (UA) were analyzed by RP-HPLC with diode-array and electrochemical detection. Dehydroascorbic acid (DHAA) was calculated by subtraction from total AA. Total antioxidant capacity (TAC) was measured using the ORAC assay. Cytokines were quantified using a multiplex immunoassay. γ-H2AX foci were visualized using immunofluorescence. Mean pre- and post-CT changes were compared using t-tests; P-levels < .05 indicated significance. All major plasma lipid soluble antioxidant levels were lower post- vs pre-CT (P < .05) possibly from the scavenging of free radicals formed by CT-induced IR. Average AA levels increased (134%) while DHAA levels were decreased (29%) post-CT, probably due to intracellular recycling of AA from DHAA. TAC levels in lipophilic and hydrophilic extracts were unchanged, suggesting that other antioxidants may have assisted in free radical quenching, which would corroborate their lower concentrations post-CT. Cytokine levels were unchanged and dose-dependent increases in γ-H2AX foci, a measure of double strand DNA breaks, were observed (P = .046, n = 3 children). Our results suggest that CT-derived IR can influence the antioxidant system and may elicit detrimental responses on the cellular level of young children. When possible and if appropriate non-IR based techniques such as ultrasound or magnetic resonance imaging should be used.

In vivo changes in plasma coenzyme Q10, carotenoid, tocopherol, and retinol levels in children after computer tomography.

BACKGROUND: Low dose X-irradiation (IR) from computer tomography (CT) can generate free radicals, which can damage biologically relevant molecules and ultimately lead to cancer. These effects are especially concerning for children owing to their higher radiosensitivity and longer life expectancy than adults. The lipid phase micronutrients (LPM) coenzyme Q10, carotenoids, E vitamers, and vitamin A are potent radical scavengers that can act as intracellular antioxidants. METHODS: We investigated changes in circulating levels of these LPM in 17 children (0.25-6 y) undergoing medically indicated CT scans involving relatively low IR doses. Blood was drawn before and 1h after CT scans and analyzed using HPLC with electrochemical and UV/VIS detection. RESULTS: We found significant decreases (p<0.05) in post-CT plasma levels in several LPM which suggests that these LPM can serve as biodosimeters and may protect against damage from IR during clinical procedures such as CT. The strongest predictors for pre- to post-CT changes for many LPM were their baseline levels. CONCLUSION: Future larger studies are warranted to confirm our findings and to test whether high circulating antioxidant levels protect against IR damage in vivo with an ultimate goal of establishing prophylactic modalities for CT-induced IR damage.

Simultaneous analysis of circulating 25-hydroxy-vitamin D3, 25-hydroxy-vitamin D2, retinol, tocopherols, carotenoids, and oxidized and reduced coenzyme Q10 by high performance liquid chromatography with photo diode-array detection using C18 and C30 columns alone or in combination.

Circulating lipid-phase micronutrients (LPM) such as 25-hydroxylated D vitamers, retinol, tocopherols, carotenoids including their isomers, and coenzyme Q10 play important roles in health maintenance and disease prevention and can serve as useful biomarkers. We developed fast, affordable, and accurate HPLC assays that simultaneously measured all above LPM in a single run using UV/VIS detection at 265nm, 295nm, and 480nm with (1) a C18 column alone; (2) a C30 column alone; or (3) each of these columns connected in series. The C18 column alone could separate all major LPM of interest in less than 17min but insufficiently resolved the lycopene isomers, the 25-hydroxylated D vitamers, lutein from zeaxanthin and ß- from γ-tocopherol. The C30 column alone separated all LPM of interest including many isomeric analytes but failed to resolve the Q10 compounds, which co-eluted with carotenoids. Connecting the C18 and C30 columns in series with a detector after the C30 column and a pressure resistant detector between the columns resulted in ideal resolution and accurate quantitation of all LPM of interest but required software capable of processing the acquired data from both detectors. Connecting the C18 and C30 columns in series with exclusively one detector after the C30 column resulted in carotenoid-Q10 interferences, however, this was remedied by heart-cutting 2D-LC with a 6-port valve between the columns, which resolved all analytes in 42min. Faster run times led to some analytes not being resolved. Many variations of these methods are possible to meet the needs of individual requirements while minimizing sample material and turn-around-times.

Effects of vitamin D3 and calcium supplementation on serum levels of tocopherols, retinol, and specific vitamin D metabolites.

γ-Tocopherol (γT) protects against DNA-damaging effects of nitrogen oxides, yet its physiologic regulation in vivo is unknown. Observational studies indicate inverse associations of 25[OH]-vitamin D with γT and leptin. To determine whether vitamin D(3) supplementation alters levels of lipid-soluble micronutrients, serum samples (N = 85 subjects) from a randomized, double-blind, placebo-controlled clinical trial of vitamin D(3) (800 IU) and calcium (2 g), alone and in combination, were analyzed for lipid micronutrients and specific vitamin D metabolites at baseline and after 6 mo of supplementation. Serum 25[OH]-vitamin D(3) levels increased 55% (P < 0.0001) and 48% (P = 0.0005), whereas 25[OH]-vitamin D(2) levels were lower by 48% (P = 0.26) and 21% (P = 0.36) in the vitamin D(3) and vitamin D(3) plus calcium groups, respectively. At baseline, γT levels were inversely associated with 25[OH]D (r = -0.31, P = 0.004). With vitamin D(3) plus calcium treatment, serum α-tocopherol decreased 14% (P = 0.04), whereas similar changes in γT (19% lower, P = 0.14) were observed. No significant effects were observed for D(3) supplementation on leptin or retinol levels. These results are consistent with the hypothesis that vitamin D(3) ± calcium affects serum tocopherol and 25[OH]D(2) levels; however, studies using larger, more homogeneous populations are warranted.

Analysis of urinary estrogens, their oxidized metabolites, and other endogenous steroids by benchtop orbitrap LCMS versus traditional quadrupole GCMS.

Estrogens and other endogenous steroids are known risk markers for cancer. Gas chromatography (GC) with mass spectrometry (MS) has traditionally predominated the analysis of estrogens and other endogenous steroids, but liquid chromatography (LC) MS is increasingly favored. Direct comparisons of the two technologies have hitherto not been performed. Steroids were analyzed from 232 urine samples of 78 premenopausal women in a blinded fashion by benchtop orbitrap LCMS and single quadrupole GCMS. Sixteen steroidal estrogens including oxidized metabolites could be analyzed by LCMS. LCMS-GCMS Spearman rank correlations of the major estrogens E(1), E(2), E(3), 16α-OHE(1), and 2-OHE(1) were very high (r = 0.72-0.91), and absolute concentrations also agreed (<5% difference for E(1), E(2), E(3), 16α-OHE(1)). LCMS allowed reinterrogation of the acquired data due to orbitrap technology, which permitted post-analysis quantitation of progesterone, cortisol, and cortisone (LCMS-GCMS Spearman rank correlations = 0.80-0.84; absolute difference, <7%; n = 137). GCMS allows the measurement of a wide range of steroids including non-polar analytes that escape the presented LCMS assay. In contrast, orbitrap-based LCMS can detect more estrogens, is faster, less costly, allows post-data acquisition reinterrogation of certain analytes that had not been targeted a priori, and requires much less urine.

Coenzyme Q10 in human blood: native levels and determinants of oxidation during processing and storage.

Coenzyme Q10 (Q10) is present in the circulation mainly in its reduced form (ubiquinol-10; UL10), but oxidizes quickly ex vivo to ubiquinone-10 (UN10). Therefore, native UL10:UN10 ratios, used as markers of redox status and disease risk, are difficult to measure. We established an RP-(U)HPLC method with coulometric detection to measure natively circulating UL10 and UN10 concentrations by adding a ubiquinol/ubiquinone mixture as an internal standard immediately after plasma preparation. This allowed adjustment for unavoidable artificial UL10 oxidation as well as for total losses (or gains) of analytes during sample storage, processing, and analysis because the internal standards exactly paralleled the chemical behavior of Q10. This technique applied to blood (n = 13) revealed Q10 levels of 680-3300 nM with a mean UL10:UN10 ratio of 95:5, which was inversely associated with total Q10 (r=-0.69; p=0.004). The oxidation of UL10 to UN10 was equimolar, increased by O(2), and decreased by lower temperatures or various degassing methods. Although UL10 was stable in blood or when pure in organic solvents at 22 degrees C, its oxidation was catalyzed dose dependently by alpha-tocopherol and butylated hydroxytoluene, particularly when present in combination. Key structural features for the catalytic pro-oxidant properties of phenolic antioxidants included two substituents vicinal to the phenolic hydroxyl group.

Phytoestrogenic isoflavonoids in epidemiologic and clinical research.

Isoflavones (IFLs) are natural products to which humans have been traditionally exposed predominantly through soy foods; more recently humans are also exposed to them through soy protein addition to processed foods or through supplements. They are structurally similar to steroidal estrogens and can exert estrogenic or antiestrogenic effects depending on their concentrations and on the tissue considered. These properties qualify IFLs to be classified as phytoestrogens and are believed to account for many of the biological effects observed for soy and/or IFL exposure including benefits for bone and heart health or prevention of menopausal symptoms and certain types of cancer. In order to evaluate the function of IFLs, alone or when exposure happens through soy intake, pharmacokinetics and bioavailability are critical issues to be considered in epidemiologic and clinical research. For this purpose precise, accurate, robust, fast, and affordable techniques for IFL analyses are required.

Tocopherol and tocotrienol levels of foods consumed in Hawaii.

Because of the individual biological effects and the uncertain or missing information on levels of tocopherols (T) and tocotrienols (T3) in foods frequently consumed in Hawaii, 79 food items (50 in duplicate) were analyzed for alpha-, beta-, gamma-, and delta-tocopherol (alphaT, betaT, gammaT, and deltaT) and alpha-, beta-, gamma-, and delta-tocotrienol (alphaT3, betaT3, gammaT3, and deltaT3) in addition to alpha-tocopheryl acetate (alphaTac). Foods from local markets were stored according to usual household habits, freeze-dried, homogenized, and extracted three times with hexane containing butylated hydroxytoluene as a preservative and tocol as an internal standard. A normal-phase high-pressure liquid chromatography system was applied with fluorescence and photodiode array detection that resulted in baseline separation of all eight analytes and the internal standard tocol (To). The sum of all E vitamer concentrations, or total E vitamers (TEV), in all foods analyzed ranged an average from 0.6 to 828 mg/kg (T < or = 542 mg/kg and T3 < or = 432 mg/kg) and showed the following ranges: oils, 497-828 mg/kg (mainly alphaT and gammaT); margarines, 359-457 mg/kg (mainly gammaT); salad dressings, 20-291 mg/kg (mainly gammaT, except alphaT when soy oil was the main ingredient); cookies, 54-138 mg/kg (mainly gammaT); snacks, 101-220 mg/kg (mainly gammaT); nuts, 22-201 mg/kg (mainly alphaT); vegetables, 2-152 mg/kg (mainly alphaT); pasta, 24-90 mg/kg; cereals, 4-56 mg/kg (mainly betaT3 followed by alphaT); fish, 2-39 mg/kg (mainly alphaT); fried tofu, 64 mg/kg (mainly gammaT); breads, 20-22 mg/kg (mainly betaT3); fat-free mayonnaise, 5 mg/kg (mainly alphaT); poi (fermented taro root), 2 mg/kg (mostly alphaT); and fruits, 2 (papaya) to 13 mg/kg (canned pumpkin) with alphaT predominating. Cereals fortified with alphaTac ranked third and eighth among all foods assayed regarding alphaT and TEV levels, respectively. As compared to the few data available in the literature, our values agreed with some (corn flakes, mango fruit, fat-free mayonnaise, dry-roasted macadamia nuts, dry-roasted peanuts, mixed nuts, spaghetti/marinara pasta sauce, oils, and red bell pepper) but differed for many other items. Our results provide new information on the E vitamer content in foods, emphasize the vast differences of bioactivities of individual E vitamers, and confirm the need for analyses of foods consumed in specific study populations.

Isoflavones in breastfed infants after mothers consume soy.

BACKGROUND: The bioavailability of isoflavones in children after soy exposure is uncertain. OBJECTIVE: We aimed to compare isoflavone patterns in infants exposed to isoflavone-containing breast milk (BF), in tofu-fed (TF) infants, and in mothers consuming a soy beverage. DESIGN: Eighteen nursing mothers who were not feeding soy foods to their infants consumed one daily serving of a soy protein beverage for 2-4 d and collected their own milk and urine and infant urine. Plasma was collected from infants if venous blood draws were ordered by pediatricians. Blood and urine were collected from additional children after they consumed tofu. Isoflavones were measured by liquid chromatography-mass spectrometry. RESULTS: In 7 subjects, isoflavone values increased significantly from baseline after mothers ate soy: in maternal urine (x +/- SEM) from 18.4 +/- 13.0 to 135.1 +/- 26.0 nmol/mg creatinine, in breast milk from 5.1 +/- 2.2 to 70.7 +/- 19.2 nmol/L, and in infant urine from 29.8 +/- 11.6 to 111.6 +/- 18.9 nmol/mg creatinine. The mean isoflavone concentration in plasma obtained from 11 BF infants was 19.7 +/- 13.2 nmol/L. TF infants had much higher mean isoflavone values (urine, 229 +/- 129 nmol/mg creatinine; plasma, 1049 +/- 403 nmol/L). Statistically significant correlations were observed between the types of fluids investigated within mothers, between mothers and infants, and within infants. Urinary isoflavone excretion per hour adjusted for dose per body weight was 81% lower for BF infants and 24% higher for TF infants than for their mothers after eating soy. CONCLUSIONS: More isoflavones appear in children than in adults after adjustment for isoflavone intake. Systemic isoflavone exposure in infants can be determined by urinary analysis.

Bioavailability and antioxidant effects of orange juice components in humans.

Seven healthy females and six males consumed daily 256 mg of vitamin C, 271 mg of flavanones (mainly as glycosides), 6 mg of carotenoids (mainly xanthophylls and cryptoxanthins), and 0.16 mg of folate by incorporation of daily three times 236 mL of not from concentrate orange juice (OJ) into their habitual diet. At the end of 3 weeks, mean vitamin C, folate, carotenoid, and flavanone plasma concentrations increased significantly relative to baseline by 59% (p < 0.001), 46% (p = 0.018), and 22% (p < 0.001), and 8-fold (p = 0.045), respectively. Flavanones were excreted in urine 9-fold more at the end of the intervention (p = 0.01) but returned to baseline 2 days after study completion. After the 3 week intervention, plasma concentrations of vitamins A and E did not change. 8-Hydroxydeoxyguanosine in white blood cells declined by 16% (p = 0.38; n = 11), and in individuals with high baseline concentrations by 29% (p = 0.36; n = 7), respectively. Low-density lipoprotein (LDL)-/high-density lipoprotein (HDL)-cholesterol ratios decreased but cholesterol (HDL, LDL, total) and thiobarbituric acid reactive substance plasma concentrations did not change significantly. We conclude from this pilot study that OJ is an excellent food source to enhance circulating concentrations of valuable hydrophilic as well as lipophilic phytochemicals.

Determinants for urinary and plasma isoflavones in humans after soy intake.

Consumption of soy foods leads to a biphasic appearance pattern of isoflavones (IFLs) in blood and urine, with peaks appearing at 1-2 h and 4-8 h after intake, but its causes are not understood. IFLs were measured repeatedly from plasma and/or urine after intake of soy foods, IFL glucosides, or aglycons without or with a mildly or radically reduced gut flora as a result of oral antibiotic (AB) treatment, or this combined with mechanical bowel preparation (AB+MBP). The typical biphasic IFL pattern in blood and/or urine was observed when a soy protein drink without (control) or with AB treatment or when IFL glucosides or aglycons were consumed. Soy intake combined with AB+MBP or consumption of puerarin led to a shift of the second peak to much later times. The first peak was absent after puerarin intake. Total urinary IFL recovery was more than 50% lower in the first 24 h, but overall 61% higher after AB+MBP vs. the control. When the area under the curves for corresponding time intervals were compared, individual or total urinary IFL excretion rates were highly correlated with individual or total plasma IFL levels (r=0.85-0.91; P <0.001). At the same urinary excretion rate three times more genistein than daidzein remained in the circulation. We conclude that urinary IFL excretion rates reflect circulating IFL levels, with daidzein appearing less in blood and more in urine than genistein. The first and second IFL peaks are due to uptake in the small and large intestine, respectively. The latter is the major locus of uptake (90%) at usual dietary IFL doses (0.15-1.5 mmol/kg body weight). A reduced gut flora delayed IFL uptake but led overall to increased urinary recovery because of less bacterial degradation in the intestine.

Serum isoflavones and soya food intake in Japanese, Thai and American end-stage renal disease patients on chronic haemodialysis.

BACKGROUND: Soya foods, a staple in several Asian countries, have received increasing attention because of their nutritional properties and their high isoflavone content. We have shown recently abnormal pharmacokinetics of soya isoflavones following acute oral intake, in soya-naive end-stage renal disease (ESRD) patients. No information is available, however, about blood levels of soya isoflavones in ESRD patients with habitual soya intake. Additionally, no information is available about the conjugation profile of these compounds in ESRD patients. METHODS: To assess the relationship between habitual soya intake on blood isoflavone levels in ESRD patients, we recorded dietary soya food intake and analysed circulating levels of soya isoflavones in randomly selected, clinically stable haemodialysis patients from the United States (n = 20), Thailand (n = 17) and Japan (n = 20). Dietary records and three weekly blood samples were collected from each participant. Combined isoflavones and individual genistein, daidzein, glycitein and O-desmethylangolensin (DMA) were analysed in serum by liquid chromatography/mass spectrometry. Lipid phase micronutrients, including tocopherols, carotenoids and retinol were also measured to compare ethnic differences in isoflavones with those of more common lipid soluble antioxidant micronutrients. RESULTS: Soya intake was higher in Japanese than in Thai patients and it was negligible in the US patients. Blood levels of genistein were very elevated and significantly higher in the Japanese patients (1128 +/- 205 nM), as compared with the Thai and US patients (258 +/- 64 and 168 +/- 49 nM, respectively; P < 0.001). The other isoflavones followed the same trend. Daidzein was more concentrated than genistein in the dialysis patients. Robust correlation was present between weekly soya intake and blood isoflavone levels (r = 0.56, P < 0.001). Despite very high total isoflavone concentrations, the levels of unconjugated and sulphated isoflavones in the Japanese patients were comparable to those described in healthy subjects. Compared with the striking difference in isoflavones, more easily accessible dietary antioxidants, including tocopherols, carotenoids and retinol, differed only minimally or not at all in the three groups. CONCLUSIONS: ESRD patients appear to accumulate isoflavones as a function of dietary soya intake, resulting in blood concentrations that are higher than those reported in subjects with preserved kidney function. Even in the presence of very elevated total isoflavone levels, the concentrations of the unconjugated and sulphated fractions are comparable to those of healthy subjects. A discrepancy is noted between accumulation of soya isoflavones and other more common lipid-soluble antioxidant micronutrients.

Urinary phytoestrogen excretion and breast cancer risk: evaluating potential effect modifiers endogenous estrogens and anthropometrics.

Steroid sex hormones play a central role in breast carcinogenesis. Evidence from in vitro and animal studies suggests that phytoestrogens may inhibit the development of mammary tumors through their role in regulating the synthesis, metabolism, and signal transduction of steroid hormones. In a study of 117 case-control pairs of postmenopausal women in Shanghai, we investigated whether the association between urinary phytoestrogen excretion and breast cancer risk may differ by levels of endogenous steroid sex hormones, sex hormone binding globulin (SHBG), body mass index (BMI), and waist:hip ratio (WHR). Fasting morning blood and urine samples were collected for the analysis of urinary isoflavonoids and mammalian lignans, as well as blood levels of SHBG and selected steroid hormones. For cancer patients, samples were collected before any cancer therapy. Conditional logistic regression models were used to estimate odds ratios and 95% confidence intervals after adjusting for potential confounding factors. The inverse associations between urinary phytoestrogens and breast cancer risk were found to be more evident among women with a high BMI or WHR than those with a low level of these anthropometric measurements. Although a reduced risk of breast cancer was observed among women with a high excretion rate of urinary isoflavonoids in all of the strata defined by blood SHBG and steroid hormones, the inverse association was more pronounced among women with a high blood concentration of estradiol, a low level of estrone sulfate, or a low level of SHBG. The risks of breast cancer were also reduced with increasing excretion rate of mammalian lignans, although no test for a linear association was statistically significant in stratified analyses. Findings from this study suggest that the potential protective association of phytoestrogens may be modified by BMI, WHR, and blood levels of SHBG, and steroid hormones.

Urinary excretion of phytoestrogens and risk of breast cancer among Chinese women in Shanghai.

Although the majority of ecological and experimental studies have suggested a potential role of phytoestrogens in breast cancer prevention, findings from epidemiological studies have been inconsistent. Part of the inconsistencies may be attributable to the difficulty in measuring intake levels of phytoestrogens. Overnight urine samples from 250 incident breast cancer cases and their individually matched controls were analyzed for urinary excretion rates of isoflavonoids, mammalian lignans, and citrus flavonoids. The study subjects were a subset of the participants in the Shanghai Breast Cancer Study, a large population-based case-control study conducted in Shanghai from 1996-1998. To minimize potential influence of treatment on the exposure of interest, urine samples from breast cancer cases were collected before cancer therapy. Urinary excretion of total isoflavonoids and mammalian lignans was substantially lower in breast cancer cases than in controls. The median excretion rate of total isoflavonoids was 13.97 nmol/mg creatinine in cases and 23.09 in controls (P = 0.01), and the median excretion rate of total lignans was 1.77 in cases and 4.16 in controls (P < 0.01). The risk of breast cancer was reduced with increasing excretion of total isoflavonoids (P for trend, 0.04) and total lignans (P for trend, <0.01), with adjusted odds ratios of 0.62 (95% confidence interval, 0.39-0.99) and 0.40 (95% confidence interval, 0.24-0.64) observed for the highest versus the lowest tertile of total isoflavonoid and lignan excretion, respectively. The adjusted odds ratio was 0.28 (95% confidence interval, 0.15-0.50) for women who had a high excretion rate of both total lignans and isoflavonoids compared with those with a low excretion of both groups of phytoestrogens. No association was observed with citrus flavonoids. The results from this study suggest that high intake of certain phytoestrogens may reduce the risk of breast cancer.

Well-done red meat, metabolic phenotypes and colorectal cancer in Hawaii.

Heterocyclic amines (HAAs) and polycyclic hydrocarbons are suspected colorectal cancer (CRC) carcinogens that are found in well-done meat. They require metabolic activation by phase I enzymes, such as the smoking-inducible CYP1A isoenzymes. N-acetyltransferase 2 (NAT2) also play a role in the further activation of HAAs. We conducted a population-based case-control study in Hawaii to test the associations of preference for well-done red meat and HAA intake with colon and rectal cancers, as well as the modifying effects of NAT2 and CYP1A2. We interviewed 727 Japanese, Caucasian or Native Hawaiian cases and 727 controls matched on sex, age, and ethnicity. HAA intake was estimated based on consumption of meat and fish for each of several cooking methods and doneness levels. A subgroup of 349 cases and 467 controls was phenotyped for CYP1A2 by a caffeine test. We found that preference for well-done red meat was associated with a 8.8-fold increased risk of CRC (95% CI: 1.7-44.9) among ever-smokers with the NAT2 and CYP1A2 rapid phenotypes, compared to ever-smokers with low NAT2 and CYP1A2 activities and who preferred their red meat rare or medium. A dose-dependent association was also found between the HAA intake estimates and male rectal cancer, with a two- to three-fold increase in risk from the low (T(1)) to high (T(3)) tertile of intake for each HAA. This association was strongest for MeIQx. HAA intake was not associated with male colon cancer or colon or rectal cancer in women. These data provide support to the hypothesis that exposure to pyrolysis products through consumption of well-done meat increases the risk of CRC, particularly in individuals who smoke and are genetically susceptible (as determined by a rapid phenotype for both NAT2 and CYP1A2). An attempt to examine the risk associated with specific HAAs suggested that the main HAAs increase risk of rectal cancer in men and that they do not appreciably affect risk of rectal cancer in women or of colon cancer in either sex.

Liquid chromatographic-photodiode array mass spectrometric analysis of dietary phytoestrogens from human urine and blood.

Dietary phytoestrogens have been implicated in the prevention of chronic diseases. However, it is uncertain whether the phytoestrogens or the foods associated with phytoestrogens account for the observed effects. We report here a new liquid chromatography photodiode array mass spectrometry (LC-PDA-MS) assay for the determination of nanomolar amounts of the most prominent dietary phytoestrogens (genistein, dihydrogenistein, daidzein, dihydrodaidzein, glycitein, O-desmethylangolensin, hesperetin, naringenin, quercetin, enterodiol, enterolactone) in human plasma or serum and urine. This assay was found to be suitable for the assessment of quercetin exposure in an onion intervention study by measuring urinary quercetin levels. Other successful applications of this assay in clinical and epidemiologic studies validated the developed method and confirmed previous results on the negative association between urinary isoflavone excretion and breast cancer risk.