Boobis et al. 1994 Lynch et al. 1992 Feeding study Feeding study 6 healthy males rece-ived either placebo or 125 mg furafylline 2 h prior to a test meal of fried beef with a known amount of HCA 10 normal, healthy ma-les consu-med a stan-dard cooked meat meal (containing 2.2±0.2 ng MeIQx/g Urine Urine Furafylline on dietary HCA Dietary MelQx/g, DiMelQx/g and PhlP/g MeIQx, PhIP MelQx/g, DiMelQx/g and PhlP/g GC/MS GC/MS Intraindividual coefficients of variation were 28.4% (MeIQx) and 23.7% (PhIP) Interday (intrasubject) variation: Precision was deter-mined by analyzing 6 aliquots of one sub-ject’s urine: coefficients of variation CYP 1A2--catalyzed metabolism accounted for 91% of the elimination of inges-ted MeIQy and 70% of ingested PhIP (most likely via N-hydro-xylation)
136 Appen d ix 2. He te ro c y clic Lynch et al. 1992 — cont. Stillwell et al. 1994 Reistad et al. 1997 Feeding study Feeding study meat, 0.7 0.1 ng DiMeIQx/g meat, and 16.4±2.1 ng PhIP/g meat) on four sepa-rate occa-sions over a period of 14 months 7 subjects consumed pan-fried fish, beef, or bacon 8 subjects consumed fried minced beef patties (295 g con-Urine Urine Food PhlP MelQx and DiMelQx in food MeIQx (free) DiMeIQx, MeIQx, PhIP GC/MS after HPLC separa-tion and acid hydrolysis GC/MS with a novel deriva-tization procedure (Limits pooled coefficients of variation for both compounds = = 19% (MeIQx) and 3.4% (PhIP) MeIQx = 3.2%, PhIP = 9.8% Coefficient of variation < 5%
DiMeIQx was below the limit of detection Table A2.1. HCA — cont.
Study Study design Sample size Tissue /cells Exposure Biomarker Variability of the test intra-individual
Confounders Other source
of variation Laboratory technique Repeatability Dose-response Notes Reliability of the test
137 Kulp et al. 2000 Feeding study 8 volunteers were fed 200 g of cooked
Urine PhlP in food N2
-OH-PhIP-N2 -glu-curonide, PhIP-N2 -glu-LC/MS/MS Variation between injections: Variation activity, whereas total PhIP in urine showed no association to CYP1A2 activity. Linear regression analysis: no association between NAT2 activi-ty and levels of MeIQx or PhIP excret-ed in urine N2-OH-PhIP-N2
-glu-curonide is the most abundant urinary metabolite, followed
138 Appen d ix 2. He te ro c y clic Kulp et al. 2000 — cont. Sinha et al. 1995 Stillwell Feeding study Feeding chicken containing 27 µg PhIP 66 healthy subjects ate lean ground beef cooked at low tem-perature for 7 days; on day 8, sub-jects ate lean ground beef cooked at high tem-perature (9.0 ng MeIQx/g meat) See Sinha Urine Urine Melqx in meat cooked at hight tem-perature Ground curonide, 4'-PhIP-sul-fate and N2 -OH-PhIP--N3-glucuronide MeIQx (free) N-OH-GC/MS NCI-GC-MS within the same sample ranged from 20–30%
Precision of the
Signifi-CYP 1A2 activity was inversely associated with free MeIQx con-centration in urine Excretion by PhIP-N2
-glucuro-nide, Both account for 92–98% of the total metabolite excreted
Table A2.1. HCA — cont.
Study Study design Sample size Tissue /cells Exposure Biomarker Variability of the test intra-individual
Confounders Other source
of variation Laboratory
technique Repeatability Reproducibility
Dose-response
Notes Reliability of the test
139 2002 cooked al low tem-perature and at high tem-perature iduronyl)-2- hydroxy- amino-1- -methyl-6- -phenylimi- dazo(4,5--b]pyridine assessed by duplicate analy-sis of represen-tative samples (n = 12): rela-tive SD between the replicates 5.5–22.2% 1-glucosi- duronyl)-2- -hydroxy- amino-1- -methyl-6- pheny- limida- zo(4,5--b]pyridine in the 0–12 h urine was signifi-cantly related to the quantity of PhIP ingested for all subjects (r = 0.52, P < 0.0001)
140 Appen d ix 2. He te ro c y clic Strick-land et al. 2001 Friesen et al. 2001 Feeding study Feeding study See Sinha et al. 1995 10 healthy non-smoking males con-sumed iden-tical amounts of broiled beef on five con-secutive days Urine Urine Ground beef cooked al low tempera-ture and at high tem-perature Broiled beef PhIP PhIP HPLC with fluorescence detection after immuno-affinity chro-matography GC/MS (Gas chro- matography-negative ion chemical ionization mass spec-trometry). Limit of de-tection 0.1 pmol/ml Coefficient of variation of the assay 7–10% Interassay variability < 16% Significant correlation between the amount of fried meat ingested and urinary PhIP: r = 0.61 (p < < 0.0001) After broiled beef con-sumption, urinary concentra-tion of PhIP increased 14–38 fold above Table A2.1. HCA — cont.
Study Study design Sample size Tissue /cells Exposure Biomarker Variability of the test intra-individual
Confounders Other source
of variation Laboratory
technique Repeatability Reproducibility
Dose-response
Notes Reliability of the test
141 land et al. 2001 Dingley et al. 1998 study Feeding study et al. 2001 Oral admin-istration of a single dose of either 21.3 (n = 5) or 228.0 (n = 2) µg (14C]MeIQx) Blood beef MeIQx MeIQx-albu-min and -Hb adducts fluorescence detection after immuno-affinity chro-matography and GC/MS AMS Dose-dependent levels of MeIQx-albumin and MeIQx-Hb adducts (regres-sion coeffi-cient of the dose-response curve was approx.1)
between the two assays: r = 0.87 (p < 0.0001)
142 Appen d ix 2. He te ro c y clic Mauthe et al. 1999 Dingley et al. 1999 Feeding study Feeding study Oral admi-nistration of a single dose of 21.3 µg (14C]MeIQx (n = 5) before surgery for removal of colon tumors Oral admin-istration of a single dose of 70 µg (n = 2) or 84 µg (n = 3) (14C) PhIP 48– –72 h be-fore surgery for removal Colon Blood, colon MeIQx PhIP MeIQx tis-sue level. MeIQx DNA--adducts PhIP-protein adducts; PhIP tissue level; PhIP DNA--adducts AMS AMS No correla-tion between PhIP levels in tissue and DNA adduct lev-els in that tissue WBC DNA adducts (up to 1.4 adducts/109 nu-cleotides) were unstable & declined substantially over 24 h Table A2.1. HCA — cont.
Study Study design Sample size Tissue /cells Exposure Biomarker Variability of the test intra-individual
Confounders Other source
of variation Laboratory
technique Repeatability Reproducibility
Dose-response
Notes Reliability of the test
143 et al. 2000 Magag-notti et al. 2003 Observation al study different die-tary habits 76 incident colorectal cancer patients Lymphcytes phip Diet likely to contain phip &-globulin adducts PhIP-DNA adducts in lympho-cytes tandem mass spectrometry Liquid chro- mato-graphy-tandem mass spectrometry Sex, smoking, age NAT1, NAT2, SULT1A1, GSTM1, GSTA1 Adduct lev-els were non-signifi-cantly higher in smokers, young subjects and high meat consumers. None of the genetic poly-morphisms studied sig-nificantly affected
144 Appen d ix 2. He te ro c y clic Magag-notti et al. 2003 — cont. PhIP-DNA adducts. High vege-table intake significantly reduced PhIP-DNA adducts (Mann-Whitney U, p = 0.044) Table A2.1. HCA — cont.
Study Study design Sample size Tissue /cells Exposure Biomarker Variability of the test intra-individual
Confounders Other source
of variation Laboratory
technique Repeatability Reproducibility
Dose-response
Notes Reliability of the test
