Evaluation of a 24-Hour Caffeine Intake Assessment Compared with Urinary Biomarkers of Caffeine Intake among Young Adults in Canada.

BACKGROUND: Caffeine is a widely consumed stimulant, and caffeine-containing products are increasingly available on the market. Few tools are available to capture caffeine intake, particularly among young adults. To estimate caffeine consumption in the previous 24 hours, the 24-Hour Caffeine Intake Recall (CIR-24) was modeled after the Automated Self-Administered 24-Hour Dietary Assessment Tool, using a brand-specific database of caffeine-containing foods, beverages, and supplements. OBJECTIVE: To evaluate the accuracy of the CIR-24 compared with caffeine concentration biomarkers in urine and a caffeinated beverage intake frequency screener (CBQ) designed to assess usual intake among a young adult population in Canada. DESIGN/PARTICIPANTS: In all, 79 young adults, aged 18 to 29 years, provided 24-hour urine samples and completed the CIR-24 and CBQ. MAIN OUTCOME MEASURES: Excretion for caffeine and eight caffeine metabolites were quantified from urine samples using high-performance liquid chromatography-polarity switching electrospray ionization-tandem quadrupole mass spectrometry with stable isotope-labeled internal standards. STATISTICAL ANALYSES PERFORMED: Pearson correlations and weighted κ coefficients were calculated for the self-report tools and caffeine biomarkers. RESULTS: The CIR-24 was significantly positively associated with all caffeine biomarkers (rp=0.28 to 0.52, κ=0.39 to 0.59), and the CBQ was significantly positively associated with all but one biomarker (rp=0.21 to 0.40, κ=0.32 to 0.45). The CIR-24 yielded a higher mean intake of caffeine than the CBQ. There was strong linear correlation between the CIR-24 and CBQ (rp=0.60, P<0.001), but poor agreement in absolute caffeine consumed (t=2.83, P=0.006); quartile ranking concordance was 0.44 (P<0.001). The CIR-24 performed better than the CBQ across all biomarkers in both linear correlation and quartile ranking. CONCLUSIONS: Although both the CIR-24 and CBQ performed reasonably well in capturing caffeine intake compared with urinary biomarkers of caffeine consumption, the CIR-24 had stronger agreement than the CBQ. The results suggest that the CIR-24 is a promising tool for evaluating caffeine intake among this population.

Urine excretion of caffeine and select caffeine metabolites is common in the U.S. population and associated with caffeine intake.

BACKGROUND: Caffeine is a widely consumed psychoactive stimulant and is of epidemiologic interest. Major sources of caffeine are challenging to standardize, and the use of biomarkers is proposed as an alternative means of assessing intake. OBJECTIVE: We described urine caffeine and caffeine metabolite concentrations (n = 2466) and excretion rates (n = 2261) in the US population ≥6 y by age, sex, race-ethnicity, and caffeine intake (from foods, beverages, and dietary supplements). METHODS: We measured caffeine and 14 of its metabolites in spot urine samples from the cross-sectional NHANES 2009-2010 by use of LC-tandem mass spectrometry. RESULTS: Caffeine and its metabolites were detectable in the urine of most persons, generally at concentrations ≥1 µmol/L. Median concentrations (95% CI) ranged from 0.560 (0.497, 0.620) µmol/L to 58.6 (48.6, 67.2) µmol/L; median excretion rates from 0.423 (0.385, 0.468) nmol/min to 46.0 (40.7, 50.2) nmol/min. Urine concentrations and excretion rates for 9 analytes (caffeine, theophylline, paraxanthine, 1-methylxanthine, 1-methyluric acid, 1,3-dimethyluric acid, 1,7-dimethyluric acid, 1,3,7-trimethyluric acid, and 5-acetylamino-6-amino-3-methyluracil) had moderate correlations with caffeine intake (Spearman ρ = 0.55-0.68, P < 0.0001); the remaining analytes had low correlations (ρ = 0.15-0.33, P < 0.0001). We observed larger differences in geometric mean concentrations and excretion rates between the highest vs. lowest quartiles of caffeine intake for these 9 compounds than the rest. Consistent with dietary caffeine intake, we observed that urine concentrations and excretion rates for most compounds were significantly (P < 0.05) higher in men than women, non-Hispanic whites than Hispanics and non-Hispanic blacks, and highest in persons aged 40-59 y. CONCLUSION: Excretion of caffeine and its metabolites in urine is common in the US population. According to the observed associations between spot urine concentrations or excretion rates with caffeine intake, several of these compounds show promise as potential biomarkers of caffeine intake.

Determination of urine caffeine and its metabolites by use of high-performance liquid chromatography-tandem mass spectrometry: estimating dietary caffeine exposure and metabolic phenotyping in population studies.

We have developed and validated a high-performance liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for determining urine caffeine and 14 caffeine metabolites suitable for estimating caffeine exposure and metabolic phenotyping in population studies. Sample preparation consisted solely of a series of simple reagent treatments at room temperature. Stable isotope-labeled analogs were used as internal standards for all analytes. We developed rapid LC-MS/MS separations for both positive and negative ion mode electrospray ionizations to maximize measurement sensitivity. Limits of detection were 0.05-0.1 µmol/L depending on the analytes. Method imprecision, based on total coefficients of variation, was generally <7 % when analyte concentration was >1 µmol/L. Analyte recoveries were typically within 10 % of being quantitative (100 %), and good agreement was observed among analytes measured across different MS/MS transitions. We applied this method to the analysis of a convenience set of human urine samples (n = 115) and were able to detect a majority of the analytes in ≥99 % of samples as well as calculate caffeine metabolite phenotyping ratios for cytochrome P450 1A2 and N-acetyltransferase 2. Whereas existing LC-MS/MS methods are limited in number of caffeine metabolites for which they are validated, or are designed for studies in which purposely elevated caffeine levels are expected, our method is the first of its kind designed specifically for the rapid, sensitive, accurate, and precise measurement of urine caffeine and caffeine metabolites at concentrations relevant to population studies.

Detection and quantitation of subgroup C adenovirus DNA in human tissue samples by real-time PCR.

Advances in amplification techniques have revolutionized the ability to detect viruses both quantitatively and qualitatively and to study viral load. Real-time polymerase chain reaction (PCR) amplification depends on the ability to detect and quantify a fluorescent reporter molecule whose signal increases in proportion to the amount of amplification product generated. Recent advances have been made by using probes, such as TaqMan probes, to detect amplified products. Use of these probes offers confirmation of specificity of the PCR product. Here we describe a sensitive real-time PCR assay to quantify subgroup C adenoviral DNA in human lymphocytes derived from mucosal tissues removed in routine tonsillectomy or adenoidectomy. This chapter will describe in detail the methods used for these analyses.