Ten healthy Japanese men were recruited in this study. Exclusion criteria were liver or renal dysfunction, alcohol dependence, and receiving treatment for hyperuricemia or gout. Subjects who had alcohol intolerance were also excluded. The clinical and biological characteristics of the subjects are shown in Table 1. The mean values ± standard deviation (SD) of age and body mass index were 25.0 ± 4.5 years and 22.1 ± 2.4 kg/m2, respectively.
The present study was performed after obtaining written informed consent from all subjects and was approved by the Ethics Committee of the University of Shizuoka and registered with UMIN (UMIN registration number: UMIN000040076). The study was performed in accordance with the Helsinki Declaration.
We used a randomized crossover study design. The experiment was conducted so that the test days were separated by a washout period of at least 7 days. All the subjects were asked to avoid heavy exercise and any intake of alcohol and purine-rich foods (>200 mg/100 g) for 3 days prior to each study day. All the subjects were instructed to eat from 20:00 to 21:00 h prior to each test day. After an overnight fast, the subjects were provided with the same prescribed foods (breakfast) at 08:00 h and were required to consume breakfast within 20 min. At first visit, the body weight and height were measured. The subjects were instructed to abstain from foods and beverages other than those prescribed from 08:00 h to 13:00 h. The subjects were provided with their test meals (lunch) at 13:00 h and required to consume each test meal within 20 min. During the experimental period, all subjects were instructed to drink water, 100 mL/h. All subjects underwent a 5 h urine collection from 13:00 to 18:00 h. Venous blood samples were collected at 15:30 h, which was the mid-point of the 5 h urine collection.
Two different test meals were used: a Japanese distilled spirit (Shōchū) with water (SW) and Shōchū with catechin-rich green tea (SC). The SW and the SC each contained 20 g alcohol and were made up to produce a final total volume of 500 mL. We used catechin-rich green tea, which is a commercial beverage, and the SC contained 617 mg of total catechin. All test meals were ingested with 60 g steamed chicken and 10 g sesame dressing (Table 2). Steamed chicken contained about 85 mg/60 g purines and Shōchū did not contain purine according to the guidelines for the management of hyperuricemia and gout in Japan.(19)
Blood and urine analysis methods and anthropometric measurements
Blood samples were centrifuged at 2,400 rpm for 10 min at 4°C and separated into serum and stored at −80°C until analysis of serum creatinine (Cre), UA, Xa/Hx, aspartate aminotransferase, alanine aminotransferase, γ-glutamyl transpeptidase, and blood urea nitrogen concentrations. Urine samples were used for analysis of pH, Cre, UA, and Xa/Hx concentrations. The analyses of serum and urine samples were performed by a blood test company, SRL, Inc. (Tokyo, Japan), except for the analyses of urine pH and Xa/Hx concentrations. The pH was measured using a portable pH meter (LAQUA act, D-71, Horiba Scientific, Kyoto, Japan). The concentration of Xa/Hx was measured using a Xa/Hx colorimetric assay kit (Bio Vision, USA). Anthropometric measurements were determined using a bioelectrical impedance analysis method (innerscan DUAL RD-909, TANITA Corporation, Tokyo, Japan). Height was measured using a stadiometer.
We calculated the creatinine clearance (Ccr), filtered UA load (FUA), UA clearance (CUA), urinary UA excretion per kilogram of body weight per hour (U-UA excretion), and renal fractional UA excretion (FEUA) using the following formulas (U denotes urine, S denotes serum, BSA denotes body surface area, and BW denotes body weight)(19, 20):
Ccr = U-volume × U-Cre / (S-Cre × min) × 1.73 / BSA
FUA = (S-UA / 100) × Ccr
CUA = U-volume × U-UA / (S-UA × min) × 1.73 / BSA
U-UA excretion = U-UA × (U-volume/100) / BW / h
FEUA = (U-UA × S-Cre) / (U-Cre × S-UA) × 100
We also calculated filtered Xa/Hx load (FXa/Hx), Xa/Hx clearance (CXa/Hx), urinary Xa/Hx excretion per kilogram of body weight per hour (U-Xa/Hx excretion), and renal fractional Xa/Hx excretion (FEXa/Hx) using the following formulas:
FXa/Hx = S-Xa/Hx × Ccr
CXa/Hx = U-volume × U-Xa/Hx / (S-Xa/Hx × min) × 1.73 / BSA
U-Xa/Hx excretion = (U-Xa/Hx/1000) × U-volume / BW / h
FEXa/Hx = (U-Xa/Hx × S-Cre) / (U-Cre × S-Xa/Hx) × 100
All data are shown as means ± SD. The Shapiro–Wilk statistic was used for data normality testing. Parametric analysis was used for normal distribution data, and non-parametric analysis was used for data exhibiting a non-normal distribution. Differences between the SW and SC groups were identified using a paired t-test or the Wilcoxon signed-rank test. Probability (P) values less than 0.05 were considered statistically significant in all analyses. Statistical analyses were performed using SPSS for Windows, release 26.0 (SPSS, Chicago, IL).