J Phys Fitness Sports Med, 9 (3): 115-125 (2020) DOI: 10.7600/jpfsm.9.115 JPFSM: Regular Article Effects of black beverage intake on exercise-induced fatigue in untrained healthy adults: a randomized, double-blind, placebo-controlled trial

Shun Inagaki1*, Yoshitake Baba1, Takayuki Ochi2, Yoshie Sakurai2, Takanobu Takihara1 and Yuko M. Sagesaka1

1 Central Research Institute, ITO EN, Ltd., 21 Mekami, Makinohara-shi, Shizuoka 421-0516, Japan 2 1st Production Development Department, ITO EN, Ltd., 21 Mekami, Makinohara-shi, Shizuoka 421-0516, Japan

Received: August 16, 2019 / Accepted: December 24, 2019

Abstract In this study, healthy participants aged 30 to 45 years, who do not exercise regularly, were administered black vinegar beverages (containing ) daily for 7 days and as- sessed for feelings of physical fatigue following exercise. The study was a randomized, double- blind, placebo-controlled crossover study. The evaluation was conducted mainly using the Visual Analog Scale (VAS), which assesses feelings of physical fatigue, to measure levels of serum creatine kinase (CK). Feelings of fatigue were determined using the VAS with an er- gometer before, immediately, and 30 and 60 min after exercise, as well as at bedtime, and the following morning after awaking. Our results showed that 7 days of test beverage (containing 666 mg acetic acid) intake significantly decreased feelings of physical fatigue 30 min follow- ing exercise and at bedtime, compared to intake of the placebo beverage (containing 54.8 mg acetic acid). Furthermore, in the test beverage condition, feelings of shoulder stiffness were significantly decreased at bedtime on the day of exercise and the following morning. However, the test beverage had no significant effects on subjective feelings of fatigue or pain in the leg muscles or levels of serum CK after exercise. Our findings suggested that acetic acid may con- tribute to a more rapid recovery of physical fatigue after exercise. Keywords : acetic acid, black vinegar, endurance exercise, fatigue, stiffness, randomized con- trolled study

on the reasons for the lack of regular exercise, 39.9% of Introduction respondents selected “Too busy with work or home du- The Ministry of Health, Labor, and Welfare of Japan ties” and 55.3% selected “Because it is tiring” as a reason conducted a health awareness survey in 2014 and found for disliking exercise3). This observation suggests that that the most common health concern was decline in fatigue, possibly lasting throughout the day after sporadic physical fitness, accounting for 49.6% of the surveyed or even regular exercise, is a deterrent to more sustained subjects. By age groups, it was the primary concern for exercise habits. Regular exercise is, therefore, considered 46.4% of participants aged between 20 to 39, 50.4% be- challenging to insert within a busy day-to-day schedule, tween 40 to 64, and 51.7% for 64 and over1). This survey and fatigue from daily work is thought to decrease further shows that participants from diverse age groups feel a the will to exercise. Therefore, decreasing the amount of decline in physical fitness regardless of their age. physical fatigue experienced throughout everyday life or Exercise is a healthy lifestyle habit that effectively im- as a result of recreational exercise may make it easier for proves or maintains strength and relieves stress. However, people to introduce regular exercise into their schedules. based on the 2017 Japan National Health and Nutrition Such a change in lifestyle would ultimately lead to the Survey, only 35.9% of men and 28.6% of women exercise maintenance and improvement of health. regularly, with no significant increase in these proportions Brewed vinegar, which is made by the fermentation of over the past 10 years. When numbers are stratified by carbohydrates from grains or fruits, differs depending on age, the proportion of both men and women who regularly the raw ingredients and method of fermentation. is exercise during their working years from 20 to 50 years traditionally used in East Asia as the raw ingredient for old is even lower2). Based on a Japan Sports Agency study black vinegar production. Black vinegar is aged for a longer period than other types of vinegar, and is charac- *Correspondence: [email protected] terized by a rich flavor and lower sourness. In addition 116 JPFSM: Inagaki S, et al. to being used as a , diluted black vinegar has conducted at the Tokyo Skytree Station Medical Clinic recently and increasingly been consumed as a beverage to (Tokyo, Japan) from February 4, 2018 to March 1, 2018 promote health. (UMIN000030886). In Japan, vinegar is traditionally known to have fatigue- relieving effects, and it has been used to alleviate feelings Test beverage and placebo beverage. Each bottle of test of day-to-day fatigue. Acetic acid has also been reported beverage (200 ml) comprised 13.6 g brown rice black vin- to promote fatigue recovery, boost the supplementation of egar produced by Yokoi Vinegar Brewing Co., Ltd. (To- glycogen in muscles and kidneys, which store energy in kyo, Japan) and 185 ml distilled water containing brown the body4), and suppress the accumulation of blood lactic sugar , artificial sweeteners, and flavorings resulting acid, which is a biomarker of intense physical fatigue5). in 666 mg acetic acid per bottle. The placebo beverage Naito et al.6) showed that in both male and female college (200 ml) was prepared using the same product as the test students enrolled in the Faculty of Physical Education, beverage; however, the black vinegar was concentrated consumption of 500 ml of vinegar solution containing under reduced-pressure distillation, which decreases the 1 % acetic acid and glucose led to lower levels of post- concentration of acetic acid (i.e., a low-acetic-acid black exercise serum lactic acid after 1 h of endurance exercise vinegar) to 54.8 mg acetic acid per bottle. The beverages using an ergometer. As described above, to improve ex- were flavored with flavorants to ensure that the test and ercise performance in athletes, the functional evaluation placebo beverages tasted identical. The measured calories of acetic acid has been conducted, focusing on the effect and contents of carbohydrates, proteins, fats, organic ac- of recovering from exhaustingly intense physical activity. ids (8 types), and amino acids (18 types) for both the test On the other hand, few studies have examined the effects and placebo beverages are shown in Table 1. of moderate physical fatigue, wherein blood lactic acid levels do not increase. Furthermore, the effect of acetic acid on trial participants’ subjective symptoms of fatigue Table 1. Nutritional information per serving (200 ml/bottle) has not been fully verified. Therefore, we aimed to com- of test and placebo beverages of black vinegar con- prehensively examine the effects of acetic acid intake on taining acetic acid. fatigue among the general working population who do not exercise regularly and may feel a decline in physi- cal fitness. It has been shown that continuous intake of acetic acid is effective in promoting lipid metabolism7,8), improving sugar metabolism7,9), and increasing blood flow10,11), and it is suggested that intake of acetic acid may promote energy production in the body. We hypothesized that these effects might contribute to the recovery of physical fatigue. Therefore, this study assessed the effects of the daily in- take of a black vinegar beverage containing acetic acid as an active ingredient over 7 days on the feelings of fatigue following moderate exercise in healthy participants who do not exercise regularly.

Methods Ethics committee. This study was conducted in strict accordance with the Declaration of Helsinki (October 2013, Fortaleza revision), Ethical Guidelines for Human Medical Research (2014, Ministry of Education, Culture, Sports, Science, and Technology / Ministry of Health, Labor, and Welfare Notification No. 3), and the Law Concerning the Protection of Personal Information (May 30, 2003, Law No. 57). Prior to commencing the study, the study protocols were examined and approved by the Ethics Committees of Nihonbashi Egawa Clinic (Tokyo, Japan; approval number: food-17121102). The lead phy- Amino acids represent the total value of aspartic acid, glutamic acid, asparagine, serine, histidine, glycine, threonine, arginine, sician of the study explained the details of the study to alanine, gamma-aminobutyric acid, tyrosine, tryptophan, phe- all participants and obtained their voluntary written con- nylalanine, isoleucine, lysine, and proline. ND indicates “not sent before commencement of the study. The study was detected.” JPFSM: Black vinegar reduces physical fatigue after exercise 117

Subjects. Study subjects comprised Japanese men and participants were prohibited from performing strenuous women between the ages of 30 and 45 (at the start of the or prolonged exercise that would rapidly cause shortness study) who had not exercised regularly in the year prior to of breath, overeating, excessive diet restriction, sleep-de- the study. In this study, “lack of regular exercise” was de- privation, excessive exertion, and intake of liquids would fined as less than once per week and less than three times deviate from ordinary daily routine. Participants should per month. Prior to the study, assessments were conducted have completed their last meal at least 6 hours before the to determine the social history, body mass index (BMI), visit; and after that, only water was available until the end vital signs (systolic and diastolic blood pressure, pulse of the test. A flowchart demonstrating the study protocols rate), hematology (white blood cell count, red blood cell is shown in Fig. 1. The evaluation scheme of the clinical count, hemoglobin, hematocrit, mean corpuscular vol- trial is shown in Table 2. ume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration platelet count), and biochemi- Exercise task. Study subjects performed a specific exer- cal blood parameters (triglyceride, total cholesterol, blood cise task on day 7 of test or placebo beverage consump- urea nitrogen, total bilirubin, total protein, albumin, alka- tion, which is shown in Fig. 2. On the day of the exercise line phosphatase, lactate dehydrogenase, gamma-glutamyl task, participants banned eating and drinking 6 hours transpeptidase, aspartate aminotransferase, alanine amino- before the visit. Participants were given two rice balls transferase, creatinine, uric acid, high density lipoprotein– (~360 kcal) about 30 minutes before exercising. During cholesterol, low density lipoprotein–cholesterol, glucose, the task, participants exercised for 50 min maintaining a and hemoglobin A1c) of each study subject. Based on target heart rate equivalent to 60% heart rate reserve (HR the social history assessments, the following participants reserve), (calculated individually based on each partici- were excluded (currently undergoing any medication or pant’s resting heart rate) using an ergometer [STB-3200, outpatient treatment, have a history of liver, kidney, en- STB-3300, or STB-3400, Nihon Kohden Corporation, docrine system, cardiovascular system, gastrointestinal Tokyo, Japan). The exercise task started around noon. tract, lung, blood, or a severe metabolic disease, health The target heart rate was calculated using the Karvonen foods and supplements that are considered to reduce fa- formula as follows: target heart rate = (220 – age – resting tigue such as medicines, nutrition drinks, vitamins, etc., heart rate) × target exercise intensity (%) + resting heart extremely irregular in diet and sleep, suspected insom- rate. A test schedule on day 7 of consumption of the bev- nia, have mental disorders and alcoholism, participate in erage is shown in Fig. 2. other clinical trials within 3 months, irregular work hours such as night shifts). Those with BMI