The Prevalence of Dietary Supplements That Claim Estrogen-like Effects in Japanese Women

Recently, adverse events, such as irregular vaginal bleeding and menstrual disorders, associated with the use of dietary supplements containing Pueraria mirifica, have been reported in Japan. P. mirifica contains phytoestrogens, such as deoxymiroestrol and miroestrol. Therefore, we investigated the use of supplements that claim to have estrogen-like effects (i.e., estrogen-like supplements) in Japanese women aged from 15 to 69 years old in an online survey. The prevalence of estrogen-like supplement use was 5%, accounting for approximately 15% of the sample, including ex-users. The majority of the users were in their 40s and 50s, mainly using these supplements for the treatment of menopausal symptoms. In contrast, the younger generation mainly used them for beauty purposes, such as weight loss, mastogenic effects, and skin care. Many of them visited a clinic or took medicines for menstrual-related troubles. In all age groups, soybeans/isoflavones were the most commonly used, followed by equol and placenta. Participants in their teens and 20s also used P. mirifica. Among them, 16.2% had experienced adverse events, including irregular vaginal bleeding, breast swelling and pain, and heavy menstruation. In conclusion, estrogen-like supplement use is associated with adverse events; thus, it is necessary to pay attention to the use of these supplement. Furthermore, because the purpose of use differs depending on generation, caution according to each generation is necessary.


Introduction
Dietary supplements are used for a wide range of purposes other than nutritional supplementation, and the purpose differs depending on the user's characteristics, such as gender, age, and health condition. It has been reported that the prevalence of dietary supplement use is higher in women compared with men [1,2]. Specifically, women are more likely than men to use dietary supplements for weight loss and beauty purposes [3]. Recently, many products targeting women that claim a mastogenic effect via an estrogen-like action are commercially available on the market. One of the major ingredients of these products in Japan is Pueraria mirifica, which contains phytoestrogen, such as deoxymiroestrol and miroestrol. Phytoestrogens are plant-derived compounds that have a similar structure to estradiol and have weak affinity to the estrogen receptor [4]. However, it has been reported that the estrogenic activity of deoxymiroestrol and miroestrol is almost the same as that of 17beta-estradiol [5]; therefore, products containing P. mirifica might possess estrogen-like effects. Indeed, the effects of P. mirifica on the reproductive organs, bones, cardiovascular diseases, and other symptoms related to estrogen deficiency in menopausal women have been studied [6]. In contrast, there have been a series of reports of adverse events related to the use of products containing P. mirifica among young women in Japan, and the National Consumer Affairs Center of Japan cautions consumers on P. mirifica [7,8]. This caution from the National Consumer Affairs Center of Japan is mainly for young women, but some dients of dietary supplements, and the experience of adverse events. The questionnaire in this survey is presented as Supplementary Materials.

Statistical Analysis
Differences in the distribution among age groups were compared using a chi-squared (χ 2 ) test using IBM SPSS Statistics ver. 28.0.1.0 (IBM Corporation, Armonk, NY, USA). A p-value of <0.05 was considered statistically significant. Estrogen-like supplement usage is influenced by consumers' status (age, gestational condition, and menopause condition). These factors were entered into a bivariate logistic regression, and candidate variables were entered into a multivariable logistic regression model. To identify factors of the prevalence of estrogen-like supplement usage, multivariable logistic regression was conducted, and odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated using EZR ver. 4.0.3 (Saitama, Japan), which is a modified version of R commander designed to add statistical functions frequently used in biostatistics.

Characteristics of the Participants
To study estrogen-like supplement users, a preliminary survey was conducted on 61,021 females aged 15-69 years (Table 1). Regarding gestational condition, 2.8% were pregnant and 4.0% were lactating. In terms of menopause, 1.7% were before the first menstruation, 66.4% were premenopausal, and 31.9% were post-menopausal.

The Prevalence of Dietary Supplement Use
The prevalence of dietary supplement use is shown in Table 2. Regarding any type of dietary supplement, 28.9% of the participants were currently using dietary supplements, whereas 18.8% used to use dietary supplements. In terms of estrogen-like supplements, 5.0% of the participants were currently using estrogen-like supplements, accounting for 10.5% of the dietary supplement users. Additionally, 2.8% of the participants had used estrogen-like supplements within the last year, accounting for 5.8% of the dietary supplement users. Then, the 1200 participants who were using or had used estrogen-like supplements progressed to the actual survey (Table 1). The characteristics of the participants in the actual survey were almost the same as those in the preliminary survey.

Factors Associated with Estrogen-like Supplement Use
Estrogen-like supplement use might be associated with one's hormonal condition. Thus, we conducted a logistic regression analysis to identify the factors influencing estrogenlike supplement usage (Table 3). No multicollinearity among the independent variables was suggested by the correlation analysis. In this survey, age, gestational condition, menstruation, menstruation-related problems, dedication for menstruation-related problems, and subjective symptoms suggestive of menopause were identified as significant factors. Among these factors, only menstruation suppressed estrogen-like supplement usage.

Health Conditions among Estrogen-like Supplement Users
Data on health conditions in terms of menstruation are shown in Table 4. Only 19.2% of the premenopausal participants did not have any problem with menstruation, whereas the other 80.8% expressed some problems, such as feeling irritable, depressed, or sleepy before menstruation (46.6%), irregular cycles (42.4%), and severe pain during menstruation (42.3%). In this situation, 11.3% of them were seeing a doctor and 20.8% of them were taking medicine for menstrual-related symptoms (Table 5). In addition, 10.0% of them had subjective symptoms suggestive of menopause, and they were visiting a clinic, whereas 29.2% of them had subjective symptoms suggestive of menopause, but they were not visiting a clinic (Table 6).

The Purpose of Estrogen-like Supplement Use
The purpose of estrogen-like supplement use is shown in Table 7. The main purpose was "Anti-aging (36.8%)", which was higher in older generations compared with younger generations. Other key purposes were "Treatment for menopause symptom (33.5%)", "Skin care (31.0%)", and "Treatment for menstruation-related symptoms (24.5%)", and "Weight loss (20.0%)". Except for "Treatment of disease", there were significant differences among generations. "Weight loss", "Mastogenic effect", "Skin care", and "Treatment for menstruation-related symptoms" were higher in younger generations; on the other hand, "Treatment for menopause symptoms", "Prevention of diseases", and "Anti-aging" were higher in older generations.

The Major Ingredient of Estrogen-like Supplements
The main ingredients of estrogen-like supplements which the participants used are shown in Table 8. The most popular ingredient was soybeans/isoflavones (43.3%). The second most popular ingredient was equol (24.4%), which is a metabolite of daidzein by enterobacterium. The third most popular ingredient was the placenta (22.7%), and the prevalence of these three ingredients was higher in older generations compared with younger generations. The other ingredients comprised less than 10%, although the prevalence of P. mirifica was 5.2% and was higher in the younger generations compared with the older generations.

Statement of Estrogen-like Supplement Use
In terms of frequency, most of the participants used estrogen-like supplements for five to seven days a week (69.6%), followed by three to four days a week (12.2%), and one to two days a week (9.8%) ( Table 9). Regarding duration, most of the participants had used estrogen-like supplements for more than one year (34.3%), followed by one to three months (17.8%) and three to six months (14.3%) ( Table 10).

Information Sources and Ways to Obtain Estrogen-like Supplements
The information sources pertaining to estrogen-like supplements are shown in Table 11. The most popular information source was the Internet (58.5%), then television or radio (26.8%), then family, friends, or acquaintances (19.8%). There were significant differences among generations in information sources, but only SNS showed a clear tendency, in that it was higher in the younger generations and decreased age-dependently. According to information source, most obtained estrogen-like supplements via the Internet (57.5%), followed by a pharmacy or drug store (44.2%) and by mail order (9.9%) ( Table 12). There was a tendency toward more of the younger generations obtaining estrogen-like supplements at physical stores, such as a pharmacy or drug store, clinic, or supermarket/convenience store compared with the older generations.

Adverse Events
Adverse events that might be associated with estrogen-like dietary supplement use are shown in Table 13. The major adverse event was nausea/vomiting (4.7%), followed by diarrhea/constipation (3.6%) and breast swelling and pain (2.6%). We also asked about supplement use after having experienced adverse events. When they had experienced palpitation/shortness of breath, irregular vaginal bleeding, or eczema/itching, more than half of subjects stopped immediately. However, when they had experienced breast swelling and pain, heavy menstruation, and hot flashes, a lot of them kept using the supplements without changing or decreasing the amount or frequency.

Discussion
In 2017, a lot of adverse events associated with use of supplements containing P. mirifica were reported to the National Consumer Affairs Center of Japan [7]. The major adverse events included irregular menstruation and vaginal bleeding, potentially caused by the estrogenic effects of P. mirifica. In addition to P. mirifica, there are several other estrogen-like supplements on the Japanese market, resulting in a risk of adverse events if consumers use them inappropriately. To clarify this issue, we conducted an Internet survey regarding estrogen-like supplement usage. In this study, the prevalence of estrogen-like supplement use was 5%, accounting for approximately 15% of the total sample, including ex-users; the most popular ingredients were soybeans/isoflavones. Among them, 16.3% of the participants had experienced adverse events, including symptoms mainly related to physiological effects peculiar to women, such as irregular vaginal bleeding.
We clarified that the prevalence of estrogen-like supplement usage was associated with age, menstruation and menopause conditions, and related problems. Under-menstruation (premenopausal) was a negative factor of estrogen-like supplement use (OR = 0.75, 95% CI = 0.64-0.86); in other words, menopause was one of the factors for estrogen-like supplement usage. In addition, menstruation-related problems (OR = 1.40, 95% CI = 1.24-1.58) and menopause symptoms (OR = 2.54, 95% CI = 2.34-2.75) were factors of estrogen-like supplement use. Consistently, more than 30% of those aged between their teens and 40s used such supplements for the treatment for menstruation-related symptoms, and more than 50% of those in their 50s-60s used them for the treatment of menopause symptoms. This trend could be explained by the age at onset of menopause. It has been reported that the average age of menopause is 48.3 years old in Japan [27] and 51 years old in the USA [28], and 75% of these women are annoyed by symptoms such as hot flashes and night sweats [28]. Postmenopausal women desire to alleviate these symptoms, such as hot flashes, sleep problems, mood disorders, sexual dysfunction, weight gain, and declines in cognitive functioning, and hormone-replacement therapy is an effective treatment for such problematic symptoms. However, there are potentially undesirable health consequences of hormone-replacement therapy in terms of cardiovascular health and the risk of ovarian and breast cancer [29,30]; thus, a lot of post-menopausal women desire to alleviate these symptoms and seek out complementary and alternative medicine, including phytoestrogens [15,[31][32][33][34].
In addition, numerous participants used estrogen-like supplements for weight loss and mastogenic effects. In the case of weight loss, 20.0% of the participants, especially those of the younger generations (33.8% in their 10s; 34.5% in their 20s), used estrogen-like supplements for this purpose (Table 7). At this time, there is limited evidence of the efficacy of phytoestrogens on body weight. It has been reported that soy products decrease the body weight, BMI, and body fat ratio in overweight or obese Asian populations [35]. In contrast, however, one systematic review showed no association between phytoestrogen supplementation (e.g., soy isoflavones, daidzein, red clover isoflavones, and flaxseed extract) and body weight, body mass index, waist-to-hip circumference, total fat mass, or percentage of body fat in post-menopausal women [36]. In this regard, there is not enough evidence to encourage the use of estrogen-like supplements for this purpose. In the case of mastogenic effect, 15.7% of the participants (40.0% in their 10s; 37.4% in their 20s) used estrogen-like supplements for this purpose. P. mirifica is the most popular ingredient for this purpose in Japan. P. mirifica contains a lot of phytoestrogens; miroestrol and deoxymiroestrol are known to be responsible ingredients due to their estrogenic activity [5]. Even though the estrogenic activities of miroestrol and deoxymiroestrol are almost same as those of 17beta-strradiol, quality control has not been conducted on products containing P. mirifica which claim mastogenic effects. This situation has caused adverse events in many young women. In light of this, the Japanese government has specified P. mirifica as a "Designated Ingredient, etc." [37]. However, various P. mirifica products with attractive claims for women are still on the market, and adverse events are constantly being reported [38].
Soybeans/isoflavones were the most popular ingredients in this survey; there are several studies about the efficacy of soybeans/isoflavones via estrogenic activity on not only menopausal symptoms, but also breast cancer, prostate cancer, cardiovascular diseases, and osteoporosis [39]. In addition, it is reported that isoflavones combination with probiotics improved bone mineral density in postmenopausal women [40]. Prebiotics not only affect the balance of the intestinal flora-this alteration increases the absorption of minerals such as calcium and magnesium, and influences inflammation and the immune system, affecting bone metabolism [41]-but also increase the bioavailability of isoflavones [40]. Probiotics and isoflavones might coordinately prevent osteoporosis. Equol was the second most popular ingredient in this survey. Equol is a metabolite of daidzein which is a soybean isoflavone, and it is produced through intestinal bacterial metabolism [42,43].
There are many reports about the beneficial characteristics of equol products. However, it is also known that approximately 50% of Asians and 25% of non-Asians host the intestinal bacteria that convert daidzein into equol [44]. In addition to soy isoflavones, the market for equol products as supplements has expanded, and studies about equol intake are ongoing.
Beneficial effects of equol intake on hot flashes in postmenopausal women have been reported [45], but there is little scientific evidence of the beneficial effects of equol, other than on menopausal symptoms.
We previously reported that patients use dietary supplements in combination with medicines for the purpose of treating diseases, among other reasons [23,46]. The interaction between dietary supplements and medicines is an important issue among patients. In this survey, 20.8% of the participants were taking medicine for menstruation-related symptoms, and these participants were potentially taking medicines that include estrogen preparations [23,24]. In this situation, pharmacodynamic interactions should be taken into consideration. In contrast, 16.4% of the participants were taking medicine for purposes other than menstruation-related symptoms. In this situation, pharmacokinetic interactions should be considered. It has been reported that soybeans/isoflavones [47], equol [48], black cohosh [49], and other herbal ingredients of dietary supplements have the potential to interact with drugs via the modification of drug metabolism [50,51]; their concomitant use can cause serious adverse events [52]. In this survey, we did not ask what kind of medicines the participants were taking; thus, we cannot speculate on the exact risks to our participants. However, we should consider the possibility that some users of estrogen-like supplements may be deterred from appropriate medical care and appropriate self-care.
The strength of this study is that it is the first report clarifying the perceptions of estrogen-like supplement use. In addition, this survey was conducted on 61,021 participants and 1200 estrogen-like supplement users in each generation. In contrast, a limitation of this study is that it was an online survey; thus, the participants were registrants of the survey company. In this regard, we have to carefully treat our data as general, even though Internet and online questionnaires have become popular across all age groups. In addition, 20.8% of the estrogen supplement users were taking medicine for menstruation-related symptoms, and 16.4% of the estrogen supplement users were taking medicine for reasons other than menstruation-related symptoms. However, we did not ask what kind of medicines they were taking, so we could not examine the risk of the adverse events by concomitant use. In addition, the participants were limited to females only. Although phytoestrogens exert health benefits for not only women, but also men, there is not enough evidence of their efficacy and safety or their potential to cause adverse effects [53]. Recently, the demand for estrogen-like supplements among men [54] and trans people [55] has also increased; thus, we should clarify the situation among these populations.

Conclusions
In this study, the use of supplements with estrogen-like effects was observed in all age groups, and it was clarified that the purpose of use differed according to age group. In addition, health hazards, particularly symptoms attributed to female hormone-like effects, are observed at a certain rate, and the possibility of interactions between estrogen-like supplements and medicines is suspected. Thus, it is necessary to pay attention to the use of these products. The purpose of use differs depending on age; therefore, it is considered necessary to provide information according to each age. Informed Consent Statement: Informed consent was obtained from all of the participants involved in this study.

Data Availability Statement:
The data presented in this study are available upon request from the corresponding author.