Tribulus terrestris (protodioscin) in the treatment of male infertility with idiopathic oligoasthenoterato-zoospermia A Adimoelja, L Setiawan, and T Djojotananjo Academic Hospital Dr. Soetomo and Airlangga University, Surabaya, Indonesia in First International Conference of Medical Plants for Reproductive Medicine in Taipei, Taiwan (1995) SUMMARY Tribulus terrestris L (TTL) extract is a natural remedy that has been successfully tested clinically for its use in the treatment of idiopathic oligoasthenoteratozoospermia (OTA). Here, we conducted a double blind study of 45 infertile, married men with OTA. 36 men were treated with 500 mg purified TTL (Libilov) 3 times daily for 3 months. Composing the controlled group, 9 men were given placebo (sugar pills) for the same period of time. Spouses of 8 of the men in the treated group successfully achieved pregnancies after treatment of their husbands, whereas no pregnancies occurred in the spouses of the men in the control group. An improvement in the sperm morphology, including that observed under conventional phase contrast microscopy as well as improvements in acrosome morphology and reaction, seemed to account for the increased fertility after treatment. In addition, TTL was also shown to increase the level of dehydroepiandrosterone (DHEA) and might also contributed to the activation of cell membrane receptors and the production of weak androgens. These, in turn, might additionally improve fertility by increasing the level of spermatogenesis. ABSTRACT Recently, herbal remedies have became popular in many Eastern as well as Western countries. Many of the active ingredients of these remedies have been successfully purified and standardized, resulting in a diversity of various extracts known as pythochemicals. Tribulus terrestris L (TTL) is one of such herbal remedies that have been successfully tested clinically for its use in the treatment of idiopathic oligoasthenoteratozoospermia (OTA) in male subjects. In this double blind study, 45 infertile married men with OTA were recruited. Thirty six men were treated with 500 mg purified TTL (Libilov) 3 times daily for 3 months. 9 men were given sugar pills (placebo) to compose the control group for the same period of treatment. Spouses of 8 men in the treated group achieved pregnancies after treatment, whereas no spouse of the males in the control group did. No significant differences (p > 0.05) in liver and kidney function tests, as well as serum sex hormone levels (FSH, LH, PRL, T) were reported before and after treatment. The results of the semen analyses done before and after treatment of both the control and the treated groups, however, showed a significant increase in sperm concentration. Hence, the increased sperm concentration did not seem to contribute to the successful pregnancies. No improvement of sperm motility were observed between the two groups. There was, however, improvement in sperm morphology in the treated group (p > 0.05), which could explain the successful pregnancies in this group. Better sperm morphology was observed microscopically by the conventional phase contrast procedure. TTL seemed to improve the acrosome morphology of the sperm cells and enhanced the acrosome reaction, thus contributing to increased fertility. The active component of TTL, termed protodioscin, was shown to increase the level of dehydroepiandrosterone (DHEA), and might also have contributed to the activation of cell membrane receptors and the activation of weak androgens, thus contributing to increased spermatogenesis and fertility. INTRODUCTION In the late 20th century, herbal medicines became more popular, especially after scientific and pharmacological efforts to identify active components were able to introduce standardization to this field. This contributed very much to our understanding of the role of the natural bioactive phytochemical components in traditional herbal medicines in benefiting human health and in understanding its possible side-effects as well. Tribulus terrestris L (TTL) is one such herbal remedy that has recently been scientifically studied, purified, standardized and clinically tested for its usefulness or benefits in treating male infertility due to oligoasthenozoospermia (Protich et al. 1984). The Tribulus plant has long been used as herbal remedies in Eastern European and Asian countries to treat sexual disorders, such as impotence or to treat infertility. Recent scientific research identified the active ingredient of TTL extracts as a non-steroidal furostanol saponin, which constituted 45% or more of the purified extract (Zarkova 1984). Recently, the purified dry powder of TTL, in form of a tablet preparation, has been studied in a multi-center study in Indonesia for treatment of infertile males diagnosed with idiopathic oligoasthenoteratozoospermia (OTA) (Moeloek et al. 1994). The active ingredient of TTL extracts has received approval by the Indonesian Food and Drugs Administration (POM), and is marketed under the name of Libilov™. A total of 45 subjects were involved in this study: 36 of which were treated and 9 of which were given placebo for the entire duration of the study. After 3 months, eight (22%) of the infertile male subjects in the treated group succeeded in impregnating their spouses, whereas no subjects in the control group did. Semen analyses were conducted to determine the factor or factors responsible for the increased fertility of the treated group, as compared to the control group. There was no significant difference in conventional laboratory sperm analyses in both groups before and after treatment. Significant improvements in sperm concentration, however, were observed for both the treated and the control groups. Although this was a significant benefit, as sperm concentration also increased in the control group, we concluded that this could not be the reason for the successful conception of the spouses of the male subjects in the treated group. Preliminary data suggested that improved sperm morphology, specifically improved acrosome reaction, might be the responsible factor. We are currently conducting further research efforts to identify factors responsible for the increased fertility of the treated group. SUBJECTS AND METHODS Six subjects from the Libilov treated group who had successfully impregnated their spouses, six other subjects taken randomly from the treated group, and six subjects from the control group were subjected to laboratory semen analyses. In vitro sperm-cervical mucus interaction test and in vivo post-coital test were conducted (WHO 1987). Preliminary study on sperm acrosome morphology was performed by the acrosin staining method (Pedersen, 1976; Schill 1976). RESULT Better penetration was observed in sperms from the treated group, which consisted of subjects that successfully or unsuccessfully impregnated their spouses, as compared to that of the control group. However, no significant differences were observed in sperm samples from within the treated groups, i.e. between those who were successful in impregnating their spouses as compared to those who were not. Post-coital tests performed also suggested improvements in the treated group as compared to the control group. There was, however, no significant difference between those who successfully impregnated their spouses and those who did not. Acrosin staining revealed a significant increase in the percentage of normal acrosomes in sperms from the treated subjects who succeeded in impregnating their spouses. In contrast, no increase was observed in sperms from both the control group subjects or from the treated subjects who were not successful in achieving pregnancy in their spouses. DISCUSSION AND CONCLUSION The clinical study described in this paper resulted in successful fertilization in 22% of subjects that took part as the treated group. Detailed analyses of the sperm from the treated and control groups revealed that there were significant improvements in sperm morphology, especially the acrosome reaction, but no significant differences in sperm concentration or motility. The improved spermcervical mucus interaction and improvements measured by the post-coital test suggested that the increased acrosome reaction led to the improved sperm functions, thus resulting in successful conception. A further study on the effect of TTL treatment on acrosome reaction is currently being conducted. The chemical structure of protodioscin is very similar to that of dehydroepiandrosterone (DHEA), one of the most important hormones in the body. We speculate that protodioscin can stimulate the production of DHEA, or can serve as a non-hormonal precursor to DHEA. Protodioscin-stimulation of DHEA levels can also contribute to increased fertility, as DHEA has been reported in several studies to be important in the process of sperm maturation in the epididymis. Furthermore, the conversion of DHEA to testosterone or dihydrotestosterone, both potent androgens or sex hormones important for spermatogenesis in the seminiferous tubules, can also contribute to increased fertility. A better understanding of the function of DHEA in general health, and specifically in the male reproduction and sexual functions, is therefore also important to our understanding of infertility and its treatment. 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