07016 Effect of fs! (Otimun Sanctum Linn} on sperm ccurt and reproductive ho:mones In ale alo rabets & International Journal of ont Ayurveda Research cc: nt. Ayurveds Re, 2010 Oct-Dec; 14): 208-210, aco: eucaosdat ot 10.41030974-7709.76762 Effect of tulsl (Ocimum Sanctum Linn.) on sperm count and reproductive hormones In male albino rabbits Jyoll Sethl, Mridul Yaday, Sushma Sood, Kiran Dahiya,’ and Veena Singh* Department of Physiology, Pt BD. Sharma University of Health Sciences, Rehtak, Haryana, Inia “Department of Biochemistry Pt BD, Sharma Unversity of Heat Sciences, Rohtak, Haryans, Ino [Address for correspondence: Dy. .yot Sot, Department of Frysioogy, PLB. D Sharma Lnversty of Heath Sclences, Rotak Haryana, India; El de jotsetnigredtfn.com Received 2010 May 15, Accoptes 2011 an 15 (Comat © ntenatona Journal of Ayurveda Research ‘his 6 an open access arte dstrbuted under ine tame of me CreatWe Commons Attn Noncommercial Share Alke 3.0 Unported, which permis unresticted use, dstibuton, and reproduction in ary medium provided the orga work's propery cite, Abstract Fresh leaves of Ocimum Sanctum (C $) were used to study its effect on male reproductive function (sperm count and reproductive hormones) in male albino rabbits. Animals nthe test group received supplementation of 2 g of ffesh leaves of OS per rabbit sor 30 days, while the control group was maintained on normal dict for the same duration. Sperm count and hormonal estimation (testosterone, flee stimulating hormone (FSH), ans! ktenizing hormone (LH)] were done in serum samples of both groups and compared. significant decrease was noted in the sperm count in test grour rabbits, Serum testosterone levels showed marked increase while FSH and LH levels were sianificantly reduced in S-treated rabbits, The resus suggest the potential use of OS as an effective male contraceptive agent Keywords: Follicle stimulating hormone. luteinizing hormone, Ocimum Sanctum, sperm count, testosterone IntrooucTion Ocimum Sanctum ( S)~ popularly known as tuisi in Hindi and holy basil in English is one of the sacred herbs for Hindus inthe Indian subcontinent, It has a versatile role in traditional medicine (1] The whole plant of Ocimum Sanctum bas medicinal value, athough mostly the leaves, and sometimes the seeds, are used [2] Farler studies with OS have indicated that the plant has hypoglyeaemic,[3] hypolipxdaemic,[4] adaptogenie, [5] anti-cancer, [6] radioprotective,(Z] analgesic and ant- inflammatory properties [8] Vilage women and Ayurvedic physicians have been reported to be using tusi eaves for anti-fertiityeflect, [9] but this type of traditional practice has been limited to rural areas of India. rational approach to this traditional practice with modern seientific methods is not available in medical iterature in spite ofthe easy availabilty of OS in India, Leaves of OS have antizygoti, anti implantation and early abortificient effect in women and in experimental animals [10] Since no stuly available te document the effect of OS on the levels of reproductive hormones, mainly luteinizing hormone (L1) and! folicle stimulating hormone (FSH), the present study was undertaken to analyse the effect of € S on the spetm count and reproductive hormones in male rabbits Marertats ANo MetHons Animals Mae albino rabbés weighing 1.5-2.5 kg were procured fiom the disease-ffee animal house of the CCS Haryana Agriculture University, Hisar (Haryana, India). The rabbits were housed under controlled conditions of light (12-h light-dark cycle) and temperature [(23 + 2)°C] with free access to respective diets and water ad libitum for a period of 30 days. Fresh tusi kaves were collected, ekzaned, and weighed and used in the study. Institutional ptt rimrih govpcfarclesiPMC3059461/7rapormorireale 107016 Etec of ts! (Ccraun Sanctum Lin) on sperm cour and errs otmones nae aloo rabkits Animal Ethics Commitee (IAEC) approval (IAEC/PATHO/08/2352-58 dated 18.09.08) was obtained before the experiment and care of animal was taken as per guidelines of CPCSEA, Department of Animal Weltare Government of India. EXPERIMENTAL DESIGN Rabbits were divided into two groups of 10 each, Group I (Control) was maintained on standard chow diet, ‘whereas Group II (Test) was administered the same diet that the control group rabbits received along with oral supplementation with 2g fiesh leaves of Ocimum Sanctum daily for 30 days. Afier 30 days, blood samples were taken ffom the pinna vein for assessinent of hormone levels (testosterone, FSH, and LH) ffom both groups, The honmones were assayed by principk: of chemiluminescence using, chemiluminometer (Advia Centaur CP, Siemens) [1] and sperm count was assessed in all animals. The cauda ‘epiidymis flom both sides were removed and washec repeatedly in 10 ml of normal physiological saline. Spermatozoa were counted by using I-ml aliquots of sperm suspension with the help of'a haemocytometer. [12] ‘The animals were sacrificed by phenobarbitone 30 mwkkg IV dose. Data obtained were analysed statstically by applying Student's t test using SPSS version 14.0. Resut In the present study. a significant (P <0.001) decrease in sperm count was observed in rabbits fed on OS leaves. ‘as comparee to control animals, A marked increase in serum testosterone level was observed in OS-treated rabbits as compared t¢ control, However LH level was significantly reduced in this group. In fact, LH level was undetectable in the serum ofall animals, FSH levels in the test group decreasee (0.13 + 0,03 mIUimb as compared tc that in the control group (0.64 + 0.05 mIU/m) [Table Lh Discussion ‘Mak reproductive process is regulated by intricately balanced mechanisms involving the hypothalamas-pitutary- testis axis and accessory sex organs It is believed that for initiation as well as maintenance of spermatogenesis in ‘humans, both FSH and testosterone are needed. The gonadotropin-releasing hormone (GnRH) seereted by the hypothalamus regulates the synthesis and release of FSH and LH fiom the pituitary. FSH aets on the Sertoli ces, which are located within the seminiferous tubules in close proximity of developing germ cells, and stimuiates production of various proteins inchuling inhibin, androgen-binding protein (ABP), aromatase, anti- Mullerian hormone (AMI), ete, The LH acts on the Leydig cells located in the itertubular space and stimulates production of testosterone, Intraesticular testosterone concentration in humans is about 200-300 times higher than that in peripheral circulation. Testosterone has profound influence on germ cell development and differentiation It exerts ‘a negatne feedback action on LH secretion, and also on FSH (at higher concentration) acting on hypothalamic- pituitary axis [131 Results ofthe present study clearly show that tusi treatment (2 g/day) brings about a reduction in sperm court, ‘which isin agreement with the earlier reports. Khanna et al, have reported significant decrease in sperm count and motity as well as decrease in the weigh of testes, epididyms, seminal vesicle, and ventral prostate after long-term feeding of OS leaves [14] The long-term feeding of fresh tulsi eaves (465 merkg/day) have shown to increase the body weight, while decrease the weights oftestes, prostate, and adrenal glare in rats. The results suggested that infertlty in male rats seems to be due to impairment of spermatogenesis as well as changes like decrease in pll, hypotonic environment, andl chemical substances Ike mucoproteins, alkaline phosphatase and acid phosphatase in spermatogenic cell leading to formation of non-viabke spermatozoa [15] However, Seth ef al, reported that benzene extract of ( S leaves significantly altered the weight of testes but did not have any significant effet on epididymis, semial vesicle, prostate, and vas deferens (16) Treatment with OS leaves led to a highty significant increase in testosterone levels, FSH and LH levels also showed a significant decrease inthe test group aflertulsi treatment. A possible hypothesis to exphin this pattem of changes in hormone levels could be that tus leaves: probably contain some androgenic analogue, which increased the circulating testosterone levels suliciently to inhibi LI but not sufficient to accumulate inthe testis at the requited concentration for normal spermatogeness However, the decreased 11 levels will diminish intratesticular production of testosterone by Leydig cells, which results in reduced levels of spermatogenesis. For suppression of spermatogenesis, diferent androgens and ptt rimrih govpcfarclesiPMC3059461/7rapormorireale07016 Etec of ts! (Ccraun Sanctum Lin) on sperm cour and errs otmones nae aloo rabkits progestins have been wed ether alone or in combination. Reddy and Rao administered testosterone propionate 100 mg daily intramuscularly to normal volunteers an achieved azoospermia in 100% subjects 17] Weekly IM ‘nections of 200 mg testosterone enanthate given to 399 normal healthy fertile males produced azpospermia or severe oligospermia (spetm density <3 milion/mnl) in more than 95% vokateers. This contraceptive effect was comparable to female contraceptive pill, ark! was reversible [18] Itcan thus be concluded that antispermatogenic effect of OS is brought about by modulation of levels of| reproductive hormones. Since this isa prelimiary study, further studies are required to establish the role of CS as ‘an effective herbal male contraceptive Footnotes Source of Support: Nil Confilet of Interest: None declaree REsERENCES: 1. Ghosh GR, Tulasi (N. Tulasi (N.C, Labiatae, Genus-Deimum) New approaches to Medicine ané Health (NANAH) 1995,323-9, 2, Gupta SK, Prakash J, Srivastava S. Validation of traditional claim of Tulsi Ocimum Sanctum Linn. 98 ¢ ‘medicinal plant. Indian J Exp Biol 2002:40:765~73. [PubMed: 12597545] 3. GiriJ, Suganthi B, Meera G. Effect of Tulasi (Ocimum Sanctum) on diabetes mellitus. Indian J Nutr Dietet. 1987.24:193-8, 4. RaiV, Fien U, Mani V, Iyer UM. Efféct of Ocimum Sanctum leat power on blood lipoproteins, elveated proteins and total amino acids in patients with nor-insulin dependent diabetes mellitus, J Nutr Environ Med 1997,7113-8 5. Bhargave KP, Singh N. Anti-siress activity of Ocimum Sanctum Lin. Indian J Med Res, 1981;73.443-51 [PubMed: 7275241] 6. Aruna K, Sivaramakrishnan VM. Ante carcinogenic effécts of some Indian plant products. Food Chem Toxicol 1992:30.953-6. [PubMed: 1473788] 7. Devi PU, Ganasoundari A. Radinprotective effect of leaf extract of Indian medicinal plat Qcinzum Sanctum. Indian J Exp Biol, 1992;30:953-6. [PubMed: 1473788] 8, Singh S, Majumdar DK, Rehan HM. Evaluation of antéinflammatory potential of fixed oil of einum Sanctum (hoty basil) and its possible mechanism of action. J Ethanopharmacol, 1996:54:19-26. 9, Batla SK, SanthakumeariG. Anti&rtldy effect of Ocimum Sanctum and Hibiscus Rose Sinensis. Indian J Med Res. 1971:59:777-81. [PubMed: 5097076] 10. Vora SB, Garg SK, Chaudhary RR. Antifentily sereening of plants Part III. Effect of six indigenous plants on carly pregnancy in albino rats, Indian J Mec Res, 1969.57:893-9, [PubMed 5820437] 11. Rojanasakul A, Udomsubpayakul U, Chinsomboon S. Chemiuminescence versus ragioimmunoassay for the measurement of reproductive hormones. Int J Gynaecol Obstet. 1994:45:141-6. [PubMed: 7915682] 12, Seal LC, Oliyami KA, C motuyi IO. Tocopherol attenuates testicular toxicity associated with experimental eryptorchidism in ats, A ican J Biotechnology, 2007:6:1 373-7. 13. White BA. Beme arxi Levy Physiology. In: Koeppen BM, Stanton BA, editors. ‘The male and female reproductive systems. 6th ed. Pennsyhania: Ebevier; 2009. pp. 758-97. 14. Kham S, Gupta SK. Grover JK. Filet oflong term feeding of Tuisi (Ocimum Sanctum L) on reproductive performance of aduit albino rats. Indian J Exp Biol. 1986;24:302—4. [PubMed: 3770821] 15, Kashinathan $, Ramakrishnan $, Basu SL. Antiferily effet of Ocimum Sanctum L. Indian J Exp Biol 1972;10:23-5. [PubMed: 4344433] ptt rimrih govpcfarclesiPMC3059461/7rapormorireale07016 Effect of fs! (Otimun Sanctum Linn} on sperm ccurt and reproductive ho:mones In ale alo rabets 16, Seth SD, JohriN, Sundaram KR. Antispermic effect of Ocimum Sanctum. Indian J Exp Biol. 1981;19:975— 6, [PubMed: 7309144] 17. Reddy PR, Rao IM. Reversible antiterilty action of Testosterone Propionate in hurnan males. Contraception, 1972;5:295-302. [PubMed: 4650654] 18, WHO. task force on Methods for reguiation of male fertility Contraceptive efficacy of Testosterone induced! “Azoospetmic and Oligospermis in men, Fertil Steril 1996,65:821-9. [PubMed: 8654646] Figures and Tables Table 1 Grove ‘Sperm coure(milionm) SeramTertrtrone nga) Serum FSH (miUimip ‘Serum LH lV) Conor waazs0 262102 0642005 0522002 Test s0as57" >aseac0 on soot Undetectabie ‘esonreeieeh alan a enced he mar Sani ae nat eee LM le ndeasae, Pear ap onowedle Sperm count and reproductive hormones in rabbits of control and test groups (mean: SD) ‘Aaticles from International Journal of Ayurved: Research are provided here courtesy of Medknow Publications pub rimih govpmctericlestPMC30SB441/7reportorintae