Journal of Human Reproductive Sciences

: 2012  |  Volume : 5  |  Issue : 1  |  Page : 43--47

Antifertility activity of Cryptolepis sanguinolenta leaf ethanolic extract in male rats

Ayodeji F Ajayi, Roland E Akhigbe 
 Department of Physiology, College of Health Sciences, Ladoke Akintola University of Technology, Ogbomoso, Oyo state, Nigeria

Correspondence Address:
Roland E Akhigbe
Department of Physiology, College of Health Sciences, Ladoke Akintola University of Technology, Ogbomoso, Oyo State


Background: Complementary medicine has grown over time with more botanicals emerging and remaining integral parts of medicare. Such botanicals include Cryptolepis sanguinolenta. AIM: This study investigated the effect of Cryptolepis sanguinolenta leaf ethanolic extract on male reproductive system using rat model. Materials and Methods: Control and treated rats were maintained on control diet. Treated rats also received graded doses of the extract. RESULTS: When compared with the controls, Cryptolepis sanguinolenta treatment led to significant testosterone suppression associated with consequent significant rise in luteinizing hormone (LH) and decrease in sperm count. Treatment with Cryptolepis sanguinolenta did not result in significant attenuation of follicular stimulating hormone (FSH) levels and testicular morphometry. Sperm viability, motility, and morphology were also comparable in all groups. Conclusion: These results suggest that Cryptolepis sanguinolenta possesses anti-androgenic and anti-spermatogenic properties with potential anti-aphrodisiac activity.

How to cite this article:
Ajayi AF, Akhigbe RE. Antifertility activity of Cryptolepis sanguinolenta leaf ethanolic extract in male rats.J Hum Reprod Sci 2012;5:43-47

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Ajayi AF, Akhigbe RE. Antifertility activity of Cryptolepis sanguinolenta leaf ethanolic extract in male rats. J Hum Reprod Sci [serial online] 2012 [cited 2023 Mar 29 ];5:43-47
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The use of plants in the management of illnesses has been since time antiquity, and has continuously grown over time. Though western medicine has influenced the use of herbal remedies, most rural communities still practice complementary medicine as they are readily and cheaply available healthcare alternatives. [1] Complementary medicine co-exists with the medicare of most societies and is based on the use of natural and local products related to the people's perspective on the world and life. [2],[3] Plants thus remain a major constituent of life in many communities in the world [4],[5] and their utilization in medicare is still well-disseminated around the world. [6],[7],[8],[9]

Cryptolepis sanguinolenta is one of the commonly used plants for its anti-malarial [10],[11],[12],[13],[14],[15],[16] and anti-diabetic activities, particularly in Nigeria and Ghana. [17],[18],[19],[20] It has also been reported to have anti-cancer, [21] anti-microbial, [22],[23],[24],[25],[26],[27],[28] anti-thrombotic, [29] and anti-inflammatory potentials. [30],[31] The biological activities of its different morphological parts have been attributed to its alkaloid constituents. Cryptolepine, an alkaloid, is the major bioactive principle of the plant. [32] In addition to cryptolepine, other minor alkaloids and their salts that have been isolated include the hydrochloride and the 11-hydroxy derivatives of cryptolepine, cryptoheptine, iso- and neo-cryptolepine, quindoline, biscryptolepine, cryptoquindoline, cryptospirolepine, cryptosanguinolentine, cryptotakienine, and cryptomisrine. [33],[34],[35],[36],[37]

Though the therapeutic efficacy of C. sanguinolenta extract in the treatment of a plethora of human illnesses has been established, it is pertinent to evaluate its effects on other systems. This study consequently sought to determine the effect of ethanolic extract of C. sanguinolenta leaf on male reproductive profile in experimental paradigm.

 Materials and Methods

Plant material

Fresh leaves of C. sanguinolenta were obtained from Womirere, Iresi, Osun state and identified by Ugbogu A, Chukwuma E.C, and Shasanya O.S, Forestry Herbarium, Ibadan, Nigeria where a specimen has been deposited (voucher number FHI.108847).

Preparation of extract

C. sanguinolenta leaves air-dried and milled. 526 g of the milled leaves was extracted in 65% v/v ethanol. After the 3 rd day, the leaf extract was separated from the leaf with a cloth sieve. For absolute separation of the leaf from the extract, filter paper was used to sieve the extract into a bottle. The extract was then taken to the laboratory for the process of evaporation. The evaporation process involved the total removal of ethanol and water with which the extraction took place from the extract. The extract was concentrated using a rotary evaporator at 40°C. 0.1 g/ml stock solution was then used for the experiment.


Experiment was performed with male albino rats of Wistar strain of comparable weight. The animals were allowed to acclimatize to the laboratory condition (12:12h light/dark cycle at 25°C ± 2) for 2 weeks and fed on rat chow and water without restriction. The study was approved by the ethical committee of the department, and all procedures were in accordance to the National Institute of Health Guidelines for the Care and Use of Laboratory Animals (NIH, department of Health and Human services publication no. 85-23, revised 1985).

Experimental design

Rats were randomly divided into 4 equal groups. The control was given 1 ml of distilled water (vehicle for extract). Group I, II, and III were given 50, 150, and 250 mg/kg of the extract, respectively. The vehicle and extract were administered orally for 21 days. After the experimental period, blood samples were collected from each rat into plain bottles via cardiac puncture for hormonal assays, and testes were removed from post-euthanized rats.


This study was approved by the ethics committee. All animals received humane care in compliance with the institution's guideline and criteria for humane care as outlined in the National Institute of Health Guidelines for the Care and Use of Laboratory Animals.

Determination of testicular morphometry

The testes were excised, blotted with tissue paper, and weighed. The length and diameter were also measured.

Determination of FSH, LH, and testosterone

Serum FSH, LH, and testosterone concentrations were estimated by the enzyme-linked immunosorbent assay (ELISA) using standard assay kits following the manufacturers' instructions.

Estimation of sperm quality/semen analysis

The testes were removed along with the epididymis. The caudal epididymis was separated from the testis and lacerated to collect semen onto the microscope slide. [38] Sperm characteristic was evaluated as described by Woode et al.[39] using 2.9% sodium citrate as buffer and 10 mL phosphate buffer saline for sperm motility and count, respectively. Briefly, sperm motility was examined under the microscope, and sperm count was carried out in the improved Neubauer hemocytometer and calculated. Sperm viability was evaluated using the eosin-nigrosin stain technique. 2 drops of the stain was mixed with semen. A thick smear was prepared, air-dried and examined under microscope. Live/viable sperm cells were unstained while dead/non-viable sperm cells appeared stained. The live and dead sperm were counted and the percentage of each calculated. Sperm morphology was done using 2 drops of Walls and Ewas stain and air-dried and examined under the microscope. The normal sperm cells were counted and the percentage calculated.

Histological study

Testicular tissues were transferred into 10% formalin after being fixed in Bouin's fluid for 6h. They were dehydrated with varying percentage of ethanol; sections were cleared in xylene and embedded in molten wax. Thin sections were cut (5 μm), stained with hematoxylin and eosin, and microscopically analyzed.

Statistical analysis

Results are expressed as Mean ± SEM (n = 6). The difference between the means was determined by one-way Analysis of Variance (ANOVA) complemented with unpaired t-test. In all statistical tests, a value of P < 0.05 was considered significant.


Testicular morphology was comparable in all groups. Though testicular weight, length, and diameter were altered following Cryptolepis sanguinolenta leaf extract administration, the morphometric changes were not statistically significant [Figure 1].{Figure 1}

FSH was statistically similar in all groups while LH was significantly raised in the treated groups when compared with the control. On the other hand, testosterone was significantly reduced in the treated groups when compared with the control in a dose-related manner. The rise in LH and fall in testosterone observed in the treated groups were statistically comparable across the treated groups [Figure 2].{Figure 2}

Sperm motility, viability, and morphology was not statistically different across all groups, however, sperm count was statistically reduced in the treated groups when compared with the control. Similar to LH and testosterone changes, sperm count was reduced in a dose-dependent manner in the treated groups. Treatment of animals with 50 mg/kg of the extract showed the highest reduction in testosterone level, sperm count and a higher rise in LH concentration. However, hormonal and sperm count changes observed across the treated groups were not statistically different [Figure 3].{Figure 3}

Histomorphological observations revealed that administration of the extract did not cause any alteration in the testicular tissues of rats treated with 50 and 150 mg/ kgBW of the extract though rats treated with 250 mg/kgBW of the extract showed mild distortion of the seminiferous tubules.


Plants and their products are integral parts of medicare. They are also a major source of most formulated drugs in western medicine. None of these forms of therapy are, however, without side effects ranging from mild to severe. Though the side effects of a drug could be used for therapeutic purposes in other conditions, it is necessary to evaluate the effects of medicinal plants, their products, and formulated drugs commonly used in the treatment of ailments on other organs/systems to determine their side effects/adverse effects and possibly their beneficial effects on other pathological conditions. This led us to study the pharmacological effect of C. sanguinolenta on the male reproductive system. This seems to be the first study to account for the influence of C. sanguinolenta on the male reproductive indices.

The male reproductive system is a complex that consists of the hypothalamus, anterior pituitary gland, the testes, [40] and intimately related organs like prostate gland, seminal vesicle and bulbourethral glands. These structures work together to maintain the potentness, fertility, and male secondary sexual characteristics. Results from this study showed that C. sanguinolenta did not alter testicular morphometry. This suggests that the plant extract does not affect the gross anatomy of testes. This is in conflict with the previous study of Ansah et al. [41] that documented reductions in testes weights following C. sanguinolenta treatment, especially in animals treated with 1000 mg/kgBW of the extract. The inconsistency seen in this study could be dose-dependent.

The results from this study demonstrate that administration of C. sanguinolenta caused a significant reduction in testosterone level, and a rise in LH concentration, but a normal FSH level. The increase in LH observed in the treated rats is in attendant with the C. sanguinolenta-induced testosterone suppression. Suppression of testosterone is expected to be consequently accompanied with increase in LH and FSH concentrations in an attempt to stimulate the production of more testosterone. The normal level of FSH seen in association with testosterone suppression suggests that the hypothalamic cells, which are responsible for the synthesis and release of gonadotropin-releasing hormone (GnRH), do not function correctly when testosterone levels decrease. [42] Though this feedback seems to be maintained for LH release as it increased following testosterone decline. A potential mechanism through which C. sanguinolenta may reduce testosterone levels is its aromatization into estradiol or impairment of the conversion of one or more of its precursors. Low testosterone levels have been associated with decreased reproductive ability. [42] Hence, it appears that the anti-fertility potential of C. sanguinolenta is mediated at all 3 levels of the male reproductive axis: The hypothalamus, pituitary, and testes.

Sperm characteristics are important reproductive indices as they account for male fecundity. The observation that low sperm count induced by C. sanguinolenta treatment in animals is associated with testosterone suppression is consistent with previous findings. [41] A possible explanation for these observations could be attributed to the alkaloids contained in the botanical extract. Alkaloids-containing C. sanguinolenta has been documented to possess anti-muscarinic, [43] α-adrenoceptor antagonistic, [30] or/and cytotoxic [21],[44] activities. Anti-muscarinic agents have been reported to reversibly impair male fertility by an unknown mechanism. [45],[46] a-adrenoceptor antagonists have been documented to inhibit sperm emission [47],[48] via inhibition on both the neutrally-evoked contractions on vas deferens and sperm transport from the caudal epididymis to the distal vas deferens. [49],[50],[51] Its cytotoxic effect may cause a direct destructive effect on the sperm cells, [52] with consequent reduced sperm count. However, the botanical did not cause any alteration in the sperm motility, viability, and morphology. The distortion of the seminiferous tubules observed in C. sanguinolenta treatment could also be ascribed to its cytotoxic activities.

In conclusion, we have demonstrated that the ani-fertility activity of C. sanguinolenta is associated with its anti-muscarinic,α-adrenoceptor antagonistic and cytotoxic effects. This study suggests that C. sanguinolenta treatment exerts quantitative ani-androgenic and ani-spermatogenic effect.


1Bussmann Rainer W, Paul S, Aserat W, Paul E. Plant use in Odo-Bulu and Demaro, Bale Region, Ethiopia. J Ethnobiol Ethnomed 2011;7:28-81.
2Toledo BA, Galetto L, Colantonio S. Ethnobotanical knowledge in rural communities of Cordoba (Argentina): The importance of cultural and biogeographical factors. J Ethnobiol Ethnomed 2009;5:40.
3WHO. Armonización de los sistemas de salud indígenas y el sistema de salud convencional en las Américas. Washington, DC: 2003.
4Sindiga I, Nyaigotti-Chacha C, Kanunah MP. Traditional Medicine in Africa. Nairobi: East African Educational Publishers; 1995.
5Sindiga I, Kanunah MP, Aseka EM, Kiriga GW. Kikuyu traditional medicine. In: Sindiga I, Nyaigotti-Chacha C, Kanuna MP, editors. Traditional Medicine in Kenya. Nairobi: East African Educational Publishers; 1995.
6Begossi A, Hanazaki N, Peroni N. Knowledge and use of bodiversity in Brazilian hot spots. Environ Dev Sustain 2001;2:177-93.
7Hilgert NI. Plants used in home medicine in the Zenta River basin, Northwest Argentina. J Ethnopharmacol 2001;76:11-34.
8Sarpa G. Plantas empleadas contra trastornos digestivos en la medicina tradicional criolla del Chaco noroccidental. Dominguezia 2002;18: 36-50.
9Chunlin L, Sumei L, Bo L, Shana S, Benxi L. Medicinal plants used by the Yi ethnic group: A case study in central Yunnan. J Ethnobiol Ethnomed 2009;5:13-18.
10Bugyei KA, Boye GL, Addy ME. Clinical Efficacy of a Tea-Bag Formulation of Cryptolepis Sanguinolenta Root in the Treatment of Acute Uncomplicated Falciparum Malaria. Ghana Med J 2010;44:3-9.
11Boye GL. Studies on antimalarial action of Cryptolepis sanguinolenta extract. Proceedings of the International Symposium on East-West Medicine, October 10-11, Seoul, Korea; 1989. p. 243-51.
12Wright CW, Phillipson JD, Awe SO, Kirby GC, Warhurst DC, Quertin-Leclerq J, et al. Antimalarial activity of cryptolepine and some other anhydronium bases. Phytother Res 1996;10:361-3.
13Cimanga K, De Bruyne T, Pieters L, Vlietinck AJ, Turger CA. In vitro and in vivo antiplasmodial activity of cryptolepine and related alkaloids from Cryptolepis sanguinolenta. J Nat Prod 1997;60:688-91.
14Grellier P, Ramiaramanana L, Milleriox V, Deharo E, Shrevel J, Frappier F. Antimalarial activity of cryptolepine and isocryptolepine, alkaloids isolated from Cryptolepis sanguinolenta. Phytother Res 1996;10:317-21.
15Kirby GC, Paine A, Warhurst DC, Noamesi BK, Phillipson JD. In vitro and in vivo antimalarial activity of cryptolepine, a plant-derived indoloquinoline. Phytother Res 1995;9:359-63.
16Noamesi BK, Paine A, Kirby GC, Warhurst DC, Phillipson JD. In vitro antimalarial activity of cryptolepine, an indoquinoline. Trans Roy Soc Trop Med Hyg 1991;85:315.
17Bierer DE, Fort DM, Mendez CD, Luo J, Imbach PA, Dubenko LG, et al. Ethnobotanical-directed discovery of the antihyperglycemic properties of cryptolepine: Its isolation from Cryptolepis sanguinolenta, synthesis, and in vitro and in vivo activities. J Med Chem 1998;41:894-901.
18Bierer DE, Dubenko LG, Zhang P, Lu Q, Imbach PA, Garofalo AW, et al. Antihyperglycemic activities of cryptolepine analogues: An ethnobotanical lead structure isolated from Cryptolepis sanguinolenta. J Med Chem 1998;41:2754-64.
19Luo J, Fort DM, Carlson TJ, Noamesi BK, nii-Amon-Kotei D, King SR, et al. Cryptolepis sanguinolenta: An ethnobotanical approach to drug discovery and the isolation of a potentially useful new antihyperglycaemic agent. Diabet Med 1998;15:367-74.
20Ajayi AF, Akhigbe RE, Adewumi OM, Okeleji LO, Mujaidu KB, Olaleye SB. Effect of ethanolic extract of Cryptolepis sanguinolenta stem on in vivo and in vitro glucose absorption and transport: Mechanism of its anti-diabetic activity. Indian J Endocr Metab 2012; 16:S91-6.
21Charles A, Gooderham Nigel J. The Popular Herbal Antimalarial, Extract of Cryptolepis sanguinolenta, Is Potently Cytotoxic. Toxicol Sci 2002;70:245-51.
22Boakye-Yiadom K. Antimicrobial properties of some West African medicinal plants II. Antimicrobial activity of aqueous extracts of Cryptolepis sanguinolenta (Lindl.) Schlechter. Quart J Crude Drug Res 1979;17:78-80.
23Paulo A, Duarte A, Gomes ET. In vitro antibacterial screening of Cryptolepis sanguinolenta alkaloids. J Ethnopharmacol 1994;44:127-30.
24Silva O, Duarte A, Cabrita J, Pimentel M, Diniz A, Gomes E. Antimicrobial activity of Guinea-Bissau traditional remedies. J Ethnopharmacol 1996;50:55-9.
25Boakye-Yiadom K, Herman Ackah SM. Cryptolepine hydrochloride: Effect on Staphylococcus aureus. J Pharm Sci 1979;68:1510-4.
26Cimanga K, De Bruyne T, Pieters L, Totte J, Tona L, Kambu K, et al. Antibacterial and antifungal activities of neocryptolepine, biscryptolepine and cryptoquindoline, alkaloids isolated from Cryptolepis sanguinolenta. Phytomed 1998;5:209-14.
27Paulo A, Pimentel M, Viegas S, Pires I, Duarte A, Cabrita J. Gomes Cryptolepis sanguinolenta activity against diarrhoeal bacteria. J Ethnopharmacol 1994;44:73-7.
28Sawer IK, Berry MI, Brown MW, Ford JL. Antimicrobial activity of cryptolepine. J Pharm Pharmacol 1993;45:1108-11.
29Oyekan AO, Okafor JP. Effects of cryptolepine alone and in combination with dipyridamole on a mouse model of arterial thrombosis. J Ethnopharmacol 1989;27:141-8.
30Bamgbose SO, Naomesi BK. Studies on cryptolepine II: Inhibition of carrageenan induced oedema by cryptolepine. Planta Med 1981;41:392-6.
31Olajide OA, Ajayi AM, Wright CW. Anti-inflammatory properties of cryptolepine. Phytother Res 2009;23:1421-5.
32Gellert E, Raymond-Hamet, Schlittler E. Die Konstitution des Alkaloids Cryptolepin. (The structure of the alkaloid cryptolepine) Helv Chim Acta 1951;34:642-51.
33Paulo A, Gomes ET, Houghton PJ. New alkaloids from Cryptolepis sanguinolenta. J Nat Prod 1995;58:1485-91.
34Tackie AN, Boye GL, Sharaf MH, Cryptospirolepine, an unique spiro-nonacyclic alkaloid isolated from Cryptolepis sanguinolenta. J Nat Prod 1993;56:653-70.
35Pousset JL, Martin MT, Jossang A, Bodo B. Isocryptolepine from Cryptolepis sanguinolenta. Phytochem 1995;39:735-6.
36Sharaf MH, Schiff PL Jr, Tackie AN, Phoebe CH Jr, Martin GE. Two new indoloquinoline alkaloids from Cryptolepis sanguinolenta: Cryptosanguinolentine and cryptotackieine. J Heterocyclic Chem 1996;33:239-43.
37Sharaf MH, Schiff PL Jr, Tackie AN, Phoebe CH Jr, Johnson RL, Minick D. The isolation and structure determination of cryptomisrine, a novel indolo [3,2-b] dimeric alkaloid from Cryptolepis sanguinolenta. J Heterocyclic Chem 1996;33:789-97.
38Costa LG, Hodgson E, Lawrence DA, Reed DJ. Current Protocol in Toxicology. United States: John Wiley and Sons. Inc.; 2005.
39Woode E, Alhassan A, Chrissie S. Abaidoo. Effect of ethanolic fruit extract of Xylopia aethiopica on reproductive function of male rats. Int J Pharm Biomed Res 2011;2:161-5.
40Emanuele MA, Emanuele NV. Alcohol's effects on male reproduction. Alcohol Health Res World 1998;22:195-201.
41Charles A, Boadu MK, Eric W, Mahama D. Reproductive and Developmental Toxicity of Cryptolepis sanguinolenta in Mice. Res J Pharmacol 2010;4:9-14.
42Emanuele MA, Emanuele N. Alcohol and the male reproductive system. Alcohol Res Health 2001;25:282-7.
43Rauwald HW, Kober M, Mutschler E, Lambrecht G. Cryptolepis sanguinolenta: Antimuscarinic properties of Cryptolepine and the alkaloid fraction at M1, M2 and M3 Receptors. Planta Med 1992;58:486-8.
44Bonjean K, De Pauw-Gillet MC, Defresne MC, Colson P, Houssier C, Dassonneville L, et al. The DNA intercalating alkaloid cryptolepine interferes with topoisomerase II and inhibits primarily DNA synthesis in B16 melanoma cells. Biochemistry 1999;37:5136-46.
45Ratnasooriya WD. Effect of atropine on fertility of female rat and sperm motility. Indian J Exp Biol 1984;22:463-6.
46Sato T, Ban Y, Uchida M, Gondo E, Yamamoto M, Sekiguchi Y, et al. Atropine-induced inhibition of sperm and semen transport impairs fertility in male rats. J Toxicol Sci 2005;30:207-12.
47Homonnai ZT, Shilon M, Paz GF. Phenoxybenzamine: An effective male contraceptive pill. Contraception 1984;29:479-791.
48Ratnasooriya WD, Wadsworth RM. Tamsulosin, a selective alpha 1-adrenoceptor antagonist, inhibits fertility of male rats. Andrologia 1994;26:107-10.
49Doggrell SA. Prazosin selectively inhibits the responses to field stimulation in the rat vas deferens. Eur J Pharmacol 1981;71:447-53.
50Bradley L, Doggrell SA. Modification by desipramine of the effects of 1-adrenoceptor antagonists on the contractile responses of the trisected rat vas deferens. Gen Pharmacol 1985;16:475-82.
51Solomon HM, Wire PJ, Ippolito DL, Toscano TV. Effect of prazosin on sperm transport in male rats. Reprod Toxicol 1997;11:627-31.
52Klaassen CD. Casarett and Doull's Toxicology: The Basic Science of Poisons. 5th ed. New York: McGraw Hill; 1996. ISBN: 0071054766.