American Journal of Neuroscience 3 (1): 17-24, 2012 ISSN 1948-9900 2012 Science Publications

Moringa Oleifera Leaves Extract Attenuates Male Sexual Dysfunction

1,3

Thawatchai Prabsattroo, 2,3Jintanaporn Wattanathorn,

23 13

34
,

Sitthichai Iamsa-ard, , Supaporn Muchimapura and , Wipawee Thukhammee 1 Department of Physiology and Graduate School (Neuroscience Program), Faculty of Medicine, 2 Department of Physiology, Faculty of Medicine, 3 Intergrative Complementary Alternative Medicine Research and Development Group, 4Department of Anatomy, Faculty of Medicine, Khon Kaen University, Khon Kaen, Thailand

Abstract: Problem statement: At present, the novel agent that is effective, cheap and easy to approach for treating male sexual dysfunction is required due to the current poor therapeutic efficacy. Though Moringa oleifera is reputed for aphrodisiac activity in traditional folklore, no scientific evidence is available. Therefore, we aimed to determine the effect of M.oleifera leaves extract on male sexual behaviors in animal model of sexual dysfunction. Moreover, the possible underlying mechanisms were also investigated. Approach: Male Wistar rats, weighing 200-250 g, had been orally given M.oleifera leaves extract at doses of 10, 50 and 250 mg kg-1 BW once daily at 30 min before the exposure to 12-h immobilization stress for 14 days. They were assessed male sexual behaviors including mounting, intromission and ejaculation numbers and latencies after single administration and every 7 days until the end of experiment. To further investigate the possible mechanisms of action, we also determined serum testosterone level of all rats at the end of experiment together with the determination of suppression effect of the plant extract on MAOB and PDE-5 activities. Results: The results showed that after single administration, rats subjected to M.oleifera extract at dose of 10 mg kg-1 BW significantly enhanced mounting number. When the treatment was prolonged to 7 days rats subjected to the low dose of extract showed the enhanced intromission number whereas rats subjected to high dose of extract showed the enhanced mounting number. Our data also showed no significant change in serum testosterone level. However, the extract could also suppress MAOB and PDE-5 activities. Taken all together, the extract could enhance male sexual desire and performance via the suppression of MAOB and PDE-5 activities. Conclusion: M.oleifera can be the potential sexual enhancer particularly for acute and short term application. However, further researches are necessary. Key words: Numerous health products, scientific evidence, sexual enhancer particularly, Erectile Dysfunction (ED), harvested during, room temperature INTRODUCTION Sexual dysfunction has been recognized as one of the important social and biological relationship in human life. This condition affects the sexual life of millions of men worldwide. The prevalence of male sexual dysfunction is increased as the age advanced (Wattanathorn et at., 2012; Moreira et al, 2006). Therefore, the sexual dysfunction is still increasing its prevalence and importance. Sexual dysfunction caused by various factors including stress. Despite the importance of sexual dysfunction, the therapeutic efficacy is still not in satisfaction level. The most well known drug used nowadays appears to target only at intromission phase, the main problem in Erectile Dysfunction (ED) which is the most commonly found sexual dysfunction. However, sexual dysfunction refers to a problem during any phase of the sexual response cycle. Therefore, the searching for new agent targeting at numerous phases of sexual response cycle which is more cheap has gained much concentration. Moringa oleifera Lam. or Drumstick tree, a plant in a family of Moringaceae, is widely cultivated in Thailand. Both leaves and fruit of this tree are edible. It is believed to be miracle herbs because it can be used not only as food but also as medicine which can cure

Corresponding Author: Jintanaporn Wattanathorn, Department of Physiology, Faculty of Medicine and Integrative Complementary Alternative Medicine, Khon Kaen University, Khon Kaen 40002, Thailand

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numerous ailments. The leaves of M.oleifera have been used as used as antiulcer, diuretic, anti-inflammatory and for wound healing (Kirtikar and Basu, 1935; Caceres et al., 1992; Udupa et al, 1994; Pal et a l , 1995). Moreover, it has been used to enhance male sexual functions including libido, improve sperm quality and anti-erectile dysfunction. Nowadays, numerous health products of M.oleifera are available in the market and claimed for the effect on male sexual functions as mentioned earlier, no scientific evidence was observed until now. Therefore, the current study aimed to determine the effect of M.oleifera leaves extract on male sexual behaviors and the possible underlying mechanism in sexual dysfunction rats induced by stress. MATERIALS AND METHODS Plant material: The fresh Moringa oleifera Lam (Moringaceae) were harvested during November to December, 2010 from the Khon Kaen province Thailand. The plant specimen was authenticated by Associate Professor Dr. Panee Sirisa-ard, Faculty of Pharmaceutical Sciences, Chiangmai University, Thailand. The voucher specimen was kept at Integrative Complementary Alternative Medicine Research and Development Group, Khon Kaen University, Khon Kaen, Thailand. Plant material preparation: The fresh leaves were immediately cleaned, than cut into small pieces and dried at the temperature less than 50C. The dried plant material was ground into fine coarse powder and extracted with 50% alcoholic. After that evaporation of solvent in rotary evaporator affords a crude extract of the soluble components and filtrate was lyophilized. The percent yield of extract was 17.49%. The extract contained total phenolic compounds at concentration of 86.73-93.60.51 mg of GAE/g extract. The extracts were stored at -25C in a dark bottle until used. The crude extract was suspended in distilled water. Animals: Healthy male Wistar rats (200-250 g) were obtained from National Animal Center, Salaya, Nakorn Pathom and were housed in group of 6 per cage in standard metal cages at 222C on 10:14 h light-dark cycle. All animals were given access to food and water ad labium The experiments were performed to minimize animal suffering in accordance with the internationally accepted principles for laboratory use and care of European Community (EEC directive of 1986; 86/609/EEC).The experimental protocols were approved by the Institutional Animal Care and Use Committee. Stress procedure: The restraint stress was performed during the light cycle from 6.00 a.m. to 6.00 p.m. The restrainer was made of transparent perforated plastic tube, 20 cm long and 7 cm in diameter. The rats were put into the restrainer, head first and once in, the tubes were closed with plexiglass lids. The animals fit tightly into the restrainers and it was not possible for them to turn around. None stressed control rats were at the same time briefly handled and returned to their home cages.

Am J. Neuroscience 3 (1): 17-24, 2012 Evaluation of sexual behaviors: Male Wistar rats of proven
fertility were randomly divided into 4 groups of 6 animals each as following; (1) Vehicle plus stress (2)- (4) M.oleifera plus stress. Rats in group 1 were administered with distilled water which used as vehicle for the plant extract then expose to the 12 h-restraint stress whereas rats in group (2)-(4) were administered with the plant extract at doses of 10, 50 and 250 mg kg-1 BW plus stress exposure as mentioned earlier. All substances treatments were administered 45 m prior to the 12 h-restraint stress exposure. The treatment and the stress-exposure were performed once daily and the sexual behaviors assessments were performed blindly. The animals were allowed to rest in order to refresh the animals 3 h after the removal from restraint cage and then they were assessed the sexual behaviors between 9.00 p.m. to 12.00 a.m. at room temperature 26-28C after single dose,1 and 2 weeks of treatment. In order to assess the sexual behaviors, estrous female rats were paired with male treated with single or repeated doses of extract. Female rats were induced to estrous by sequential administration of estradiol benzoate (Sigma, St. Louis, MO) at dose of 2 ^g kg-1 BW and progesterone (Sigma, St.Louis, MO) at dose of 500 ^g kg-1 BW were injected before the determination of copulatory behaviors via subcutaneous route 48 h and 6 h respectively. Sexual behaviors were monitored in a separate room for 3 h in a clear plastic box via blind observer 30 min at the start of first hour whereas the whole duration of observation (3h) was recorded by digital video recording. The assessed sexual parameters were including the following parameters: Mounting number: The number of mounts without intromission from the time of introduction of the female until ejaculation: Intromission number: The number of intromissions from the time of introduction of the female until ejaculation Mount latency: The time interval between the introductions of the female to the first mount by the male Intromission latency: The interval from the time of introduction of the female to the first intromission by the male Ejaculation number: The number of ejaculation which characterized by longer, deeper pelvic thrusting and slow dismount followed by a period of inactivity Ejaculation latency: The time interval between the first intromission and ejaculation

Determination of testosterone level: Separate groups of animals were used for the measurement of plasma testosterone level of rats subjected to 12-h immobilization stress. At the end of 14-day experimental period, the rat blood samples were collected and kept on ice and then centrifuged immediately at 2000*g at 4C for 15 min. The obtained plasma was kept at-80C until analysis. Testosterone levels were measured using a commercially available radioimmunoassay (RIA) kit (TESTO-CT2, Cisbio International, France). Determination of monoamine oxidase type B inhibition: The inhibitory action of the plant extract on monoamine

oxidase type B was determined by incubating a series of concentrations of the test samples in the reaction mixture including rat brain homogenates. In brief, 2.75 mL Tris buffer (0.1 M, pH 7.4) and 100 ^L of 0.1 M benzylamine was mixed in quartz cuvette which was placed in double beam spectrophotometer and followed by the addition of 150 ^L solution of brain homogenate to initiate the enzymatic reaction. The change in absorbance was recorded at wavelength of 249.5 nm for 5 min against the blank containing Tris buffer and 5-hydroxytryptamine (Xu et al., 2005).

Am J. Neuroscience 3 (1): 17-24, 2012

Statistic analysis: All data were expressed as meanSEM value. The significant differences among various groups were compared by ANOVA and followed by Duncan's test. The statistical difference was regarded at p<0.05. RESULTS Effect of M.oleifera leaves extract on male sexual behavior: The effects of M.oleifera leaves extract on male sexual behavior in animal model of sexual dysfunction induced by 12-h immobilization stress were shown in Fig. 1-6. The current results showed that rats subjected to M.oleifera leaves extract at dose of 10 mg kg-1 BW significantly increased mounting number after single administration whereas other parameters failed to show the significant difference (p-value<0.05; compared to vehicle+stress). When the treatment duration was increased further to 7 days, the rats which obtained M.oleifera leaves extract at dose of 10 mg kg-1 BW showed the enhanced intromission number while the rats which exposed to high dose of M.oleifera leaves extract (250 mg kg-1 BW) showed the increased mounting number (p-value<0.05; compared to vehicle +stress). Unfortunately, no significant changes were observed at 14 days of treatment. Effect of M.oleifera leaves extract on phosphodiesterase (PDE) activity: We had determined the effect of the plant extract on the activity of MAOB and the results were shown in Fig. 7. It was found that Sildenafil citrate or Viagra which was used as positive control in this study could significantly suppress PDE activity. The plant extract both at dose of 500 and at dose of 2000 mg mL-1 also significantly suppressed PDE activity (p- value <0.05 and .01 respectively compared to control). Interestingly, M.oleifera leaves extract at dose of 2000 mg mL-1 could suppress PDE activity at the same magnitude as Sildenafil citrate or Viagra.

Determination of Phosphodiesterase (PDE) activity: Testis was collected from healthy Wistar male rat in order to Determine Phosphodiesterase Enzyme (PDE) activity. The testis was washed with PBS and weighted before cut to small pieces. Then, it was homogenized with 5 volumes of lysate RIPA buffer. The testicular solution was centrifuged at 14,000*g for 15 min at 4C and the supernatant was collected and used as PDE substrate. M.oleifera leaves extract at the various concentrations ranging from 2, 10, 100, 500, 2000 mg mL-1 were prepared together with the positive control sildenafil at dose of 10 mg mL-1. The standard curve was prepared from PDE (testicular lysate) at the various concentrations. The experiment was divide into 7 groups as following (1) control group (untreated group), (2-6) M.oleifera leaves extract treated groups at the various concentrations (2, 10, 100, 500, 2000 mg ml) and (7) positive control (10 mg mL-1 of sildenafil citrate). PDE-Glo Phosphodiesterase assays were performed in a white 96-well microplate. In brief, phosphodiesterase substrate or testicular lysate was incubated with cGMP. Then, PDE reaction solutions were added and incubated for 20 min at room temperature. The cGMP in the mixture then drives a kinase reaction leading to a reduction of ATP levels. Following the kinase reaction, an Kinase Glo reagent was added and reactions were mixed and incubated for 10 min at room temperature. Luminescence was measured using a SpectraMax L microplate luminometer (MSD AT (US) Inc). The luminescent signal produced is directly related to the amount of ATP remaining and correlates with phosphodiesterase activity.

Fig. 1:Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on mount latency. Data were presented as mean SEM (n=6 group-1). * p<0.05 compared with vehicle plus stress treated group

Am J. Neuroscience 3 (1): 17-24, 2012

Fig. 2: Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on mount number. Data were presented as mean SEM (n = 6 group-1). * p<0.05 compared with vehicle plus stress treated group

Fig. 3: Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on intromission latency. Data were presented as mean SEM (n = 6 group )

Fig. 4: Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on intromission number. Data were presented as mean SEM (n = 6 group-1). * p<0.05 compared with vehicle plus stress treated group

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Am J. Neuroscience 3 (1): 17-24, 2012

Fig. 5:Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on ejaculation latency. Data were presented as mean SEM (n = 6 group-1)

Baseline
D

Single dose

Day 7
n

Day 14

Vehicle + stress

M.oleifera 50 mg kgi BW + stress

M.oleifera 10 mg kg"1 BW + stress M.oleifera 250 mg kg i BW + stress

Fig. 6: Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on ejaculation number. Data were presented as mean SEM (n = 6 group-1).

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Am J. Neuroscience 3 (1): 17-24, 2012

Fig. 7: Effect of M.oleifera leaves extract (0, 2, 10, 100, 500, 2000 ^g ml-1) and Sildenafil citrate (10 ^g mL-1) on phosphodiesterase (PDE) activity. Data were presented as mean SEM. a aa p<0.05 and 0.01 respectively, compared with control group respectively SEM (n = 6 group-1)

Fig. 8:Effect of M.oleifera leaves extract (0, 50, 100, 250, 500, 1000 ^g ml-1) on monoamine oxidase type B (MAO-B) activity. Data were presented as mean SEM. ^ a1, aaa P-value < 0.05, 0.01 and 0.001 respectively, compared with control group

Fig. 9: Effect of M.oleifera leaves extract (10, 50, 250 mg kg 1 BW) on testosterone. Data were presented as mean

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Am J. Neuroscience 3 (1): 17-24, 2012

Effect of M.oleifera leaves extract on monoamine oxidase type B (MAOB) activity: The effect of M.oleifera leaves extract on MAOb activity was evaluated and the results were shown in Fig. 8. Our data clearly revealed that the extract at concentration of 50,100, 250, 500 and 1000 mg mL-1 could significantly suppress MAOB activity (p-value<0.05, .05, .01, .01, 0.001 respectively; compared to control. Effect of M.oleifera leaves extract on testosterone: Figure 9 demonstrated the effect of M.oleifera leaves extract on testosterone level. It was found that no significant changes in testosterone levels were observed in all rats subjected to M.oleifera leaves extract plus stress. DISCUSSION The present results provide, for the first time, information concerning the ability of M.oleifera leaves extract to improve male sexual behavior in rats. We have revealed that the plant extract could enhance mounting number and intromission number. Accumulative lines of evidence have demonstrated that male sexual behavior is regulated mainly by neuroendocrine system. It has been reported that male sexual behaviors were under the influence of testosterone level (Seftel et al., 2004; Wang et al., 2004) and dopamine (Dominguez and Hull, 2005; Wattanathorn et al., 2012). Therefore, in this study we also determined the effect of the plant extract on both parameters. Based on the information that dopamine is metabolized mainly by MAOB, the activity of MAOB is used to reflect the available dopamine (Glover et al., 1977). Our data clearly demonstrated that rats subjected to M.oleifera leaves extract could enhance mounting and intromission numbers. Unfortunately, we could not detect the elevation of testosterone level. However, our in vitro data also demonstrated that the extract possessed monoamine oxidase type B inhibitor (MAOBI). Since dopamine plays a crucial role on the regulation of male sexual function in many aspects including motivation and reinforcement, motor response to sexual stimuli and male genital reflex (Dominguez and Hull, 2005), it could be possible that the plant extract suppress MAOB and gave rise to the elevation of dopamine which in turn enhanced libido and copulatory behavior both mounting and intromission numbers. Besides dopamine, it has been found that phosphodiesterase type 5 (PDE 5) also plays a pivotal role on penile tumescence and intromission phase. PDE-5 is the enzyme contributing the important role on the hydrolysis of Guanosine 3'5'-cyclic monophosphate (cGMP), an important second messenger in cellular signal transduction processes including the smooth muscle relaxation. PDE-5 is also abundantly present in the penile corpus cavernosum (Ballard et al., 1998; Moreland et al., 1998) and plays a major role in the relaxation of the corpus cavernosal smooth muscle during sexual stimulation (Andersson and Wagner, 1995). In this study, we have found that M.oleifera leaves extract also significantly suppressed PDE-5 activity. Therefore, the possible underlying mechanism of M.oleifera leaves to enhance intromission might be due to its ability to suppress PDE-5 activity together with the suppression of MAO B activity. Our current data failed to show the alteration of testosterone level. It has been believed that normal adult testosterone levels are not required for normal erections to occur (Bagatell et al., 1994). The age of rats which used as experimental animals in this study were young adult rats and there was no pathological state and they were in eugonadal state. Since testosterone contributed minor role during this period, we have found the enhanced mounting and intromission numbers without the significant change of testosterone level. The current results also showed that the prolonged treatment duration to 14 days failed to show beneficial effect. Although the precise mechanism was not understood, we suggested that this might be due to the adaptation of the nervous system which has been recognized as high plasticity organ. This study provides evidence that M.oleifera leaves can enhance male sexual desire and performance. This enhancement can be ascribed to the suppression of MAOB and phosphodiesterase activities. Therefore, Moringa oleifera may be served as the natural resource for developing functional food and food supplement to enhance sexual function particularly for acute and short term application. In order to provide optimum benefit, the screening of activities of this plant using other organic solvent should also perform in order to select the most suitable fraction as natural resource for further development of sexual enhancer food and related products. In addition, the possible active ingredient and safety evaluation are also very much necessary. CONCLUSION
Moringa oleifera is a potential agent to manage sexual dysfunction induced by stress especially for acute and short term application. Further researches are warranted to confirm this activity before moving forward to clinical trial.

ACKNOWLEDGEMENT This study was supported by the Higher Education Research Promotion and National Research University

Am J. Neuroscience 3 (1): 17-24, 2012

Project of Thailand, Office of the Higher Education Commission, through the Food and Functional Food Research Cluster of Khon Kaen University, Integrative Complementary Alternative Medicine Research and Development Group, Khon Kaen University. Moreover, we also would like to express our sincere thank to Associate Professor Bungorn Sripanidkulchai, Director of Center for Research and Development of Herbal Health Product, Khon Kaen University for her kindly management through Functional Food Cluster and Associate Professor Panee Sirisa-ard for her authentication. REFERENCES Andersson, K.E. and G. Wagner, 1995. Physiology of penile erection. Physiol. Rev., 75: 191-236. PMID: 7831397 Bagatell, C.J., J.R. Heiman, J.E. Rivier and W.J. Bremner, 1994. Effects of endogenous testosterone and estradiol on sexual behavior in normal young men. J. Clin. Endocrinol. Metab., 78: 711-716. PMID: 8126146 Ballard, S.A., C.J. Gingel, K. Tang, L.A. Turner and M.E. Price et al, 1998. Effects of sildenafil on the relaxation of human corpus cavernosum tissue in vitro and on the activities of cyclic nucleotide phosphodiesterase isozymes. J. Urol., 159: 21642171. DOI: 10.1016/S0022-5347(01)63299-3 Caceres, A., A. Saravia, S. Rizzo, L. Zabala, E.D. Leon and F. Nave, 1992. Pharmacologie properties of Moringa oleifera. 2: Screening for antispasmodic, antiinflammatory and diuretic activity. J. Ethnopharmacol., 36: 233-237. DOI: 10.1016/0378-8741 (92)90049-W Dominguez, J.M. and E.M. Hull, 2005. Dopamine, the medial preoptic area and male sexual behavior. Physiol. Behav., 86: 356-368. PMID: 16135375 Glover, V., M. Sandler, F. Owen and G.J. Riley, 1977. Dopamine is a monoamine oxidase B substrate in man. Nature, 265: 80-81. DOI: 10.1038/265080a0 Kirtikar, K.R. and B.D. Basu, 1935. Indian Medicinal Plants. 2nd Edn., Lalit Mohan Basu, Allahabad.
PHYTOTHERAPY RESEARCH Phytother. Res. 21, 17-25 (2007) Published online 6 November 2006 in Wiley InterScience (www.interscience.wiley.com) DOI: 10.1002/ptr.2023

Moreira, E.D., S.C. Kim, D. Glasser and C. Gingell, 2006. Sexual activity, prevalence of sexual problems and associated help-seeking patterns in men and women aged 40-80 years in Korea: data from the Global Study of Sexual Attitudes and Behaviors (GSSAB). J. Sex Med., 3: 201-211. DOI: 10.1111/j.1743- 6109.2006.00210.x Moreland, R.B., I. Goldstein and A. Traish, 1998. Sildenafil, a novel inhibitor of phosphodiesterase type 5 in human corpus cavernosum smooth muscle cells. Life Sci., 62: 309-318. PMID: 9600334 Pal, S.K., P.K. Mukherjee and B.P. Saha, 1995. Studies on the antiulcer activity of Moringa oleifera leaf extract on gastric ulcer models in rats. Phytother Res., 9: 463-465. DOI: 10.1002/ptr.2650090618 Seftel, A.D., R.J. Mack, A.R. Secrest and T.M. Smith, 2004. Restorative increases in serum testosterone levels are significantly correlated to improvements in sexual functioning. J. Androl., 25: 963-972. PMID: 15477371 Udupa, S.L., A.L. Udupa and D.R. Kulkarni, 1994. Studies on anti-inflammatory and wound healing properties of Moringa oleifera and Aegle marmelos. Fitoterapia, 65: 119-123. Wang, C., G. Cunningham, A. Dobs, A. Iranmanesh and A.M. Matsumoto et al., 2004. Long-term testosterone gel (AndroGel) treatment maintains beneficial effects on sexual function and mood, lean and fat mass and bone mineral density in hypogonadal men. J. Clin. Endocrinol. Metab., 89: 2085-2098. DOI: 10.1210/jc.2003-032006 Wattanathorn, J., P. Pangphukiew, S. Muchimapura, K. Sripanidkulchai and B. Sripanidkulchai, 2012. Aphrodisiac activity of Kaempferia parviflora. Am. J. Agric. Biol. Sci., 7: 114-120. DOI: 10.2844/ajabssp.2012.114.120 Xu, Y., B.S. Ku, H.Y. Yao, Y.H. Lin and X. Ma et al., 2005. The effects of curcumin on depressive-like behaviors in mice. Eur. J. Pharmacol., 518: 40-46. DOI: 10.1016/j.ejphar.2005.06.002
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REVIEW ARTICLE

Moringa oleifera: A Food Plant with Multiple Medicinal Uses

Farooq Anwar1, Sajid Latif1, Muhammad Ashraf2 and Anwarul Hassan Gilani3*
1

Department of Chemistry, University of Agriculture, Faisalabad-38040, Pakistan 'Department of Botany, University of Agriculture, Faisalabad-38040, Pakistan 3 Department of Biological and Biomedical Sciences, Aga Khan University Medical College, Karachi-74800, Pakistan

Moringa oleifera Lam (Moringaceae) is a highly valued plant, distributed in many countries of the tropics and subtropics. It has an impressive range of medicinal uses with high nutritional value. Different parts of this plant contain a profile of important minerals, and are a good source of protein, vitamins, -carotene, amino acids and various phenolics. The Moringa plant provides a rich and rare combination of zeatin, quercetin, j3- sitosterol, caffeoylquinic acid and kaempferol. In addition to its compelling water purifying powers and high nutritional value, M. oleifera is very important for its medicinal value. Various parts of this plant such as the leaves, roots, seed, bark, fruit, flowers and immature pods act as cardiac and circulatory stimulants, possess antitumor, antipyretic, antiepileptic, antiinflammatory, antiulcer, antispasmodic, diuretic, antihypertensive, cholesterol lowering, antioxidant, antidiabetic, hepatoprotective, antibacterial and antifungal activities, and are being employed for the treatment of different ailments in the indigenous system of medicine, particularly in South Asia. This review focuses on the detailed phytochemical composition, medicinal uses, along with pharmacological properties of different parts of this multipurpose tree. Copyright 2006 John Wiley & Sons, Ltd.
Keywords: Moringa oleifera; phytomedicine; food plant; medicinal uses; pharmacological properties; natural coagulant.

INTRODUCTION Moringa oleifera Lam (syn. M. ptreygosperma Gaertn.) is one of the best known and most widely distributed and naturalized species of a monogeneric family Moringaceae (Nadkarni, 1976; Ramachandran et al., 1980). The tree ranges in height from 5 to 10 m (Morton, 1991). It is found wild and cultivated throughout the plains, especially in hedges and in house yards, thrives best under the tropical insular climate, and is plentiful near the sandy beds of rivers and streams (The Wealth of India, 1962; Qaiser, 1973). It can grow well in the humid tropics or hot dry lands, can survive destitute soils, and is little affected by drought (Morton, 1991). It tolerates a wide range of rainfall with minimum annual rainfall requirements estimated at 250 mm and maximum at over 3000 mm and a pH of 5.0-9.0 (Palada and Changl, 2003). Moringa oleifera, native of the western and sub- Himalayan tracts, India, Pakistan, Asia Minor, Africa and Arabia (Somali et al., 1984; Mughal et al., 1999) is now distributed in the Philippines, Cambodia, Central America, North and South America and the Caribbean Islands (Morton, 1991). In some parts of the world M. oleifera is referred to as the 'drumstick tree' or the 'horse radish tree', whereas in others it is known as the

good source of natural antioxidants; and thus enhance the shelf-life of fat containing foods due to the presence of various types of antioxidant compounds such as ascorbic acid, flavo- noids, phenolics and carotenoids (Dillard and German, 2000; Siddhuraju and Becker, 2003). In the Philippines, it is known as 'mother's best friend' because of its utilization to increase woman's milk production and is sometimes prescribed for anemia (Estrella et al., 2000; Siddhuraju and Becker, 2003). A number of medicinal properties have been ascribed to various parts of this highly esteemed tree (Table 1). Almost all the parts of this plant: root, bark, gum, leaf, fruit (pods), flowers, seed and seed oil have been used for various ailments in the indigenous medicine of South Asia, including the treatment of inflammation and infectious diseases along with cardiovascular, gastrointestinal, hematological and hepatorenal disorders

* Correspondence to: Professor Anwarul Hassan Gilani, Department of Biological and Biomedical Sciences, Aga Khan University Medical College, Karachi-74800, Pakistan. E-mail: anwar.gilani@aku.edu

kelor tree (Anwar and Bhanger, 2003). While in the Nile valley, the name of the tree is 'Shagara al Rauwaq', which means 'tree for purifying' (Von Maydell, 1986). In Pakistan, M. oleifera is locally known as 'Sohanjna' and is grown and cultivated all over the country (Qaiser, 1973; Anwar et al., 2005). Moringa oleifera is an important food commodity which has had enormous attention as the 'natural nutrition of the tropics'. The leaves, fruit, flowers and immature pods of this tree are used as a highly nutritive vegetable in many countries, particularly in India, Pakistan, Philippines, Hawaii and many parts of Africa (D'souza and Kulkarni, 1993; Anwar and Bhanger, 2003; Anwar et al., 2005). Moringa leaves have been reported to be a rich source of -carotene, protein, vitamin C, calcium and potassium and act as a
Copyright 2006 John Wiley & Sons, Ltd.

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F. ANWAR ET AL.

Table 1. Some common medicinal uses of different parts of Moringa oleifera

Plant part Root Medicinal Uses Antilithic, rubefacient, vesicant, carminative, antifertility, anti-inflammatory, stimulant in paralytic afflictions; act as a cardiac/circulatory tonic, used as a laxative, abortifacient, treating rheumatism, inflammations, articular pains, lower back or kidney pain and constipation, Purgative, applied as poultice to sores, rubbed on the temples for headaches, used for piles, fevers, sore throat, bronchitis, eye and ear infections, scurvy and catarrh; leaf juice is believed to control glucose levels, applied to reduce glandular swelling Rubefacient, vesicant and used to cure eye diseases and for the treatment of delirious patients, prevent enlargement of the spleen and formation of tuberculous glands of the neck, to destroy tumors and to heal ulcers. The juice from the root bark is put into ears to relieve earaches and also placed in a tooth cavity as a pain killer, and has anti-tubercular activity Used for dental caries, and is astringent and rubefacient; Gum, mixed with sesame oil, is used to relieve headaches, fevers, intestinal complaints, dysentery, asthma and sometimes used as an abortifacient, and to treat syphilis and rheumatism High medicinal value as a stimulant, aphrodisiac, abortifacient, cholagogue; used to cure inflammations, muscle diseases, hysteria, tumors, and enlargement of the spleen; lower the serum cholesterol, phospholipid, triglyceride, VLDL, LDL cholesterol to phospholipid ratio and atherogenic index; decrease lipid profile of liver, heart and aorta in hypercholesterolaemic rabbits and increased the excretion of faecal cholesterol Seed extract exerts its protective effect by decreasing liver lipid peroxides, antihypertensive compounds thiocarbamate and isothiocyanate glycosids have been isolated from the acetate phase of the ethanolic extract of M o r i n g a pods

The Wealth of India, 1962; Padmarao e t a l . ,

1996; Dahot, 1988; Ruckmani e t a l . , 1998 Morton, 1991; Fuglie, 2001; Makonnen e t a l . , 1997;

References

Leave

The Wealth of India,

1962; Dahot, 1988 Bhatnagar e t a l . , 1961; Siddhuraju and Becker, 2003

Stem bark

Gum

Fuglie, 2001

Flower

Seed

Nair and Subramanian, 1962; Bhattacharya e t a l . , 1982; Dahot, 1998; Siddhuraju Becker, Faizi e t a l . ,and 1998; Lalas 2003; Mehta 2002 e t a l . , 2003 and Tsaknis,

(The Wealth of India, 1962; Singh and Kumar, 1999; Morimitsu et al., 2000; Siddhuraju and Becker, 2003). The seeds of Moringa are considered to be antipyretic, acrid, bitter (Oliveira et al., 1999) and reported to show antimicrobial activity (The Wealth of India, 1962). The seed can be consumed fresh as peas; or pounded, roasted, or pressed into sweet, non-desiccating oil, commercially known as 'Ben oil' of high quality. The unique property is the ability of its dry, crushed seed and seed press cake, which contain polypeptides, to serve as natural coagulants for water treatment (Ndabigengesere and Narasiah, 1998). So far no comprehensive review has been compiled from the literature encompassing the efficacy of this plant in all dimensions. Its versatile utility as a medicine, functional food, nutraceutical and water purifying potential motivated us to bridge the information gap in this area, and to write a comprehensive review on the medicinal, phytochemical and pharmacological attributes of this plant of high economic value.

Purified, whole-gum exudate from M. oleifera has been found to contain L-arabinose, -galactose, -glucuronic acid, and L-rhamnose, -mannose and -xylose, while a homogeneous, degraded-gum polysaccharide consisting of L-galactose, -glucuronic acid and L-mannose has been obtained on mild hydrolysis of the whole gum with acid (Bhattacharya et al., 1982). Flowers contain nine amino acids, sucrose, D-glucose, traces of alkaloids, wax, quercetin and kaempferat; the ash is rich in potassium and calcium (Ruckmani et al., 1998). They have also been reported to contain some flavonoid pigments such as alkaloids, kaempherol, rhamnetin, isoquercitrin and kaempferitrin (Faizi et al., 1994a; Siddhuraju and Becker, 2003). Antihypertensive compounds thiocarbamate and isothiocyanate glycosides have been isolated from the acetate phase of the ethanol extract of Moringa pods (Faizi et al., 1998). The cytokinins have been shown to be present in the fruit (Nagar et al., 1982). A new 0-ethyl-4-(a-L-rhamnosyloxy)benzyl carbamate [11]

PHYTOCHEMISTRY Moringa oleifera is rich in compounds containing the simple sugar, rhamnose and a fairly unique group of compounds called glucosinolates and isothiocyanates (Fahey et al., 2001; Bennett et al., 2003). The stem bark has been reported to contain two alkaloids, namely moringine and moringinine (Kerharo, 1969). Vanillin, -sitosterol [14], -sitostenone, 4-hydroxymellin and octacosanoic acid have been isolated from the stem of M. oleifera (Faizi et al., 1994a).

Copyright 2006 John Wiley & Sons, Ltd.

Phytother. Res. 21, 17-25 (20 DOI: 10.1002

MORINGA OLEIFERA

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14: R = H 15: R = 6'-0-oleoyl-P-D-glucop5Tanosyl 16: R = P-D-glucopyranosyl

Fiqure 1. Structures of selected phytochemicals from Moringa: niazinin A [1], 4-(4'-0-acetyl-a-L-rhamnopyranosyloxy)benzyl isoth- iocyanate [2], 4-(-L-rhamnopyranosyloxy)benzyl isothiocyanate [3], niazimicin [4], 4-(a-L-rhamnopyranosyloxy)benzyl glucosinolate [5], benzyl isothiocyanate [6], aglycon of deoxy-niazimicine (N-benzyl, S-ethylthioformate) [7], pterygospermin [8], niaziminin [9 + 10], 0-ethyl-4-(a-L-rhamnosyloxy)benzyl carbamate [11], niazirin [12], glycerol-1-(9-octadecanoate) [13], ^-sitosterol [14], 3-0-(6'-0-oleoyl- ^-D-glucopyranosyl)-3-sitosterol [15], ^-sitosterol-3-O-^-D-glucopyranoside [16].

together with seven known bioactive compounds, 4(aL-rhamnosyloxy)-benzyl isothiocyanate [3], niazimicin [4], 3O-(6'- O-oleoyl--D -glucopyranosyl)- -sitosterol [15], ^-sitosterol-3-O-^-D-glucopyranoside [16], niazirin [12] , ^-sitosterol [14] and glycerol-1-(9-octadecanoate) [13] have been isolated from the ethanol extract of the Moringa seed (Guevara et al., 1999). Figure 1 shows the structures of selected phytochemicals from Moringa.
Copyright 2006 John Wiley & Sons, Ltd.

Lately, interest has been generated in isolating hormones/growth promoters from the leaves of M. oleifera. Nodulation of black-gram (Vigna munga L.)

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Sterol

F. ANWAR ET AL.

Table 2. Sterol composition (grams per 100 g of fatty acids) of the M. oleifera oils
Anwar and Bhanger, 2003 Not reported Not reported 1.49 16.00 Not reported 0.50 19.00 Not reported 1.95 1.00 46.65 0.96 10.70 0.50 Not reported Not reported Lalas and Tsaknis, 2002 0.10 0.05 0.08 15.29 0.33 Not reported 23.06 0.35 1.22 0.64 43.65 Not detected 11.61 0.25 0.39 0.85 Tsaknis e t a l . , 1999 0.13 0.06 0.88 15.13 0.35 Not reported 16.87 0.39 2.52 0.86 50.07 1.11 8.84 1.40 Not reported 0.44

Cholesterol Brassicasterol 24-methylenecholesterol Campesterol Campestanol A7-campestanol Stigmasterol Ergostadienol Clerosterol Stigmastanol ^-sitosterol A7-avenasterol A5-avenasterol 28-isoavenasterol

A7,14 Stigmastadienol A7,14 Stigmastanol

has been shown to increase vigorously with the application of an aqueous-ethanol extract (Bose, 1980) of M. oleifera leaves, although the nature of the active ingredient is still unknown. Moringa leaves act as a good source of natural antioxidant due to the presence of various types of antioxidant compounds such as ascorbic acid, flavonoids, phenolics and carotenoids (Anwar et al., 2005; Makkar and Becker, 1996). The high concentrations of ascorbic acid, oestrogenic substances and -sitosterol [16], iron, calcium, phosphorus, copper, vitamins A, B and C, a-tocopherol, riboflavin, nicotinic acid, folic acid, pyridoxine, -carotene, protein, and in particular essential amino acids such as methionine, cystine, tryptophan and lysine present in Moringa leaves and pods make it a virtually ideal dietary supplement (Makkar and Becker, 1996). The composition of the sterols of Moringa seed oil mainly consists of campesterol, stigmasterol, -sitosterol, A5-avenasterol and clerosterol accompanied by minute amounts of 24-methylenecholesterol, A7-campestanol, stigmastanol and 28-isoavenasterol (Tsaknis et al., 1999; Anwar and Bhanger, 2003; Anwar et al., 2005; Table 2). The sterol composition of the major fractions of Moringa seed oil differs greatly from those of most of the conventional edible oils (Rossell, 1991). The fatty acid composition of M. oleifera seed oil reveals that it falls in the category of high-oleic oils (C18:1, 67.90%-76.00%). Among the other component fatty acids C16:0 (6.04%- 7.80%), C18:0 (4.14%-7.60%), C20:0 (2.76%-4.00%), and C22:0 (5.00%-6.73%) are important (Tsaknis etal., 1999; Anwar and Bhanger, 2003; Anwar et al., 2005). Moringa oleifera is also a good source of different tocopherols (a-, y- and 5-); the concentration of those is reported to be 98.82-134.42, 27.90-93.70, and 48.0071.16 mg/kg, respectively (Anwar and Bhanger, 2003; Tsaknis et al., 1999).

The widespread combination of diuretic along with lipid and blood pressure lowering constituents make this plant highly useful in cardiovascular disorders. Moringa leaf juice is known to have a stabilizing effect on blood pressure (The Wealth of India, 1962; Dahot, 1988). Nitrile, mustard oil glycosides and thiocarbamate glycosides have been isolated from Moringa leaves, which were found to be responsible for the blood pressure lowering effect (Faizi et al., 1994a; 1994b; 1995). Most of these compounds, bearing thiocarbamate, carbamate or nitrile groups, are fully acetylated glycosides, which are very rare in nature (Faizi et al., 1995). Bioassay guided fractionation of the active ethanol extract of Moringa leaves led to the isolation of four pure compounds, niazinin A [1], niazinin [1] B, niazimicin [4] and niazinin A + B which showed a blood pressure lowering effect in rats mediated possibly through a calcium antagonist effect (Gilani et al., 1994a). Another study on the ethanol and aqueous extracts of whole pods and its parts, i.e. coat, pulp and seed revealed that the blood pressure lowering effect of seed was more pronounced with comparable results in both ethanol and water extracts indicating that the activity is widely distributed (Faizi et al., 1998). Activity-directed fractionation of the ethanol extract of pods of M. oleifera has led to the isolation of thiocarbamate and isothiocyanate glycosides which are known to be the hypotensive principles (Faizi et al., 1995). Methyl p- hydroxybenzoate and -sitosterol (14), investigated in the pods of M. oleifera have also shown promising hypotensive activity (Faizi et al., 1998). Moringa roots, leaves, flowers, gum and the aqueous infusion of seeds have been found to possess diuretic activity (Morton, 1991; Caceres et al., 1992) and such diuretic components are likely to play a complementary

MEDICINAL PROPERTIES

USES

AND

PHARMACOLOGICAL

Moringa oleifera also has numerous medicinal uses, which have long been recognized in the Ayurvedic and Unani systems of medicine (Mughal et al., 1999). The medicinal attributes (Table 1) and pharmacological activities ascribed to various parts of Moringa are detailed below.

Antihypertensive, diuretic and cholesterol lowering activities

Copyright 2006 John Wiley & Sons, Ltd.

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role in the overall blood pressure lowering effect of this plant. The crude extract of Moringa leaves has a significant cholesterol lowering action in the serum of high fat diet fed rats which might be attributed to the presence of a bioactive phytoconstituent, i.e. -sitosterol (Ghasi et al., 2000). Moringa fruit has been found to lower the serum cholesterol, phospholipids, triglycerides, low density lipoprotein (LDL), very low density lipoprotein (VLDL) cholesterol to phospholipid ratio, atherogenic index lipid and reduced the lipid profile of liver, heart and aorta in hypercholesteremic rabbits and increased the excretion of fecal cholesterol (Mehta et al., 2003).

Antispasmodic, antiulcer and hepatoprotective activities M. oleifera roots have been reported to possess anti- spasmodic activity (Caceres et al., 1992). Moringa leaves have been extensively studied pharmacologically and it has been found that the ethanol extract and its constituents exhibit antispasmodic effects possibly through calcium channel blockade (Gilani et al., 1992; 1994a; Dangi et al., 2002). The antispasmodic activity of the ethanol extract of M. oleifera leaves has been attributed to the presence of 4-[a-(L-rhamnosyloxy) benzyl]o-methyl thiocarbamate [3] (trans), which forms the basis for its traditional use in diarrhea (Gilani et al., 1992). Moreover, spasmolytic activity exhibited by different constituents provides pharmacological basis for the traditional uses of this plant in gastrointestinal motility disorder (Gilani et al., 1994a). The methanol fraction of M. oleifera leaf extract showed antiulcerogenic and hepatoprotective effects in rats (Pal et al., 1995a). Aqueous leaf extracts also showed antiulcer effect (Pal et al., 1995a) indicating that the antiulcer component is widely distributed in this plant. Moringa roots have also been reported to possess hepatoprotective activity (Ruckmani et al., 1998). The aqueous and alcohol extracts from Moringa flowers were also found to have a significant hepatoprotective effect (Ruckmani et al., 1998), which may be due to the presence of quercetin, a well known flavonoid with hepato- protective activity (Gilani et al., 1997).

4-(a-L-rhamnosyloxy)benzyl carbamate [11] together with 4(a-L-rhamnosyloxy)-benzyl isothiocyanate [3], niazimicin [4] and 3-O-(6'-O-oleoyl-^-D-glucopyranosyl)- -sitosterol [15] have been tested for their potential antitumor promoting activity using an in vitro assay which showed significant inhibitory effects on EpsteinBarr virus-early antigen. Niazimicin has been proposed to be a potent chemopreventive agent in chemical car- cinogenesis (Guevara et al., 1999). The seed extracts have also been found to be effective on hepatic carcinogen metabolizing enzymes, antioxidant parameters and skin papillomagenesis in mice (Bharali et al., 2003). A seed ointment had a similar effect to neomycin against Staphylococcus aureus pyodermia in mice (Caceres and Lopez, 1991). It has been found that niaziminin [9 + 10], a thio- carbamate from the leaves of M. oleifera, exhibits inhibition of tumor-promoter-induced Epstein-Barr virus activation. On the other hand, among the isothiocyanates, naturally occurring 4-[(4'-O-acetyl-a-i-rhamnosyloxy) benzyl] [2], significantly inhibited tumor-promoter- induced Epstein-Barr virus activation, suggesting that the isothiocyano group is a critical structural factor for activity (Murakami et al., 1998).

Other diverse activities Moringa oleifera has also been reported to exhibit other diverse activities. Aqueous leaf extracts regulate thyroid hormone and can be used to treat hyperthyroidism and exhibit an antioxidant effect (Pal et al., 1995a; 1995b; Tahiliani and Kar, 2000). A methanol extract of M. oleifera leaves conferred significant radiation protection to the bone marrow chromosomes in mice (Rao et al., 2001). Moringa leaves are effective for the regulation of thyroid hormone status (Tahiliani and Kar, 2000). A recent report showed that M. oleifera leaf may be applicable as a prophylactic or therapeutic anti-HSV (Herpes simplex virus type 1) medicine and may be effective against the acyclovir-resistant variant (Lipipun et al., 2003). Table 1 depicts some common medicinal uses of different parts of this plant. The flowers and leaves also are considered to be of high medicinal value with anthelmintic activity (Bhattacharya et al., 1982). An infusion of leaf juice was shown to reduce glucose levels in rabbits (Makonnen et al., 1997). Moringa oleifera is coming to the forefront as a result of scientific evidence that Moringa is an important source of naturally occurring phytochemicals and this provides a basis for future viable developments. Different parts of M. oleifera are also incorporated in various marketed health formulations, such as Rumalaya and Septilin (the Himalaya Drug Company, Bangalore, India), Orthoherb (Walter Bushnell Ltd, Mumbai, India), Kupid Fort (Pharma Products Pvt. Ltd, Thayavur, India) and Livospin (Herbals APS Pvt. Ltd, Patna, India), which are reputed as remedies available for a variety of human health disorders (Mehta et al., 2003). Moringa seeds have specific protein fractions for skin and hair care. Two new active components for the cosmetic industry have been extracted from oil cake. Purisoft consists of peptides of the Moringa seed. It protects the human skin from environmental influences and combats premature skin aging. With dual activity, antipollution and conditioning/strengthening of hair, the M. oleifera seed extract is a globally acceptable innovative solution for hair care (Stussi et al., 2002).

Antibacterial and antifungal activities Moringa roots have antibacterial activity (Rao et al., 1996) and are reported to be rich in antimicrobial agents. These are reported to contain an active antibiotic principle, pterygospermin [8], which has powerful antibacterial and fungicidal effects (Ruckmani et al., 1998). A similar compound is found to be responsible for the antibacterial and fungicidal effects of its flowers (Das et al., 1957). The root extract also possesses antimicrobial activity attributed to the presence of 4-a-L-rhamnosyloxy benzyl isothiocyanate [3] (Eilert et al., 1981). The aglyc- one of deoxy-niazimicine (N-benzyl, S-ethyl thiofor- mate) [7] isolated from the chloroform fraction of an ethanol extract of the root bark was found to be responsible for the antibacterial and antifungal activities (Nikkon et al., 2003). The bark extract has been shown to possess antifungal activity (Bhatnagar et al., 1961), while the juice from the stem bark showed antibacterial effect against Staphylococcus aureus (Mehta et al., 2003). The fresh leaf juice was found to inhibit the growth of microorganisms (Pseudomonas aeruginosa and Staphylococcus aureus), pathogenic to man (Caceres et al., 1991).

Antitumor and anticancer activities Makonnen et al. (1997) found Moringa leaves to be a potential source for antitumor activity. O-EthylCopyright 2006 John Wiley & Sons, Ltd.

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F. ANWAR ET AL.

Moringa seeds as coagulant

Moringa seeds are one of the best natural coagulants discovered so far (Ndabigengesere and Narasiah, 1998). Crushed seeds are a viable replacement of synthetic coagulants (Kalogo et al., 2000). In Sudan, seed crude extract is used instead of alum by rural women to treat the highly turbid Nile water because of a traditional fear of alum causing gastrointestinal disturbances and Alzheimer's disease (Crapper et al., 1973; Miller et al., 1984; Martyn et al., 1989; Muyibi, 1994). Moringa seeds are very effective for high turbidity water and show similar coagulation effects to alum (Muyibi and Evison, 1995b). The coagulation effectiveness of M. oleifera varies depending on the initial turbidity and it has been reported that M. oleifera could reduce turbidity by between 92% and 99% (Muyibi and Evison, 1995b). Moringa seeds also have softening properties in addition to being a pH correctant (alkalinity reduction), as well as exhibiting a natural buffering capacity, which could handle moderately high to high alkaline surface and ground waters. The Moringa seeds can also be used as an antiseptic in the treatment of drinking water (Obioma and Adikwu, 1997). Ongoing research is attempting to characterize and purify the coagulant components of Moringa seeds (Ndabigengesere et al., 1995; Gassenschmidt et al., 1995). It is believed that the seed is an organic natural polymer (Jahn, 1984). The active ingredients are dimeric proteins with a molecular weight of about 1300 Da and an iso-electric point between 10 and 11 (Ndabigengesere et al., 1995). The protein powder is stable and totally soluble in water. Moringa coagulant protein can be extracted by water or salt solution (commonly NaCl). The amount and effectiveness of the coagulant protein from salt and water extraction methods vary significantly. In crude form, the salt extract shows a better coagulation performance than the corresponding water extract (Okuda et al., 1999). This may be explained by the presence of a higher amount of soluble protein due to the salting-in phenomenon. However, purification of the M. oleifera coagulant protein from the crude salt extract may not be technically and economically feasible. The coagulation mechanism of the M. oleifera coagulant protein has been explained in different ways. It has been described as adsorption and charge neutralization (Ndabigengesere et al., 1995; Gassenschmidt et al., 1995) and interparticle bridging (Muyibi and Evison, 1995a). Flocculation by inter-particle bridging is mainly characteristic of high molecular weight polyelectrolytes. Due to the small size of the M. oleifera coagulant protein (6.5-13 kDa), a bridging effect may not be considered as the likely coagulation mechanism. The high positive charge (pI above 10) and small size may suggest that the main destabilization mechanism could be adsorption and charge neutralization.

Moringa seeds as biosorbent

Moringa seeds could be used as a less expensive bio- sorbent for the removal of cadmium (Cd) from aqueous media (Sharma et al., 2006). The aqueous solution of Moringa seed is a heterogeneous complex mixture having various functional groups, mainly low molecular weight organic acids (amino acids). These amino acids have been found to constitute a physiologically active group of binding agents, working even at a low concentration, which because of the ability to interact with metal ions is likely to increase the sorption of metal ions (Brostlap and Schuurmans, 1988). The proteineous amino acids have a variety of structurally related pH dependent properties, generating a negatively charged atmosphere and play an important role in the binding of metals (Sharma et al., 2006).

FUTURE PROSPECTS So far numerous studies have been conducted on different parts of M. oleifera, but there is a dire need to isolate and identify new compounds from different parts of the tree, which have possible antitumor promoters as well as inhibitory properties. Although preliminary studies are under way in different laboratories to use the antispasmodic, antiinflammatory, antihypertensive and diuretic activities of M. oleifera seed, these studies should be extended to humans in view of the edible nature of the plant. Moringa roots and leaves have been used traditionally to treat constipation. Studies to verify these claims need to be carried out in the light of the reported antispasmodic activities, which are contrary to its medicinal use as a gut motility stimulant. Earlier studies on the presence of a combination of spasmogenic and spasmolytic constituents in different plants used for constipation (Gilani et al., 2000; 2005a; Bashir et al., 2006) might be of some guidance in designing experiments in which the presence of antispasmodic constituents at higher doses are explained as a possible mode to offset the side-effects usually associated with high dose of laxative therapy. Similarly, the known species differences in the pharmacological actions of medicinal plants (Ghayur et al., 2005; Ghayur and Gilani, 2006) may also be taken into account when planning studies involving contradictory results. Food plants are considered relatively safe as they are likely to contain synergistic and/or side effect neutralizing combinations of activities (Gilani and Atta-ur- Rahman, 2005). Moringa oleifera, known to be rich in multiple medicinally active chemicals, may be a good candidate to see if it contains effect enhancing and/or side-effects neutralizing combinations. Medicinal plants are relatively rich in their contents of calcium channel blockers (CCBs) which are known to possess a wide variety of pharmacological activities such as antihyper- tensive, hepatoprotective, antiulcer, antiasthmatic, anti- spasmodic and antidiarroeal (Stephens and Rahwan, 1992; Gilani et al., 1994b; 1999; 2005b; Yaeesh et al., 2006; Ghayur et al., 2006) and it remains to be seen whether such activities reported to be present in Moringa oleifera have a direct link with the presence of CCBs. Niazimicin, a potent antitumor promoter in chemical carcinogenesis is present in the seed; its inhibitory mechanism on tumor proliferation can be investigated by isolating more pure samples. The mechanism of action of M. oleifera as prophylactic or therapeutic anti- HSV medicines for the treatment of HSV-1 infection also needs to be examined. The available information on the -, - and y- tocopherol content in samples of various parts of this edible plant is very limited. -Carotene and vitamins A and C present in M. oleifera, serve as an explanation for their mode of action in the induction of antioxidant profiles, however, the exact mechanism is yet to be

Microbial elimination with Moringa seeds Moringa seeds also possess antimicrobial properties (Olsen, 1987; Madsen et al., 1987). Broin et al. (2002) reported that a recombinant protein in the seed is able to flocculate Gram-positive and Gram-negative bacterial cells. In this case, microorganisms can be removed by settling in the same manner as the removal of colloids in properly coagulated and flocculated water (Casey, 1997). On the other hand, the seeds may also act directly upon microorganisms and result in growth inhibition. Antimicrobial peptides are thought to act by disrupting the cell membrane or by inhibiting essential enzymes (Silvestro et al., 2000; Suarez et al., 2003). Sutherland et al. (1990) reported that Moringa seeds could inhibit the replication of bacteriophages. The antimicrobial effects of the seeds are attributed to the compound 4(a-L-rhamnosyloxy) benzyl isothiocynate (Eilert et al., 1981).
Copyright 2006 John Wiley & Sons, Ltd.

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elucidated. -Carotene of M. oleifera leaves exerts a more significant protective activity than silymarin against antitubercular induced toxicity. It would be interesting to see if it also possesses hepatoprotective effect against other commonly used hepatotoxic agents such as CCl4 and galactosamine, which are considered more suitable models and close to human viral hepatitis (Gilani and Janbaz, 1995; Yaeesh et al., 2006). Although Moringa leaves are considered a best protein source, it still has to be shown whether or not this protein source could compete with the more common protein sources in highly productive growing or milk- producing ruminants. Many studies have also been conducted on the performance of Moringa seeds as an alternative coagulant, coagulant aid and in conjunction with alum for treating waste water. Therefore, it is important to identify the active constituents of Moringa seed for a

better understanding of the coagulation mechanism. Reports on the antimicrobial effects of the protein purified from M. oleifera are very rare. Since this plant naturally occurs in varying habitats, it is naive to expect a great magnitude of variation in the concentration and composition of chemical ingredients in different parts of the tree. However, the extent to which the chemical composition varies in populations adapted to varying habitats is not known. Thus, detailed studies are required to examine this aspect. In view of its multiple uses, the M. oleifera plant needs to be widely cultivated in most of the areas where climatic conditions favor its optimum growth. In this way, a maximum yield of its different useable parts could be achieved to derive the maximal amount of commodities of a multifarious nature for the welfare of mankind.

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MORINGA OLEIFERA Von Maydell HJ. 1986. T r e e s a n d S h r u b s o f S a h e l , T h e i r C h a r a c t e r i z a t i o n a n d U s e s . Deutsche Gesellschaft fur Technische Zusammenarbeit, Germany: Eschborn, 334-337. Yaeesh S, Jamal Q, Khan A, Gilani AH. 2006. Studies on hepatoprotective, antispasmodic and calcium antagonist activities of the aqueous-methanol extract of A c h i l l e a m i l l e f o l i u m . P h y t o t h e r R e s 20: 546-551.

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