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Journal of Ethnopharmacology
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Effect of Moringa oleifera Lam. leaves aqueous extract therapy on hyperglycemic rats
Dolly Jaiswal, Prashant Kumar Rai, Amit Kumar, Shikha Mehta, Geeta Watal ∗
Alternative Therapeutics Unit, Drug Development Division, Medicinal Research Lab, Department of Chemistry, University of Allahabad, Allahabad, U.P. 211 002, India
a r t i c l e
i n f o
a b s t r a c t
Ethnopharmacological relevance: In Indian traditional system of medicine, Moringa oleifera Lam. Syn. Moringa pterygosperma Gaerth (Moringaceae) is commonly used as healing herb to treat diabetes. Aim of the study: The purpose of the present study was to assess the effect of M. oleifera leaves aqueous extract therapy on glycemic control, haemoglobin, total protein, urine sugar, urine protein and body weight. Materials and methods: Variable doses of 100, 200 and 300 mg kg−1 of aqueous extract were administered orally by gavage for evaluating their hypoglycemic and antidiabetic effects on fasting blood glucose (FBG), oral glucose tolerance test (OGTT) and post prandial glucose (PPG) of normal and streptozotocin (STZ) induced sub, mild and severely diabetic rats. Results: The dose of 200 mg kg−1 decreases blood glucose level (BGL) of normal animals by 26.7 and 29.9% during FBG and OGTT studies respectively. In sub and mild diabetic animals the same dose produced a maximum fall of 31.1 and 32.8% respectively, during OGTT. In case of severely diabetic animals FBG and PPG levels were reduced by 69.2 and 51.2% whereas, total protein, body weight and haemoglobin were increased by 11.3, 10.5 and 10.9% respectively after 21 days of treatment. Signiﬁcant reduction was found in urine sugar and urine protein levels from +4 and +2 to nil and trace, respectively. Conclusion: The study validates scientiﬁcally the widely claimed use of M. oleifera as an ethnomedicine to treat diabetes mellitus. © 2009 Elsevier Ireland Ltd. All rights reserved.
Article history: Received 10 October 2007 Received in revised form 2 February 2009 Accepted 19 March 2009 Available online 5 April 2009 Keywords: Antidiabetic Drumstick tree Moringaceae Moringa oleifera
1. Introduction Moringa oleifera Lam. Syn. Moringa pterygosperma Gaerth (family Moringaceae) is commonly known as Drumstick tree, indigenous to Northwest India. Most of the parts of the plant possess antimicrobial activity (Bhavasar et al., 1965; Caceres et al., 1991). They are well known for their pharmacological actions too and are used for the traditional treatment of diabetes mellitus (Bhishagratna, 1991; Sharma, 1981; Babu and Chaudhuri, 2005) hepatotoxicity (Ruckmani et al., 1998), rheumatism, venomous bites and also for cardiac stimulation (Chaudhary and Chopra, 1996). In India M. oleifera is incorporated in various marketed herbal formulations, such as Rumalya and Septilin (The Himalya Drug Company, Bangalore, India), Orthoherb (Water Bush-nell Ltd., Mumbai, India), which are available for various disorders.
Abbreviations: bw, body weight; BGL, blood glucose level; FBG, fasting blood glucose; LD50 , lethal dose50 ; M. oleifera, Moringa oleifera; OGTT, oral glucose tolerance test; STZ, streptozotocin; WHO, World Health Organization. ∗ Corresponding author. Tel.: +91 532 2462125; mobile: +09450587750. E-mail address: firstname.lastname@example.org (G. Watal). 0378-8741/$ – see front matter © 2009 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.jep.2009.03.036
Leaves of M. oleifera are lopped for fodder (Sastri, 1962) and have been used as antiulcer, diuretic, anti-inﬂammatory and for wound healing (Kirtikar and Basu, 1935; Caceres et al., 1992; Udupa et al., 1994; Pal et al., 1995). Ethanolic extract of leaves have shown antifungal activity against a number of dermatophytes (Chuang et al., 2007), whereas methanol extract has a potent CNS depressant action (Pal et al., 1996). The aqueous extract of the leaves has been found to possess antifertility activity (Shukla et al., 1981; Prakash, 1998) and is very useful in regulating the thyroid hormone status in adult Swiss rats (Tahiliani and Kar, 2000). Its leaves are also used as nutritional supplement and growth promoters due to the signiﬁcant presence of protein, Se, P, Ca, ˇ-carotene and ˛-tocopherol (Makkar and Becker, 1996; Freiberger et al., 1998; Nambiar and Seshadri, 2001; Lakshminarayana et al., 2005; Sanchez et al., 2006). N-Benzyl thiocarbamates, N-benzyl carbamates, benzyl nitriles and a benzyl ester isolated from methanol extract of its dried fruit powder has been shown to stimulate signiﬁcantly insulin release from the rodent pancreatic beta cells and have cycloxygenase enzyme and lipid peroxidation inhibitory activities (Francis et al., 2004). Hyperglycemia resulting either due to defective production or action of insulin leads to a number of complications; cardiovascular,
5) to a group of overnight fasted rats (Shanmugasundaram et al. 2. Jaiswal et al. 1981. The leaves were washed thoroughly with distilled water. Assessment of hypoglycemic activity by OGTT in normal healthy rats A different group of twenty four normal rats was divided and treated on the same pattern as mentioned above. After 3 days of streptozotocin administration.5. The Institutional Ethical Committee has approved the study. India... University of Allahabad.3. A voucher specimen no. Urine sugar and urine protein were detected by reagent based Uristix from Bayer Diagnostics and body weight of severely diabetic models was measured every week up to 21 days. crushed and extracted twice with distilled water at temperature 60–70 ◦ C repeatedly. total protein were estimated and body weight. BGLs were further checked. WHO has emphasized strongly on the rational use of traditional and natural indigenous medicines. which was then lyophilized to get powder (yield: 11.5. for treating diabetes mellitus (WHO. Group I served as control received vehicle (distilled water only).1 M citrate buffer (pH 4. but there are no reports on hypoglycemic and antidiabetic actions of its leaves. fruits and stem bark have been reported to have antidiabetic action (Kar et al.. as reference drug (El-Hilaly et al. 1 ﬁlter paper and concentrated in rotavapour under reduced pressure to give a semisolid residue. 2. In Indian ethnotherapeutic system of medicine M. Eighteen rats were divided in to three groups of six rats each. hemoglobin. male albino Wistar rats of same age group and body weight 150–200 g were selected for all the experiments. The levels of blood glucose. 6 and 8 h after administering the extract.7%. FBG was checked initially. total protein. New Delhi. Its effect on haemoglobin. 1990) in blood were also estimated weekly in severely diabetic rats. of extract powder suspended in distilled water. Thus. whereas group II and III were treated with a single dose of 200 mg kg−1 of extract and 2. 200 and 300 mg kg−1 given orally by gavage in normal and diabetic rats by conducting fasting blood glucose (FBG) and oral glucose tolerance test (OGTT) studies. urine sugar were checked regularly. this is the ﬁrst reporting of hypoglycemic and antidiabetic effect of aqueous extract of Moringa oleifera leaves. oleifera (5 kg) were collected from the Botanical garden of University of Allahabad. in the month of March 2007. urine sugar.. 200 and 300 mg kg−1 respectively. India. 2. Allahabad.5. sub. Bhishagratna. FBG was taken initially and then blood samples were collected from tail vein at 2. oleifera have been claimed to possess hypoglycemic effect in Indian traditional system of medicine (Sharma. 2.. Allahabad. mild and severely diabetic rats. 1991. A dose of 2 gkg−1 of glucose was then given orally to all the groups. Group I served as control received vehicle (distilled water only) and animals of group II. oleifera is reported to possess hypoglycemic activity (Sharma.5 mg kg−1 of Glipizide respectively.1. 2. Group V serving as a positive control received a dose of 2. ocular etc (Park. 2000... India) and water ad libitum. The leaves of other species of Moringa stenopetala are used traditionally in Ethiopia for treating diabetes mellitus and had already been explored for their hypoglycemic action (Makonnen et al. w/w) for further exploration. 1961) in serum and Total haemoglobin (Nonfon et al. and used in the experiment. urine protein and body weight were also examined in the severely diabetic models. III and IV respectively. 1991. once a day upto 21 days.D. whereas variable doses of 100. Mehta et al. Botany Department. Assessment of hypoglycemic activity by OGTT in sub diabetic and mild diabetic rats Thirty overnight fasted rats of each diabetic model (sub and mild) were divided into ﬁve groups of six rats each. Thus. 2003). Taxonomist. Experimental animals More than hundred twenty ﬁve. mild and severely diabetic rats since M. 4. Assessment of hypoglycemic activity in normal healthy rats Twenty four normal rats fasted over night were divided in to four groups of six rats each. Total protein (Stricklad et al. Plant material Fresh leaves of M.5 mg kg−1 of a known antidiabetic drug Glipizide. 1997). sub.. fruits and stem bark have been scientiﬁcally examined for their use in hypercholesterolaemia (Ghasi et al.. Materials and methods 2. Group I served as diabetic control received vehicle (distilled water only). 1994).5. depending on their blood glucose levels (BGLs) the animals were divided into three groups sub. 2007). with a dose of 200 mg kg−1 which was identiﬁed as the most effective dose by initial screening.2. Kar et al.4. upto three hours at regular intervals of 1 h each. Bhishagratna. Kar et al. 1981. Animals were fed pellet diet (Pashu Aahar Kendra. However. Animals obtained from National Institute of Communicable Disease.2. Blood and urine samples were collected initially and then weekly upto 21 days. urine protein.5. . 1990). It was considered therefore worthwhile to investigate the aqueous extract of M. Estimation BGL was estimated by glucose oxidase method (Barham and Trinder. Induction of diabetes Diabetes was induced by a single intraperitonial injection of freshly prepared streptozotocin (STZ) at the dose of 55 mg kg−1 in 0.. were housed in polypropylene cages at an ambient temperature of 25–30 ◦ C and 45–55% relative humidity with a 12 h each of dark and light cycle.4. considered as 1. The resulting extract was ﬁltered using Whatman no. 2007) 2. 200 and 300 mg kg−1 of extract were given to group II. 2003). 2. 1972) using standard kit of Bayer Diagnostics India Limited. present study deals with the scientiﬁc validation of glycemic potential of aqueous extract of M. 2. India. 2. and then BGL was taken after 90 min of treatment considered as ‘0’ h value.1. (Singh et al. III and IV received variable doses of 100. The rats were then orally administered with 2 g kg−1 of glucose and their glucose tolerance was studied up to 3 h at regular interval of 1 h each. Assessment of antidiabetic activity in severely diabetic rats Long term study of 21 days was conducted in severely diabetic rats. Its leaves. oleifera leaves in normal. neurological. 2006). 2 and 3 h values. Experimental design Initial screening of the aqueous extract of leaves for evaluating its glycemic potential was done with a range of variable doses of 100. for 48 h. FBG was checked initially and then BGL was taken after 90 min of treatment considered as ‘0’ h value. It was identiﬁed and authenticated by Dr Satya Narain. oleifera leaves for its glycemic potential by evaluating its effect on blood glucose level of normal. AD/428/07 has been submitted. Varanasi. 2003). The antidiabetic effect of the extract was also assessed in severely diabetic models. mild and severely diabetic. / Journal of Ethnopharmacology 123 (2009) 392–396 393 renal. 2003).3..
2 3. 53. each value shown in mean ± S. 2.9% at 3 h after glucose administration.).7 58.5 P < 0. oleifera. P < 0.3b 3.7% in case of sub and 15.1 76.9% was observed with the dose of 100 and 200 mg kg−1 .9 56. Hypoglycemic effect of graded doses of aqueous extract of M.2 2.D.1 and 27.9b 6h 76.1c 3.1 77. Groups/Treatment Doses Blood glucose levels (mgdl−1 ) FBG Gp I (control) Gp II (extract) Gp III (extract) Gp IV (extract) Gp V (glipizide) a b c 2h ± ± ± ± ± 2.3 64.9 DW 100 mg kg−1 200 mg kg−1 300 mg kg−1 2.05 as compared with control.5 59. A signiﬁcant increase of about 21 g after 21 days treatment was observed in body weight of extract treated animals. Two groups of six rats each of both the sex (3 females and 3 males) weighing about 150–200 g were orally administered a dose of ten and ﬁfteen times the most effective dose of aqueous extract of M.9a 4. Though.5 mg kg−1 ). Jaiswal et al.05. After 3 h of glucose administration the fall observed with the dose of 100.6 3.4 69.8.01 compared to the control at the corresponding time.6 78.8 and 30. oleifera leaves on FBG. whereas the fall of 16.7 b 8h 76. 200 and 300 mg kg−1 on OGTT of normal rats. 37.1 and 29.1 50. The fall observed after 7. 1.7a 3. and total protein of severely diabetic rats.7% in FBG after 6 h of oral administration.001 as compared with control. An increase of 10. whereas fall of 21. oleifera leaves on BGL of sub diabetic and mild diabetic rats during OGTT studies.6.4 78.1.1 ± ± ± ± ± 2. P < 0.1 4h 77.5 mg kg−1 78.9 63. LD50 experiment Toxicity of the extract was also studied by LD50 experiment.1 3. 3. Group I: control (distilled water).1 4. Moreover. * P < 0.9 4. faeces and urine were also examined at 2 h and then at 6 h intervals for 24 h.3 2.3 4.6 73. 200 and 300 mg kg−1 was 15.9. The data were analyzed with Graph Pad Prism 4. 51. As far as urine protein and urine sugar levels are concerned amazing results were noticed as both were nil only after 14 days of treatment.01 as compared with control.394 D.3 ± ± ± ± ± 2. oleifera leaves on BGL of normal rats during OGTT.7 and 30. 1 depicts the hypoglycemic effect of a single oral administration with variable doses of 100.9 and 25. even Glipizide.9% in case of mild diabetic rats respectively. oleifera leaves on FBG of normal rats. Statistical analysis The results are expressed as mean ± S. 200 and 300 mg kg−1 were evaluated on glucose tolerance of sub diabetic as well as mild diabetic rats.5. Group V: Glipizide treated (2.3. Effect on fasting blood glucose level of normal healthy rats Table 1 describes the hypoglycemic effect of graded doses of aqueous extract of M.05 or lower was considered to be statistically signiﬁcant.8 70.D.5 ± ± ± ± ± 3. similar results were found with Glipizide treatment yet the aqueous extract of M.5 71. neurologic. The difference showing a P level of 0. respectively.2.0 v for Windows (Graph Pad Software.6 72. Rats were then observed continuously for their gross behavioural.4 2. Group III: extract treated (200 mg kg−1 ). (n = 6). 3. Table 2 demonstrates the effect of graded doses of aqueous extract of M. Food consumption. oleifera was found to be more effective in all the parameters.6c 2. The body weights of all the groups (control and treated) were estimated before the treatment and then weekly during the treatment. CA.8 1.4 74. 32. 3.5.2 67. The dose of 200 mg kg−1 produced a maximum the fall of 29. LD50 No toxic effect was observed on treatment with upto 10 and 15 times the effective dose of the extract as the behavior of the Fig. different doses of aqueous extract 100. Effect on blood glucose level and other parameters of severely diabetic rats Figs.8a 3.4 a 3. 31.9 4.4 3.1 73.1% was observed with the doses of 100 and 300 mg kg−1 respectively. / Journal of Ethnopharmacology 123 (2009) 392–396 Table 1 Effect of graded doses of aqueous extract of Moringa oleifera leaves on BGL of normal rats (mean ± S. Group II: extract treated (100 mg kg−1 ). 2 and 3 reveals the effect of 21 days long treatment with extract of M. USA).5. 3. which was lesser than the fall produced by the most effective dose of the extract.4 ± ± ± ± ± 1. ** P < 0. Rats treated with 200 mg kg−1 showed a maximum fall of 26.1 63.5.7 51. San Diego.2% in FBG and 21. 69. autonomic and toxic effects up to 24 h. Effect on oral glucose tolerance of sub diabetic and mild diabetic rats In order to choose the optimum dose for the severely diabetic animals.8a 2. 2.9. PPG.4.9% in haemoglobin and 11. Group IV: extract treated (300 mg kg−1 ).3% in total protein was observed after 21 days treatment.D.4. 14 and 21 days treatment with the dose of 200 mg kg−1 of the extract was 25.4 57. . Effect on oral glucose tolerance of normal healthy rats Fig. 3.5.2% in PPG levels respectively. the reference drug produced a fall of only 26. haemoglobin. Results 3. and all statistical comparisons were made by means of one-way analysis of variance (ANOVA) followed by Newman–Keuls Multiple Comparison Test.5% in sub diabetic and mild diabetic cases respectively.
01 as compared with control.7 ± 5.7 and 25.4 85.9 ± 5. * P < 0.1 ± 4.). it is likely to be expected that the aqueous extract of leaves has some direct effect by increasing the tissue utilization of glucose (Ali et al.9 338.1c 87.6 88.6 ± 5. urine sugar.8 ± 4.3 ± 5.1 ± 3.3 ± 4.8 180. Effect of aqueous extract of Moringa oleifera leaves on Haemoglobin and total protein in severely diabetic rats Group I: control (distilled water).6 ± 5.5 mg kg−1 ).D.5 81.3b 105. 2004). It is interesting to note that the extract was more effective than reference drug.3 ± 4.7 ± 4. 2. P < 0. Fig.5 mg kg−1 ).8a 190.1 190.9 104.7 ± 4. The daily treatment with 200 mg kg−1 of extract for 21 days brought FBG and PPG levels to nearly normal range in severely diabetic animals. 300 mg kg−1 ) V (glipizide. Glipizide was used as reference drug in diabetic models for positive control.5 ± 5.induced diabetes is characterized by severe loss in body weight (Ravi et al. Effect of aqueous extract of Moringa oleifera leaves on FBG and PPG in severely diabetic rats Group I: control (distilled water). 200 mg kg−1 ) IV (extract. Signiﬁcant improvement in haemoglobin and total protein levels on long term treatment with extract for 21 days indicated that it has favourable effect in bringing down the severity of diabetes.5 mg kg−1 ) a b c 395 FBG 0h −1 1h 217.1a 103.5b 242.2 223. indirectly conﬁrming thereby the antidiabetic activity of the extract.6b 88. Groups (treatment and doses) I (control.5 ± 4.1 ± 4.6a 177.6 190. * P < 0... sub and mild diabetic animals.1% after 6 h.7 ± 3.3a 186..4a 238. STZ. 4. the weight gain after administration of the extract in severely diabetic rats is simply due to the Fig. Usually elevated levels of urine sugar and urine protein are associated with diabetes mellitus and complete elimination of these within 14 days from urine of severely diabetic animals through extract therapy. 2007. oleifera leaves aqueous extract therapy on glycemic control.4 121. P < 0. Hence. 2.9 79.2 ± 4..3 167.3 170.3a 183.6 ± 3. ** P < 0.2 2h 139.2 ± 4. / Journal of Ethnopharmacology 123 (2009) 392–396 Table 2 Effect of variable doses of Moringa oleifera on OGTT of sub diabetic and mild diabetic rats (mean ± S.2 ± 3. 200 mg kg−1 ) IV (extract..1 ± 4.1 ± 4. Gray et al. Group II: extract treated (300 mg kg−1 ). Group III: Glipizide treated (2.8 200.4 ± 4..8 ± 4.9 ± 5.2 ± 3.1 ± 4.8 189.5a 300.001 as compared with control. Trichosanthes dioica (Rai et al.01 compared to the control at the corresponding time. 2. 100 mg kg−1 ) III (extract. 1993.3b 264. 1969).5a 182.2 84. D W) II (extract. Group II: extract treated (300 mg kg−1 )..8b BGL of sub diabetic animals (mg dl ) 89. 100 mg kg−1 ) III (extract. 300 mg kg−1 ) V (glipizide.7 89.9 ± 5.8 ± 5. Jaiswal et al. urine protein and body weight. is an additional advantage.6a 85. 3. The purpose of the present study was to assess the effect of M.7 ± 4.5 ± 3.9 ± 5.5 BGL of mild diabetic animals (mg dl−1 ) 189. Aqueous extract of M. .4 ± 4. 2008) and Cinnamomum tamala (Sharma et al. by inhibiting hepatic gluconeogenesis or absorption of glucose into the muscles and adipose tissues (Kamanyi et al. 1994)..D.8b 240.7 ± 3. Cynodon dactylon (Singh et al. oleifera leaves reduces the blood glucose level in normal rats and normalizes the high blood glucose levels in sub.3 ± 4.1 188.7b 104.05.5 86..2 ± 4.05.9 ± 4. good activity has been seen in severely diabetic rats with damaged islets therefore.5 168.4 ± 4. However. 1996).2 ± 3. haemoglobin.7 ± 5.5 ± 4.6 ± 5.6 ± 5. Group III: Glipizide treated (2.9 ± 5. 2000).3a 307. D W) II (extract.4c 182.9a 375.7 ± 4.5 277.8a 87. It also improves glucose tolerance in normal. mild and severely diabetic rats. 2008).3 ± 4. Discussion STZ-induced hyperglycemia has been described as a useful experimental model to study the activity of hypoglycemic agents (Junod et al. The FBG decreases by 26. the higher concentration of the extract used.1 88.5 191. in normal rats treated with a single dose of 200 mg kg−1 and 300 mg kg−1 respectively.8b 318.0 P < 0. 2007).01 compared to the control at the corresponding time. treated rats appeared normal and no death occurred in any of these groups.2 ± 4.05 as compared with control.2 ± 4. ** P < 0.4 ± 4. Since. total protein.5 ± 5.6a 3h 124.9b 229. Such a phenomenon of less hypoglycemic response at higher doses is common with indigenous plants and has already been observed in Psidium guajava (Rai et al. against the reference drug Glipizide could be because of only a small amount of active substance present in the extract.5 mg kg−1 ) I (control.
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