Research Article

Safety Profile for Evaluation of Berberine in Sprague- Dawley Rats: Effects on Renal, Liver and Myelopoietic Functions

Attia Anwar1, Sajida Malik2*, Zobia Usman1 and Sadia Chiragh3

1Assistant Professor of Pharmacology, Faisalabad Medical University, Faisalabad; 2Associate Professor of Pharmacology, Allama Iqbal Medical College, Lahore; 3Professor of Pharmacology, Post Graduate Medical Institute (PGMI), Lahore.

Received | July 08, 2017; Accepted | November 26, 2017; Published | December 24, 2017

*Correspondence | Dr. Sajida Malik, Associate Professor of Pharmacology, Allama Iqbal Medical College, Lahore; Email: [email protected]

Citation | Anwar, A., S. Malik, Z. Usman and S. Chiragh. 2017. Safety profile for evaluation of berberine in sprague- dawley rats: effects on renal, liver and myelopoietic functions. Annals of King Edward Medical University, 23(4): 492-495.

DOI | http://dx.doi.org/10.21649/journal.akemu/2017/23.4.492.495

Keywords | Berberine, SPRAGUE-Dawley rats, Hepato-toxicity

Introduction

Berberine, an isoquinolone derivative alkaloid, is present in roots, rhizome and stem bark of many plants including Berbris equifolium, Berberis aristata, Berberis vulgaris and Hydratis canadensis. It has been widely used in traditional Chinese medicines for hundreds of years(1). Modern research has shown that berberine and its derivatives display several pharmacological effects through various mechanisms(2). Its properties include anti-microbial and anti-protozoal effects(3), anti-diabetic effects(4), anti-lipidemic effects and hence beneficial effects on cardiovascular system(5). In ayurvedic medicine, Berberis aristata and berberine is being used for prevention and treatment of cancer(6). In a recently conducted research, berberine delayed and prevented development of 7, 12Dimethylbenz (α) anthracine induced mammary carcinogenesis in rats with improvement in histopathologic parameters(7).

Berberine is an over-the-counter (OTC) drug and is used for a wide variety of diseases in China. Despite its widespread use in ayurvedic and Chinese traditional medicine, limited studies are conducted on untoward effects and safety profile of the drug. The present study was designed to evaluate the effects of berberine on various organ functions including liver functions, renal functions and myelopoietic system in Sprague-Dawley rats.

Materials and Methods

The study was carried out in the Department of Pharmacology, PGMI, Lahore during 2014. Healthy female Sprague-Dawley rats of 6 weeks age were purchased from University of Veterinary and Animal Sciences, Lahore, Pakistan. Animals were kept in animal house of PGMI for 1 week for acclimatization. They were fed on water and rat chow ad libitumand kept in iron cages at optimum room temperature according to criteria mentioned in guide for the care and use of laboratory animals(8). After 1 week, rats were divided in two groups, A and B carrying 10 animals each. The body weight was recorded in grams.Berberine hydrochloride was purchased from Sigma Aldrich. For administration, 3 g of berberine was suspended in distilled water to make volume 80 ml,producing concentration of 75 mg berberine/2 ml(9). Suspension was given to Group-B in a dose of 75 mg/kg daily by gavage for a period of 24 weeks as a morning dose whereas Group-A was given distilled water by similar route for same duration.

After 24 weeks, rats were anesthetized and blood samples collected through cardiac puncture. For CBC, blood was collected in EDTA vials. The vials were then put in roller mixer and attached to Sysmex KX-21 to get CBC reports. For alanineaminotranferase, alkaline phosphatase, urea and creatinine, clot activatedsamples were collected in vialsand centrifuged to separate serum. The serum was then put in Spectra fully automated machine to obtain results using commercially available kits.

Statistical analysis

All group data were evaluated statistically with SPSS version 20.All values were expressed as mean ± SD. Testing methods included unpaired-t test. The p-value ˂ 0.05 was considered significant.

Results and Discussion

The levels of hemoglobin and blood cells counts in control and experimental groups of rats are shown in Table 1. It is evident that there were no statistically significant differences in mean hemoglobin, and mean platelets counts in two groups. However, significant difference was observed in mean WBC and RBC counts. There was decrease in WBC count but increase in RBC count in berberine treated group.

Table 1: Effect of berberine on hemoglobin and blood cell count of female Sprague Dawley rats.

Table 2: Effect of berberine on renal function, liver enzymes and uric acid level of female Sprague Dawley rats.

Results shown in Table 2 indicate that the comparison of liver enzymes, renal function and serum uric acid levels of the rats in two groups. No statistically significant differences were seen in mean blood urea, serum creatinine, alkaline phosphatase and SGPT levels. However, there was a significant decrease in serum uric acid level in berberine treated group.

Berberine is a bitter tasting yellow plant alkaloid used for long time in Chinese and ayurvedic medicine. Several clinical trials have been conducted using berberine for various conditions in animal as well as humans. Before the present study, no information describing the toxic effects of berberine compounds on liver, kidneys and blood cells of laboratory animals was found in the literature. However, regarding developmental anomalies in rats and mice, one study has been published(10). Present study was designed with a view to observe any toxic effects of berberine on various organ systems in experimental rats.

No adverse clinical signs were observed during the treatment or the trial period. Table 1 compares the blood cell counts of control and berberine treated groups. No significant differences were found in hemoglobin level and platelet counts in the two groups. The compound has rather previously been tested for effects on platelet count and platelet functions in patients of primary and secondary thrombocytopenia showing improvement in both(11).

The mean WBC count in control group was 8475±1089 and in experimental group, it was 5820±1522 per mm3. Although this relative neutropenia observed had no apparent clinical effects but the difference is statistically significant (p=0.002). This may be attributed to a non-specific response of berberine. Leucopenia has been observed with use of some herbs like Echinacea (published as a case report) and was considered to be a reversible allergic response(12). This may be a possible explanation in case of berberine but it requires further investigation. Significantly high RBC count was observed in berberine treated rats. This effect on RBC count with concomitant decrease in WBC count suggests stimulation of one progenitor cell type in bone marrow with simultaneous suppression of other. Additional data as red cell volume and differential white cell count may have been helpful in providing an explanation.

Table 2 shows the comparison of hepatic enzymes and renal function tests of the two groups. The mean blood urea levels were 34.4 ± 4.3 and 33.0 ± 4.69 mg/dl in control and treatment groups respectively. The mean serum creatinine levels were 0.50 ± 0.24 and 0.55 ± 0.14 mg/dl respectively. These differences were not significant thus ruling out any possible nephro-toxicity of berberine in normal Spargue-Dawley rats. In one study, the drug showed nephro-protective role in streptozotocin induced diabetic rats(13). Moreover, berberine wasfound to have protecting effect on hypertensive renal impairment model rats, fed with high fat-salt-fructose diet associated with elevated antioxidant capability in body and kidney tissues(14).

Regarding hepatic enzymes, the mean SGPT was 29.1 ± 6.21 and 30.4 ± 5.4 U/l in control and treated groups respectively. Similarly, alkaline phosphatase was 206.2 ± 93.5 and 244.4 ± 43.2 U/L. These differences were not significant either, ruling out any possible hepato-toxicity. In a previously reported study, berberineshowed hepato-protective activity against CCl4 induced hepato-toxicity. The activity was both preventive and curative (15). In another study, it was concluded that berberine not only reduced elevated levels of ALT and AST, but also showed improvement in histo-pathological changes of necrosis and fibroblast proliferation in livers of rodents in which liver toxicity was induced(16).

An inhibitory effect on serum uric acid levels was observed in berberine treated group. Chang et al found an inhibitory effect of 12 phenolics including berberine on xanthine oxidase activity in experimental mice. This xanthine oxidase inhibition may be a possible explanation for reduced uric acid levels in the current study(17).

The above-mentioned results of the present study establish safety of berberine use in normal Sprague Dawley rats. Rather, a beneficial effect on serum uric acid level was observed for a possible therapeutic usage for hyperuricemia in human subjects. Berberine may also be helpful in elevated uric acid levels associated with cancers. However, further investigation in human volunteers is required to see the safety for long term human use.

Conclusions

The results of the present study showed that berberine at the dose of 75 mg/kg body weight/day for 24 weeks in Sprague- Dawley rats possesses no clinical and biochemical evidence of effects on liver and renal functions. However, a relative neutropenia was observed which provides guidance for further studies to determine the mechanisms for berberine effects.

Authors Contribution

Attia Anwar: Principal investigator topic selection and data analysis and interpretation.

Sajida Malik: Co-investigators and data collection.

Zobia Usman: Co-investigators and data collection.

Sadia Chiragh: Co-investigators and data collection.

References

1. 1. Derosa G, Mafiolla P. Alkaloids in nature: pharmacological application in clinical practice of berberine and Mate tree. Current Topics in Medicinal Chemistry. 2014; 14 (2):200-206. https://doi.org/10.2174/1568026613666131213155252
2. 2. Jin Y, Khadka DB, Cho WJ. Pharmacological effects of berberine and its derivatives: a patent update. J Expert opinion On Therpeutic Patents. 2016; 26(2):229-243. https://doi.org/10.1517/13543776.2016.1118060
3. 3. Vuddanda PR, Chakraborty S, Singh S. Berberine: A potential phytochemical with multispectrum therapeutic activities. J Expert opinion On Therpeutic Patents. 2010; 19 (10): 1297-1307. https://doi.org/10.1517/13543784.2010.517745
4. 4. Cicero AFG, Tartaghni E. Antidiabetic properties of berberine: from cellular pharmacology to clinical effects. J Hospital Practice. 2012; 40 (2):56-63. https://doi.org/10.3810/hp.2012.04.970
5. 5. Derosa G, Maffiali P, Cicerio AFG. Berberine on metabolic and cardiovascular risk factors: An analysis from preclinical evidence to clinical trials. J Expert Opinion on Biological Therapies. 2012; 12 (8):1113-1124. https://doi.org/10.1517/14712598.2012.704014
6. 6. Garodia P, Ichikawa H, Malani N, Sethi G, Agarwal BB. From ancient medicine to modern medicine: ayurvedic concepts of health and their role in inflammation and cancer. J Soc Integr Oncol 2007; 5, (1): 25-35. https://doi.org/10.2310/7200.2006.029
7. 7. Attia A, Kashif B, Farva S, Saadia C. Effects of berberine on 7, 12 Dimethylbenz (α) Anthracine induced mammary carcinogenesis in rats. APMC. 2017; 11(1):39-43.
8. 8. Guide for care and use of laboratory animals 8th ed. Washington DC: National academies press; 2013.
9. 9. Manoharan S, Sindhu G, Vinothkumar V, Kowsalya R. Berberine prevents DBMA induced Hamster buccal pouch carcinogenesis: A biochemical approach. Eu J of Cancer Prevention. 2012; 21 (2):182-192. https://doi.org/10.1097/CEJ.0b013e32834c9c3c
10. 10. Jahanke DG, Price CJ, Marr MC, Myers CB, George JD. Developmental Toxicity Evaluatio of Berberine in Rats and Mice. Birth defects research (part B). 2006; 77:195-206. DOI 101002/bdrb.
11. 11. Chekalina SI, Umerzakova RZ, Abdur-rakhmanov TR. Effects of berberine bisulphate on platelet hemostasis in thrombocytopenia patients. Gematol Transfuzini. 1994; 39, (5): 33-35.
12. 12. Kemp DE, Franco KN. Possible Neutropenia associated with long-term use of Echinacea. JABFP. 2002; 15, (5): 417-19.
13. 13. Wei hua LIU, Zei qing HIE et al. Berberine ameliorates renal injury in Streptozotosin induced diabetic rats by suppression of both oxidative stress and aldose reductase. Chin Med J. 2008; 121 (8): 706-12.
14. 14. Effects and mechanism of berberine on thehypertensive renal injury rats induced by high fat-salt-fructose enriched diet. Zhong Yao Cai. 2011; 34 (3): 412-14.
15. 15. Feng Y, Siu KY, Ye X, Kobayashi S. Hepatoprotective effects of berberine on carbon tetra chloride induced acute hepatotoxicity in rats. Chinese Medicine. 2010; 5(1); 33-38. https://doi.org/10.1186/1749-8546-5-33
16. 16. Zhang JB, Xu D, Guo Y, Ping J, Chen LB, Wang H. Protection by and anti-oxidant mechanism of berberine against rat liver fibrosis induced by multiple hepatotoxic factors. CEPP. 2008; 35 (3):303-309. https://doi.org/10.1111/j.1440-1681.2007.04819.x
17. 17. Chang WS, Chang YH, Lu FJ, Chang HC. Inhibitory effects of phenolics on xanthine oxidase. Anticancer research. 1994; 14, (2A):501-506.