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Chinnaiah Alagarasan and Ganesan Vijaiyan Siva, Oral administration of ethanolic extract of Delphinium denudatum Wall, and its bio efficacy in Wister albino rats, Int.J.Curr.Biotechnol., 2016, 4(1):1-7. International Journal of Current Biotechnology ISSN: 2321 - 8371 Journal Homepage : http://ijcb.mainspringer.com Oral administration of ethanolic extract of Delphinium denudatum Wall, and its bio effi- cacy in Wister albino rats Chinnaiah Alagarasan and Ganesan Vijaiyan Siva* Department of Biotechnology, University of Madras, Guindy Campus, Chennai - 600 025. Tamilnadu, India. ARTICLE INF O ABSTRACT Article History: Delphinium denudatum. Wall, (Dd) dried root sample were collected from authentic Received 05 January 2016 supplier and was further confirmed by taxonomist and extracted by absolute ethanol Received in revised form 14 January 2016 for oral toxicity studies. Based on this study, conclude that the extract from medicinal Accepted 22 January 2016 herb Delphinium denudatum can be administered at a dose of 1000 mg/kg/BW with- Available online 30 January 2016 out any side effects. Since, the toxicity studies in experimental animals cannot always be totally extrapolated to humans, and a reasonable estimate of the self administered Key words: dose is difficult to make such as that applied during traditional use of this plant, Delphinium denudatum, oral toxicity, additional clinical toxicological evaluations need to be performed to define a safe Animal modeling. dose and protect the population from possible toxic effects of the plant. Introduction Unani system in medicine (Kirthikar and Basu, 1999)8. About 80% of the world population, mainly in the Active components of this extract include alkaloids, developing countries, depends on herbal medicine for terpenoids. Any plant extract to be medically useful as primary health care. These medicines are more culturally therapeutic agent, it should be non-toxic or of low toxicity acceptable, have better compatibility with the human to human cells. Though Delphinium denudatum is widely body and lesser side effects (kamboj, 2000)1. India has a used, limited information is available on the toxicity of rich diversity of medicinal plants and a number of plant this plant extract. This study is designed to determine extracts are used against diseases in various medicinal the toxicity profile of the crude ethanolic extract of applications, e.g. Ayurveda, Unani, and Siddha. However, Delphinium denudatum Wall. (Jadwar) in Albino rats inspite of widespread use, only a few of these plants (Wistar strain). The acute oral toxicity study was have been scientifically explored (Malaya, 2004)2. conducted according to the OECD/OCDE guidelines 423/ 20019, using various concentrations of Delphinium Delphinium denudatum (Ranunculaceae) is a potent denudatum extract. herbal medicine used in the Indian system of medicine, was known to possess diverse biological functions like Materials and Methods antiseptic, anti diuretic, anti-inflammatory, antifungal Chemicals: All the chemicals, solvents and reagents were (G.Vijaiyan Siva et al., 20063, Rahman et al., 1997)4, of analytical reagent grade. sedative, analgesic and as a nerve tonic. Silver nanoparticals synthesized fron Dd root extract exhibits Collection of plant samples antibacterial and mosquito larvicidal activity. (G.Vijaiyan The dried roots of Delphinium denudatum was procured Siva et al., 2014)5 It is a great antidote to aconitum from authentic suppliers from Hyderabad. The identity poisoning. Alkaloid delphinine is an antidote against and confirmed by taxonomist, Center for advance studies muscarine and digitaline (Chatterji et al., 1997)6. The root in botany, University of Madras, Guindy campus, is used in Bashahr for toothache and also as an adulterant Chennai. for aconite (Stewart). Root found beneficial in rheumatism, impotency and syphilis. (Rastogi et al., 1999)7 Combined Ethanolic Extraction with other herbal stimulants, it is used as remedy in cardiac Plant extracts were prepared from dried roots of and cerebral diseases. Dried roots are a popular folk Delphinium denudatum by using the procedure remedy for the treatment of epilepsy in the traditional described by Phongpaichit et al., in 2005 10. The dried plant material was powdered by using mechanical grinder. *Corresponding author. 40 grams of dried Delphinium denudatum was packed Email address: [email protected] well in the soxlets apparatus with the help of filter paper. 1 Int.J.Curr.Biotechnol. Volume 4; Issue 1; Jan, 2016 350 ml of absolute ethanol is taken as the extraction Estimation of AST solvent. The extraction was carried out for 15 cycles at Aspartate transaminase was assayed by the method of 55ºC, the crude ethanolic extract was further concentrated King et al., 1965 11. 1.0 ml of aspartate substrate was by evaporation at 55ºC using a solvent trap. added to 0.1 ml of serum, incubated at 37oC for 1 hr to which 1.0 ml of 2, 4-dinitrophenyl hydrazine reagent was Animals added to arrest the reaction. To the blank tubes, 0.1 ml of Male Wister albino Rats (6–8 weeks old) of body weight double distilled water was added instead of serum and 100-150 g was used for the study and given standard the tubes were incubated for 15 min, followed by addition pellet diet and water ad labium. The control animals are of 10 ml of 0.4 N sodium hydroxide. The absorbance was treated with 0.9% saline, varied concentration of read immediately at 520 nm in a Shimadzu UV Delphinium denudatum extract using distilled water as spectrophotometer and the enzyme activity was the dose vehicle is administered for 21 days. All expressed in µmoles of pyruvate liberated/min/mg experiments performed as per the recommendations of protein. Institution animal ethics committee with 6 animals in each group. Estimation of ALT The activity of Alanine transaminase was assayed by Experimental design the method of (King et al., 1965)11. 1.0 ml of alanine The experimental animals were divided into four groups, substrate was added to 1ml of serum, incubated at 37oC each group comprising of six animals. Group 1 served as for 30 min, to which 1.0 ml of 2, 4-dinitrophenyl hydrazine normal control while those in group 2, 3 and 4 were fed reagent was added to arrest the reaction. To the blank orally with ethanolic extract of Delphinium denudatum tubes, 0.1 ml of double distilled water was added instead at 100, 500 and 1000mg/kg BW, respectively, for 21 days. of serum and the tubes were incubated for 15 min, The body weight changes of control and experimental followed by addition of 10 ml of 0.4 N sodium hydroxide. animals are being monitored every interval. At the end of The absorbance was read immediately at 520 nm in the experimental period, the animals were anesthetized Shimadzu UV spectrophotometer and the enzyme activity with diethyl ether followed by cervical decapitation, blood was expressed in µmoles of pyruvate liberated/min/mg was taken by cardiac puncture, the liver and kidney were protein. removed under ether anesthesia, washed thoroughly in ice-cold physiological saline [0.9% (w/v) NaCl], and Histopathological evaluation weighed. Whole blood was separated in two tubes Histological evaluation was performed in the liver tissues contain absence and presence of EDTA, for separation and a portion of specimen was fixed in 10% formalin and of serum and plasma, respectively, and the plasma and embedded in paraffin wax. Sections were cut at 4 ìm in serum was stored frozen until further analysis. thickness, stained with hematoxylin and eosin (Booran et al., 1990)12 and viewed under light microscope for Hematology: histological changes. The hematological parameters were analyzed using Horiba ABS 80 Diagnostics (ABX Pentra Montpellier, Statistical analysis France). These parameters include red blood cells, white All the grouped data were evaluated for statistical blood cells, neutrophils, monocytes, lymphocytes, significance with SPSS v.10 software. Hypothesis testing eosinophils, basophils and platelets. methods included one way analysis of variance followed by least significant difference test. P values of less than Biochemical parameters 0.05 were considered to indicate statistical significance. Estimation of SGOT, SGPT and LDH All these results were expressed as mean ± S.D for six The Serum glutamic oxaloacetic transaminase (SGOT) and animals in each group. serum glutamic pyruvic transaminase (SGPT) were estimated by UV kinetic method in which both SGOT and Results SGPT were assayed based on enzyme coupled system; The body weight of the animals treated with the extract where keto acid formed by the aminotransaminase reacts at the doses of 100, 500 and 1000 mg/kg BW and the in a system using NADH. The coenzyme is oxidized to control group were appreciably increased throughout the NAD and decrease in absorbance at 340 nm for SGOT experimental period (Fig. 1) No significant changes were malate dehydrogenase (MDH) reduces to malate with observed in the organs body weight ratio of the kidney, simultaneous oxidation of NADH to NAD. The rate of liver, lung, and heart as compared with control animals oxidation of NADH is measured, where as SGPT the (Fig. 2). pyruvate formed in the reaction is converted to lactate by lactate dehydrogenase (LDH). LDH was estimated in The hematological parameters serum samples from each group of experimental animals. The effect of oral administration of the ethanolic extract of Dd of the does investigated on hematological parameter Estimation of ALP and its function indicates in male Wister rats for 21 days. Alkaline phosphatase was assayed by the method of The extract did not significantly alter the level of RBC King et al., 1965 11. The incubation mixture of (final volume but efficiently reduced the level of WBC and its 3.2 ml) contained 1.5 ml of buffer, 1.5 ml of substrate, 0.1 differentials, basophils, monocytes and as well as ml MgCl2 and 0.1 ml serum sample. The tubes were platelets (Table 1).

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