Major Research Project Completion Report

1.    Title of the Project             :      Chemical and pharmacological analyses of Acmella sp., an indigenous phytomedicine

2.    Principal Investigator         :      K. Lalchhandama

3.   Address of the Institution :      Pachhunga University College, Aizawl 796001, Mizoram

4.    Agency                              :      University Grants Commission, New Delhi

       Approval Letter                  :      F. No. 43-47/2014(SR) of 22/8/2015 [MRP-MAJOR-BIOC-2013-36855]

5.    Date of Implementation     :      01/07/2015

6.    Date of Completion           :      30/06/2018

7.   Total Grants                       :      ₹1,446,000

8.    Project Fellow                    :      P.B. Lalthanpuii, CSIR-UGC NET, GATE, SET

9.    Summary of the work        :

Four species of Acmella were confirmed from Mizoram, namely A. oleracea, A. ciliata, A. ulinigosa and A. vazhachalensis. Biochemical tests indicated that they are chemically rich in flavonoids, saponins, tannins, and phytosterols. Chromatographic fractions of the plant extracts were prepared and bioactive compounds were tested by thin layer chromatography. HPLC analysis indicated the presence of N-alkylamides in the most bioactive fractions. Extract fraction containing the bioactive compounds showed biological activities such as anthelmintic and mosquitocidal. The plant extract at different concentrations such as 0.5, 1, 2, 5, 10 and 20 mg/ml showed concentration-dependent activity on the survival of tapeworms of chicken in vitro. Similar treatment was given with albendazole as reference drug. While the untreated tapeworms could survive for 51 h, the treated tapeworms were killed at earlier times. At the lowest concentration (1 mg/ml) it took 24.5 h to kill, while at the highest (20 mg/ml) it took only 7.7 h to kill all the tapeworms. Statistical analysis using Student’s t-test indicated that the mortality effects are significant (p<0.05) at all concentrations tested. Tapeworm tissues were prepared for histology. Under light microscope, structural damages were seen in the muscle and tegumental layers. At the lowest concentration (12.5 mg/ml) of the plant extract 78.3% of the larvae died, while 91.5% died at the highest concentration (100 mg/ml). Similar results were obtained on the adult mosquitos. Student’s t-test showed significant mosquitocidal effects (p<0.05) at all concentrations tested.
       The total antioxidant activity estimated by phosphomolybdate estimation using ascorbic acid. The reaction depended on 0.6 M sulphuric acid, 28 mM sodium phosphate and 4 mM ammonium molybdate, and the absorbance was measured at 695 nm.  It showed that 1 g of plant extract contains 12.5 mg of ascorbic acid equivalent dry weight of the sample. Total phenolic content using gallic acid was assessed using Folin–Ciocalteu reagent. The absorbance of each solution was taken at 765 nm. 1 g of plant extract was found to contain 1.38 mg of gallic acid equivalent dry weight of the sample. DPPH free radical scavenging assay was used for varying concentrations of the plant extract against. For the standard reference, butylated hydroxytoluene (BHT) was used. Absorbance was measured at 517 nm. The IC50 of extract was 13.773 mg/ml while that of the standard was 28.098 mg/ml. Nitric oxide scavenging assay was tested by using different concentrations of the plant extract and standard were mixed separately with sodium nitroprusside and incubated at 30°C for 2 hours in. The mixture was then reacted with Greiss reagent and the absorbance was measured at 550 nm using. The Inhibition concentration at half concentration (IC50) was was found to be 4.121 while that of the extract was 4.492. The reducing power was determined by using phosphate buffer and potassium ferricyanide. The reaction was stopped by adding trichloroacetic acid and the mixture was centrifuged at 3000 rpm for 10 minutes. The supernatant was mixed with distilled water, and ferric chloride solution and the absorbance was taken at 700 nm. The reducing powers of the extract showed concentration dependent activity just like the standard in all concentrations tested.