Osmotic Stress Induced Hemolytic Activity Biology Essay


Preliminary phytochemical screening of the various Ficus extract revealed the presence of triterpenoids, flavonoids, glycosides, steroids, alkaloids, saponins.

The peroxyl radical generated by AAPH on addition to erythrocyte suspension and its subsequent scavenging action produced by graded higher concentration of the extract are given in Table 2 .The results of the study were compared with the standard ascorbic acid and an increase in inhibition was noticed for all the extracts tested. FBBE exhibited significant IC 50 value (171.1± 0.52 µg/ml) compared to other extracts.

5.3. Hydrogen peroxide induced hemolysis

Lipid oxidation of erythrocyte membrane mediated by H2O2 induces membrane damage and subsequently hemolysis. The antihemolytic activity of the various extracts on erythrocytes is presented in Table 3. All the sample extracts were found to have antihemolytic activity and the values are considerably different. Interestingly, FBBE registered the highest antihemolytic activity. The antihemolytic activity of various extracts and standards are in order of ascorbic acid > FBBE > FGBE > FMBE > FRBE.

5.4. Osmotic stress induced hemolytic activity

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Hemolysis of human erythrocytes was induced by hypotonic stress. The antihemolytic activity of FBBE, FMBE, FRBE and FGBE on human erythrocytes is presented in the table 4. All the four extract have antihemolytic activity. The antihemolytic action for FBBE, FGBE, FMBE and FRBE were 164.10, 222.80, 242.30 and 234.10 µg/ml respectively. Ascorbic acid expressed the highest activity(IC 50 78.6 µg/ml)

5.5. Total anti oxidant power assay (Phosphomolybdate method)

The phosphomolybdenum method is quantitative, and the total antioxidant capacity is expressed as α Tocopherol equivalents. Among the extracts tested, the FBBE contains 10.90/ α Tocopherol equivalent /100 µg. The antioxidant activity increased in the order of FBBE> FGBE> FMBE >FRBE. (Table 12; Fig 3).

5.6. Ferric reducing power assay

Table 5 shows the reductive capabilities of methanolic extract of four different ficus species when compared to the standard, BHT. Like the anti oxidant activity, the reducing power increased considerably with increasing amount of extracts. The FBBM showed the highest reducing power ability than the other extracts tested. However, the activity of the various extracts was less than the standard. The order of reducing ability of the extracts are FBBE>FGBE>FRBE>FMBE.

5.7. Metal chelating capacity

Addition of the extracts of ficus species interfered with the ferrous-ferrozine complex and this was decreased with the increasing concentration of the drug. Among the extracted tested, the FBBM showed more pronounced ferrous chelating ability than those of other extracts(IC 50 166.8 µg/ml).The chelating action for FGBE, FMBE and FRBE were 183.6, 244.1 and 285.1 µg/ml respectively. EDTA expressed the highest ferrous ion chelating ability(IC 50 33.9 µg/ml) (Table 6).

5.8. Total peroxy radical trapping potential

The peroxy radical scavenging activity was determined for four ficus species and results were compared using α Tocopherol synthetic analogue Trolox (Table 7). Addition of increasing concentration of fractions to the solution containing AAPH decreased the luminescence produced by DCF and absorbance decreased in a linear fashion .Only FBBM exhibited significant activity with IC 50 of 166.7 µg/ml

5.9. Total flavonoid content assay

The total flavonoid content in the extracts of ficus was expressed as µg quercetin equivalent .The flavonoid content was higher for FBBE (16.9 µg/mg) and for other extracts it was found to be FGBE (16 µg/mg), FRBE (4.5 µg/mg), and FMBE (7.2 µg/mg) quercetin equivalent (Fig 14; Table 12)

5.10. Total phenolic content assay

The total phenolics content in the extracts of FBBE, FGBE, FRBE and FMBE were determined using Folin-Ciocalteu assay (Fig.15) and expressed as µg pyrocatechol equivalents (PCE).The quantity of total phenolics in the extract linked with the antioxidant activity. Among all the extract the significantly higher phenolic content amount was found in FBBE (17.8µg/mg).The phenolic content of other fractions were FGBE (13.38µg/mg), FRBE (4.68µg/mg), and FMBE (9.28µg/mg) respectively (Fig 15; Table 12).

5.11. Nitric oxide scavenging assay

Prepare solutions of sodium nitroprusside in phosphate buffered saline and incubate at 25 0c for 150 min resulted in production of nitric oxide. All the extracts effectively reduced the generation of nitric oxide radicals. The FGBE exhibited superior nitric oxide scavenging activity action with IC 50 103.7µg/ml which was lower than the standard ascorbic acid (46.8µg/ml).The IC 50 of FBBE, FMBE, FRBE were 105.1, 360.6 and 233.8 µg/ml and scavenging activity was lesser than that of standard (Table 8).

5.12. Hydroxyl radical scavenging assay

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The degradation of deoxyribose by Fe 3+ - ascorbate -ethylene diamine tetra acetic acid - hydrogen peroxide system was markedly decreased by the all extracts of ficus tested (Table 9) indicating the potential hydroxyl radical scavenging ability. The inhibitory concentration (IC 50) were higher for FGBE (24.23 µg/ml) than the other extracts employed in the study.

5.13 . Super oxide anion radical scavenging assay

The extracts of four ficus species were established as scavenger of superoxide anion in NBT reduction system (Table 10). All the extracts showed potential superoxide inhibiting activity at a concentration range of 25-400 µg/ml. The IC 50 of FBBE, FGBE, FMBE and FRBE were found to be 138.2, 183.4, 205.6 and 266.8 µg/ml respectively.

5.14. Hydrogen peroxide scavenging assay

The methanolic extracts four ficus species were scavenged hydrogen peroxide in a concentration dependent manner which could be seen by its graded increase in percentage inhibition (Table 11).

5.15. Antimicrobial activity.

The results of antimicrobial assay of four ficus species under study by agar well diffusion assay as shown on the figure 12-24.the antimicrobial activity was measured by zone of inhibition (mm).The results of antimicrobial assay showed that methanolic extract of four ficus species have potential antimicrobial activity. FBBM have considerable activity against S.aureus, P.aeruginosa and E. coli (21mm, 25mm and 20mm at 4mg conc) compared to FGBE ,FMBE and FRBE. However the activity of FBBE, FMBE, FRBE and FGBE were less than the standard ciprofloxacin.