Evaluation of anti-inflammatory, analgesic and TNF-α inhibition (upon RAW 264.7 cell line) followed by selection of extract (leaf and stem) with respect to potency to introduce anti-oral-ulcer model obtained from Olax psittacorum (Lam.) Vahl in addition to GC-MS illustration

Ethnopharmacological relevance: Olax psittacorum (Lam.) Vahl., belongs to family olacaceae claimed as an "Issan folk medicine" portray the ethnomedicinal value like curative property of infection in the urinary tract, analgesic, antipyretic, skin-ulcer, antianemic (bark) as well as food additives (leaves). Research articles have proven the presence of anti-swelling property, laxative action, and antiviral activity against poliovirus moreover the antioxidant property too.

Aim of the experiment: Evaluation of antiulcer property (induced within the oral mucosa) of the extract selected amongst two extracts based upon better property towards the ability of anti-inflammatory and analgesia through the in-vivo model as well as the inhibitory property of TNF-α (cell line RAW 246.7). To justify the presence of activity extracts were introduced for GCMS investigation.

Materials and methods: Methanolic extracts (leaf; LME and stem; SME) were collected through maceration and introduced to carrageenan-induced paw edema to evaluate the anti-inflammatory activity and formalin-induced as well as tail-flick in-vivo models to evaluate the analgesic property. Anti-oral ulcer property was analyzed through the acetic-acid induced in-vivo animal model. The cytotoxicity was performed for the test substances on mouse macrophages and fibroblast cells to find a toxic concentration of test substances and to evaluate its modulatory effect of TNF-α inhibition property against LPS induced toxicity.

Results: As compare to diclofenac (100mg/kg) only LME and SME 200mg/kg dose group have insignificant (P<0.05) difference and P-values are 0.99 and 0.88 respectively. From the overall outcome, it can be concluded that compared to the diclofenac (100 mg/kg) group from 4^th hours onwards LME (200 mg/kg) group was able to sustain the inflammation so similar. According to statistical consideration, LME (200 mg/kg) dose has also shown better results in formalin-induced analgesia as well as tail-flick. Cytotoxicity (CTC₅₀) concentrations of LME and SME are 419.60±4.09 and 230.21±0.79 μg/ml respectively on RAW 246.7 cell line. According to CTC₅₀ the highest dose of LME and SME is 400 and 200 μg/ml concentration chosen to evaluate percentage inhibition of TNF-α as compared to diclofenac 25 μg/ml and 50% inhibition was achieved by LME as well as diclofenac i.e. 51.2±2.6% and 50.3±0.8% instead of SME i.e. 45.2±1.7%. As compared to the negative group on DAY-4, LME 200mg/kg/bw dose shown proper growth of epithelial or mucosal layer which reveals proper healing of the surface of the tongue with no sign of injury. GCMS results also reveal that LME and SME both have Cyclohexasiloxane, dodecamethyl; Hexadecanoic acid, methyl ester responsible for anti-inflammatory and analgesic activity but besides, LME has more 4 compounds responsible for activities these are methyl salicylate; phytol; β-Sitosterol; 9,12,15-Octadecatrienoic acid,2,3-bis[(trimethylsilyl)oxy]propyl ester, (Z, Z, Z).

Conclusion: The overall outcomes of the study encapsulate that LME extract with a dose of 200mg/kg/bw will be a good choice to overcome the above-cited ailments. Further study upon this plant is needed to establish the importance of this plant within the human society through quantitative isolation of the metabolites and their pharmacokinetic study to establish the proper pathway of action.

Keywords: GC-MS; Inflammation; TNF-α; aphthous ulcer; hexadecanoic acid methyl ester; methyl salicylate; nociception.