Integrated In Vivo, In Vitro, and In Silico Evaluation of Spermacoce ocymoides Methanolic Extract for Antidiarrheal, Thrombolytic, and Antiarthritic Activities.

The study investigated the traditional use of Spermacoce ocymoides as an antidiarrheal, thrombolytic, and antiarthritic agent through pharmacological evaluation, phytochemical profiling, and comprehensive computer-aided drug design (CADD). The crude methanol extract of the plant was chemically profiled using GC-MS, which identified nine compounds, including 3,5-ditert-butylphenol, 1-tetradecene, 3,5-ditert-butylbenzene-1,2-diol, m-fluorothiophenol, methyl triacontanoate, and others. Qualitative analysis of phytochemicals ascertained the existence of phenols, alkaloids, flavonoids, terpenoids, saponins, and tannins. The quantitative phytochemical screening revealed that the total flavonoid and phenolic contents of the crude extract were 10.94 ± 0.27 mg QRE/g and 39.87 ± 0.33 mg GAE/g, respectively, as quantified using the aluminum chloride colorimetric and Folin–Ciocalteu reagent techniques. The in vivo antidiarrheal potential of the crude extract and its fractions was evaluated at four doses using the castor oil-induced diarrheal test. The present findings revealed that the crude extract prevented the defecation frequency by 84.2% (p < 0.001) at 500 mg/kg doses, which is significantly closer to the reference drug loperamide (89.5% inhibition) compared to the negative control group. The thrombolytic activity of the crude extract was screened using an in vitro clot lysis assay, revealing a significant clot lysis potential of 38.7%, compared to 67.2% clot lysis observed with the standard drug streptokinase. A protein denaturation assay was next performed to evaluate the in vitro antiarthritic potential of the crude extract, which showed significant protection (74.5%) against protein denaturation compared to the diclofenac sodium, with 95.9% inhibition at a concentration of 1.0 mg/mL. Molecular docking of compounds identified through GC-MS revealed excellent binding scores for most of the compounds, particularly 3,5-ditert-butylbenzene-1,2-diol and 3,5-ditert-butylphenol, regarding the active interacting sites of selected target receptors when compared to the standard drugs. Molecular orbital analysis revealed that most of the identified compounds exhibited a low energy gap between HOMO and LUMO, indicating high chemical reactivity. FT-IR analysis documented that carboxylic -OH, C = C, C = O, -SH, and -CH bonds are mainly present in identified compounds of the crude extract. Moreover, most compounds demonstrated promising profiles in ADMET and PASS prediction. This study represents the first investigation into this plant’s antidiarrheal, thrombolytic, and antiarthritic activities. In conclusion, the methanolic extract of S. ocymoides was observed to be abundant in different bioactive phytochemicals, which may contribute to its medicinal potential, and this investigation provides a significant contribution to validating the traditional use of S. ocymoides in diarrhea, thrombosis, and arthritis treatment.