The main aim of the work was to scientifically prove the anti-inflammatory property of the polyphenol-rich fraction of hydro-alcoholic leaf extract of Litsea quinqueflora (Dennst.) Suresh by protein denaturation and free radical scavenging activity. The polyphenol-rich fraction of hydro-alcoholic leaf extract was obtained via acid-alkali hydrolysis, followed by fractionation with chloroform and ethyl acetate. HPTLC profiling of the finally obtained ethyl acetate fraction and consequent derivatisation with aluminium chloride revealed the presence of flavonoids in a more purified form. LCMS/MS analysis tentatively identified the presence of bioactive polyphenolic compounds such as gallocatechin, sinapic acid, pinocembrin, paeonol and umbelliferone in the separated fraction. The polyphenol-rich fraction of hydro-alcoholic extract of leaves showed anti-denaturing activity in heat-induced bovine serum albumin denaturation with an IC50 value of 23.59 µg/ml and was statistically significant at 0.1% level. The antioxidant property of the polyphenol-rich fraction determined by its free radical scavenging ability against DPPH and ABTS showed IC50 values 122.98 and 135.44 µg/ml respectively and was also statistically significant at 0.1% level. Hence, the traditional use of Litsea quinqueflora as an anti-inflammatory agent can be attributed to the presence of polyphenols.