In this study, response surface methodology (RSM) was used to investigate the influence of independent process parameters including water to seed ratio (g/g), temperature (°C), time (min) and ultrasonic intensity (%) on extraction yield of “Barhang-e-kabir”. Chemical composition, monosaccharide composition (using HPAEC-PAD), molecular conformation, molecular weight properties, Surface tension, ζ-potential, particle size distribution, Fourier transform infrared spectroscopy (FTIR), color measurement, total phenol content (TPC), total flavonoid content (TFC), antioxidant activity (AA), antimicrobial and dilute-solution and steady-state behavior were evaluated. The optimum condition to obtain maximum extraction yield (13.1 %) was extraction temperature 70 ᵒC, extraction time 40 min, water to seed ratio of 1:10, ultrasonic power of 90 %. Plantago major gum (PMG) had 89.24% carbohydrate, 4.53% ash, 4.11% moisture and 2.12% protein. Viscometric molecular weight and average molecular weight were found to be 1.13 ×105 g/mol and 9.9 ×105 g/mol, respectively. The intrinsic viscosity of PMG was 12.56 dL/g in deionized water at 25 oC. Steady shear measurement demonstrated that PMG is a shear thinning fluid with high viscosity at low concentration. TPC, TFC and AA (IC50) tests of PMG showed 89.80 ± 1.23 mg GAE g−1dry sample, 123.25 ± 1.32 mg g−1dry sample and 470.45 ± 0.35 µg/ml, respectively. Prevention of the linoleic acid oxidation in the system of s-Carotene-linoleic acid was equal to 32.45 %. The results showed that Streptococcus pyogenes and Pseudomonas aeruginosa the most sensitive and highest resistance strain to PMG, respectively.