This study aimed to determine the antiplaque efficacy of Nisin Z extract mouth rinse (Test) as compared to 0.2% chlorhexidine gluconate mouthrinse (Positive control) and distilled water (Placebo) and to assess minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of Nisin Z against Prevotella intermedia (Pi), Porphyromonas gingivalis (Pg), and Aggregatibacter actinomycetemcomitans (Aa) in vitro.Forty-five individuals were randomly divided into three groups (15 in each group). The baseline plaque scores were brought to 0 with scaling and polishing. They were asked not to use any kind of mechanical oral hygiene aid for 4 days and advised to use mouthwashes A, B, or C (Group A: Nisin Z; Group B: distilled water; Group C: chlorhexidine) for 4 days. Following the 4-day plaque re-growth model, Plaque Index was checked on days 1 and 5.In intergroup comparison, a statistically significant difference was seen between Nisin and placebo groups and chlorhexidine and placebo groups, with Nisin and chlorhexidine showing superior antiplaque activity. There was also statistically significant difference between Nisin and chlorhexidine groups, with chlorhexidine showing superior antiplaque activity. In vitro Nisin showed effective inhibition against all three bacterial strains Pi, Pg, and Aa exhibited the highest sensitivity to Nisin with a MIC of 2.5 μg/ml and MBC of 15 μg/ml.Nisin Z mouthrinse was found to be a potent plaque inhibitor, though less effective than chlorhexidine mouthrinse. However, it can serve as a good natural alternative to the gold standard.