The relationship between the antibacterial activity of Nisin and pH value

As a natural polypeptide antibacterial agent produced by lactic acid bacteria, the antibacterial activity of nisin is highly sensitive to environmental pH. This sensitivity mainly stems from the regulation of pH on the structural stability, solubility, and target affinity of Nisin molecules. The specific mechanism and rules can be analyzed from the following aspects:

1. Impact on Molecular Structural Stability

The nisin molecule contains multiple special cyclic structures such as lanthionine and β-methyl lanthionine, which are the core for maintaining its antibacterial activity—the integrity of these cyclic structures directly determines whether nisin can bind to bacterial targets and exert its effect. In an acidic pH environment (usually pH 2.0–6.0), the amino groups (-NH₂) in the Nisin molecule undergo protonation to form positively charged amino groups (-NH₃⁺). This protonated state significantly enhances the intramolecular hydrogen bonding and hydrophobic interactions, thereby stabilizing its unique cyclic spatial conformation and preventing the loss of antibacterial activity caused by structural unfolding or breakage. However, when the pH increases to the neutral or alkaline range (pH ≥7.0), the hydroxyl groups (-OH⁻) in the environment neutralize the protonated amino groups of nisin molecules, resulting in reduced positive charge on the molecules, weakened hydrogen bonding and hydrophobic interactions, and easy unfolding or degradation of the cyclic structures. For example, at pH above 8.0, peptide bond hydrolysis may occur in Nisin molecules, generating small molecular fragments with no antibacterial activity, thus greatly reducing its bacteriostatic ability.

2. Impact on Solubility and Dispersibility

The water solubility of nisin depends on the charge state of its molecular surface. Under acidic conditions, protonated nisin molecules carry more positive charges, which enhances the electrostatic repulsion between molecules, prevents aggregation, and enables uniform dispersion in aqueous solutions or food matrices. This property makes it easier for Nisin to contact and act on bacterial cells. In contrast, in a neutral or alkaline environment, the charge of nisin molecules decreases, electrostatic repulsion between molecules weakens, and hydrophobic aggregation is prone to occur, forming precipitates. This not only reduces the effective concentration of nisin in the system but also causes some Nisin to be encapsulated inside the precipitates, preventing sufficient contact with bacteria and indirectly weakening the antibacterial effect. For instance, in a milk matrix at pH 7.5, the solubility of nisin is only about 1/3 of that at pH 4.0, and the diameter of its inhibition zone against Staphylococcus aureus is correspondingly reduced by 40%–50%.

3. Impact on Interaction with Targets

The antibacterial mechanism of nisin mainly involves binding to phosphatidylglycerol (PG) on the bacterial cell membrane and lipid Ⅱ (a key precursor for bacterial cell wall synthesis), thereby disrupting the integrity of the cell membrane and inhibiting cell wall synthesis. The phospholipid molecules on the bacterial cell membrane exhibit different charge states under different pH conditions: in an acidic environment, the carboxyl groups (-COOH) on the membrane surface are protonated, resulting in weakened overall positive charge. Meanwhile, nisin molecules are positively charged due to protonation, so the electrostatic repulsion between the two is weak. Nisin can easily insert into the cell membrane through hydrophobic interactions, specifically bind to lipid Ⅱ, and form transmembrane pores. This leads to the leakage of substances such as ions and amino acids inside bacterial cells, ultimately causing bacterial death. When the pH increases, the carboxyl groups on the membrane surface are deprotonated, enhancing the negative charge. The electrostatic repulsion between the membrane and nisin molecules (which are still positively charged but with reduced charge) is instead enhanced. At the same time, lipid Ⅱ may undergo conformational changes in an alkaline environment, reducing its binding affinity with nisin. These two factors make it difficult for nisin to effectively bind to its targets, resulting in a significant decrease in antibacterial efficiency.

4. Indirect Impact via Regulating Bacterial Physiological State

In addition, pH indirectly regulates the antibacterial activity of nisin by affecting the physiological state of bacteria themselves. The acidic environment inherently inhibits the growth and metabolism of some Gram-positive bacteria (the main targets of nisin, such as Listeria monocytogenes and Staphylococcus aureus), for example, reducing the fluidity of bacterial cell membranes and inhibiting enzyme activity, thereby increasing bacterial sensitivity to nisin. In contrast, a neutral or alkaline environment is more suitable for the growth and reproduction of such bacteria. Bacteria can enhance their resistance to Nisin by improving cell membrane repair capabilities and synthesizing drug resistance-related proteins, further offsetting the antibacterial effect of nisin. For example, in meat products at pH 5.0, the minimum inhibitory concentration (MIC) of Nisin against Listeria monocytogenes is 25 IU/mL; while in the same product at pH 7.0, the MIC needs to be increased to 100 IU/mL to achieve the same bacteriostatic effect.

In summary, the antibacterial activity of nisin exhibits a significant "acid-dependent" characteristic with pH: acidic conditions (especially pH 2.0–6.0) can maximize its antibacterial efficiency by stabilizing the molecular structure, improving solubility, and enhancing target binding ability; while neutral to alkaline conditions lead to a significant decrease in its antibacterial activity by disrupting the molecular structure, reducing solubility, and weakening target interactions. This rule provides key guidance for the application of nisin in food processing. For example, in acidic foods (such as pickles, yogurt, and acidic beverages), nisin can directly exert efficient antibacterial effects; while in neutral or alkaline foods (such as fresh meat and soy products), it is necessary to adjust the pH by adding organic acids (such as citric acid and lactic acid) or combine microcapsule encapsulation technology to protect the Nisin structure, thereby maintaining its antibacterial activity.