Factors affecting the application effect of nisin in cheese

The effectiveness of Nisin in cheese (such as antimicrobial activity, stability, and impact on product quality) is regulated by multiple factors. These factors act by influencing the stability of its molecular structure, interactions with the cheese matrix, and the sensitivity of target microorganisms, as detailed below:

I. Physicochemical Properties of the Cheese Matrix

The composition of cheese (e.g., protein, fat, moisture) and environmental parameters (pH, salt content) are core factors affecting Nisin activity.

Protein binding: High levels of casein in cheese (especially casein micelles) can bind to Nisin through hydrophobic interactions and electrostatic adsorption, forming inactive complexes. This binding reduces the concentration of free Nisin—for example, in high-protein hard cheeses (such as Cheddar), its effective antimicrobial activity may be 30%-50% lower than in soft cheeses (such as Brie). Additionally, protein hydrolysates (e.g., peptides) produced during cheese ripening may compete with Nisin for binding sites, indirectly affecting its antimicrobial efficiency.

Fat content and distribution: Fat droplets can encapsulate Nisin molecules, hindering their migration to microbial cell membranes. In high-fat cheeses (such as cream cheese), the isolating effect of the fat phase significantly reduces its inhibitory effect on waterborne pathogens (e.g., Listeria); in low-fat cheeses, however, Nisin distributes more uniformly in the aqueous phase, resulting in relatively stable antimicrobial activity.

pH impact: Nisin is more stable in acidic environments (pH 3.0-6.0), which is related to the protonation state of residues such as lysine and histidine in its molecule. The pH of cheese typically changes during ripening (e.g., from pH 4.6 during curdling to pH 5.5 in late ripening). Under neutral to alkaline conditions, Nisin molecules are prone to conformational changes, leading to decreased antimicrobial activity. For example, in blue cheese with pH > 6.0, its half-life is approximately 40% shorter than in mozzarella cheese with pH 5.0.

Salt concentration and water activity: Salt (NaCl) added during cheese processing exerts dual effects by influencing Nisin solubility and molecular charge: low salt concentrations (2%-3%) promote its dissolution and enhance interactions with microbial cell membranes; high salt concentrations (e.g., in cheese after salting, with salt content > 6%) cause Nisin molecules to aggregate and precipitate, reducing activity. Meanwhile, low water activity (e.g., < 0.90 in hard cheeses) limits its diffusion, weakening its inhibitory effect on deep-seated microorganisms.

II. Processing Parameters

Cheese production processes (e.g., heat treatment, fermentation, ripening conditions) directly affect Nisin retention and distribution uniformity.

Heat treatment intensity: Nisin is prone to thermal denaturation at high temperatures (>80℃), with lanthionine bridges in its molecule breaking, resulting in loss of activity. Therefore, in cheese processing requiring high-temperature sterilization (e.g., blanched curd technology), it must be added after heat treatment; in low-temperature fermented cheeses (e.g., yogurt cheese), it can be added during curdling to reduce heat damage.

Fermentation and ripening temperature: Nisin’s thermal stability decreases with increasing temperature—in cheeses ripened at 20℃, over 80% of its activity can be retained; while ripening at above 35℃ (e.g., in some fast-ripening cheeses) may lead to 30%-50% monthly activity loss. Additionally, high temperatures accelerate microbial degradation of Nisin (e.g., peptidases produced by certain lactic acid bacteria can decompose Nisin), further weakening its effectiveness.

Addition timing and method: Adding Nisin before curdling makes it prone to binding with whey proteins and being lost with whey; adding it after curd formation but before salting improves its retention in curds. Dispersibility is also crucial—directly adding solid Nisin may cause rapid binding to proteins due to local high concentrations, while pre-dissolving in sterile water or dilute salt solution and spraying uniformly enhances its distribution in the cheese matrix, boosting antimicrobial efficacy.

III. Microbial Factors

The type, physiological state, and drug resistance of target microorganisms affect Nisin’s effectiveness.

Differential microbial sensitivity: Nisin is most effective against Gram-positive bacteria (e.g., Staphylococcus aureus, Bacillus), but less effective against Gram-negative bacteria (e.g., Escherichia coli) due to their outer membrane barrier blocking Nisin penetration. If cheese is contaminated with Gram-negative bacteria, combination with other preservatives (e.g., EDTA) is required to enhance its activity by disrupting the outer membrane.

Impact of spores and biofilms: Bacterial spores (e.g., Clostridium botulinum spores) are far less sensitive to Nisin than vegetative cells, requiring higher concentrations (usually 2-3 times the conventional dose) to inhibit germination; microbial biofilms (e.g., Listeria biofilms on cheese surfaces) can block Nisin penetration through extracellular polysaccharide matrices, reducing antimicrobial effects. In such cases, combining with physical treatments (e.g., surface cleaning) enhances efficacy.

Development of resistance: Long-term low-dose use of Nisin may induce resistance genes (e.g., nisI) in some microorganisms (e.g., Lactococcus). Proteins encoded by these genes can bind to Nisin and expel it from cells, reducing inhibitory effects. Therefore, cheese production should avoid long-term single use of Nisin; rotating with other antimicrobials or optimizing doses reduces resistance risks.

IV. Intrinsic Properties of Nisin

Purity and formulation: High-purity Nisin (activity > 1000IU/mg) has higher antimicrobial efficiency, while impurities in low-purity products (e.g., culture medium residues) may interfere with its interaction with microorganisms. Additionally, microencapsulated Nisin (e.g., embedded in sodium alginate) reduces binding with the cheese matrix and prolongs release, making it particularly suitable for preserving long-ripened cheeses (e.g., Parmesan).

Addition dosage: Too low a dosage fails to reach the bacteriostatic threshold; too high a dosage may cause excessive binding with cheese components, reducing free activity and potentially introducing a slight bitter taste (especially in low-salt cheeses). Practical application requires adjustment based on cheese type—for example, high-moisture soft cheeses prone to spoilage typically require 25-50IU/g, while hard cheeses may need only 10-20IU/g.

Nisin’s effectiveness in cheese results from the combined action of the cheese matrix, processing techniques, microbial characteristics, and its own properties. Optimizing addition methods, regulating ripening conditions, and combining with auxiliary measures (e.g., pH adjustment, preservative compounding) in production maximizes its antimicrobial efficacy while ensuring stable cheese quality.