Nisin raw materials in food processing

As a natural antibacterial peptide, the stability of Nisin directly influences its bacteriostatic effect in food processing. Factors such as pH, temperature, metal ions, compatible components, and processing techniques in the food processing environment significantly affect the structure and activity of Nisin. The specific stability characteristics are as follows:

I. Decisive Role of pH Value on Nisin Stability

High Stability in Acidic Environments

When the pH of the food system ≤4, the molecular structure of Nisin (a cyclic polypeptide containing 5 thioether bonds) is in a protonated state. The ionic bond interaction between amino and carboxyl groups is enhanced, significantly improving its tolerance to heat, enzymes, and other factors. For example, in juice with pH 2.5, Nisin retains over 90% activity after high-pressure sterilization at 121°C for 15 minutes. In yogurt with pH 3.5, the activity loss is less than 10% after storage at room temperature for 3 months.

Rapid Inactivation under Alkaline Conditions

When pH >5, the amino groups in Nisin molecules are deprotonated, and thioether bonds are susceptible to attack and cleavage by hydroxide ions, leading to disruption of the spatial structure. For instance, in neutral canned foods with pH 7, the remaining activity is less than 50% after heating at 60°C for 30 minutes. If the pH rises to 8, it can basically lose activity within 24 hours at room temperature. Therefore, Nisin is more suitable for addition to acidic or acidified foods (such as pickles, acidic milk drinks). If used in neutral foods, the pH should be adjusted to below 4.5 with organic acids, or combined with other bacteriostatic methods (such as low temperature, vacuum packaging).

II. Dual Influence of Temperature and Heat Treatment Processes

Tolerance to Short-Term High-Temperature Treatment

Nisin has strong resistance to short-term high temperatures (especially in acidic conditions). For example, in a system with pH 3.5, about 80% of activity remains after high-pressure sterilization at 121°C for 15 minutes, which is related to the rigid structure formed by thioether bonds in its molecules. In pasteurization (70-85°C, 15-30 seconds) or high-temperature short-time sterilization (HTST, 135°C, 3-5 seconds) processes, the activity loss of Nisin is usually <20%, and it can synergistically inhibit bacteria with heat treatment.

Cumulative Inactivation at Long-Term Low or High Temperatures

If stored in a neutral system at room temperature (25°C) for a long time, Nisin will gradually inactivate due to slow peptide bond hydrolysis (half-life is about 15 days). In a high-temperature environment above 100°C, even if pH ≤4, long-term heating (such as 2 hours) will cause gradual 断裂 (cleavage) of thioether bonds, and the activity will 衰减 (decline) to below 50%. Therefore, in foods requiring long-term sterilization (such as meat cans), it is recommended to add Nisin during the cooling stage after sterilization (temperature <60°C) to avoid the continuous effect of high temperature.

III. Interference Effects of Metal Ions and Chelating Agents

Destructive Effect of High-Valence Metal Ions

Transition metal ions such as Cu²⁺ and Fe³⁺ easily undergo coordination reactions with sulfhydryl (-SH) and amino (-NH₂) groups in Nisin molecules, leading to distortion of its spatial structure and decline in bacteriostatic activity. Experiments show that 0.1 mM Fe³⁺ can reduce the bacteriostatic effect of Nisin on Staphylococcus aureus by 30%, and 0.05 mM Cu²⁺ can reduce the activity retention rate to below 60%. Therefore, in food processing, direct contact between Nisin and copper/iron containers should be avoided, and stainless steel or glass containers should be selected for storage.

Dual Role of Ca²⁺ and Mg²⁺

Low concentrations of Ca²⁺ (<10 mM) and Mg²⁺ (<5 mM) have little effect on Nisin activity, and can even enhance its bacteriostatic effect by stabilizing the bacterial cell membrane potential (such as naturally existing Ca²⁺ in dairy products can synergize with Nisin to inhibit spore germination). However, high concentrations of Ca²⁺ (>20 mM) may compete with Nisin for binding to bacterial cell membrane receptors, instead weakening the activity. In addition, adding chelating agents (such as EDTA) can complex metal ions in the system and reduce their damage to Nisin. For example, adding 0.01% EDTA to juice containing trace Fe³⁺ can increase the activity retention rate of Nisin from 70% to 90%.

IV. Inactivation Risks of Proteases and Compatible Components

Hydrolysis Effect of Proteases

As a polypeptide substance, Nisin can be recognized and hydrolyzed by proteases in foods (such as pepsin, trypsin, papain). For example, in meat products added with meat tenderizer (containing papain), the activity of Nisin will rapidly 衰减 (decline) to below 40% within 2 hours. Therefore, if proteases exist in the food system, it is necessary to protect Nisin by adjusting the pH to acidic (inhibiting protease activity), using microcapsule embedding technology, or adding it after protease inactivation (such as after heat treatment to inactivate enzymes).

Interactions with Other Components

Surfactants (such as Tween-80, sodium dodecyl sulfate) may dissociate Nisin from the bacterial cell membrane by destroying its hydrophobic interactions, reducing the bacteriostatic effect.

Proteins (such as whey protein, soybean protein) can form complexes with Nisin at low concentrations (<1%), protecting it from protease hydrolysis through steric hindrance. However, at high concentrations (>5%), they may weaken the activity due to competitive binding to bacterial cell membrane receptors.

Polysaccharides (such as pectin, sodium alginate) can 包裹 (encapsulate) Nisin by forming a gel network, delaying its release, and prolonging the activity maintenance time in acidic jellies and jams.

V. Adaptability of Food Matrix and Processing Technology

Stability Differences in Different Food Matrices

Dairy products: Casein micelles in milk can adsorb Nisin. In fresh milk with pH 6.5, the half-life of Nisin is about 48 hours (refrigerated at 4°C), but high-temperature sterilization (such as 135°C, 2 seconds) will cause casein denaturation, release Nisin, and make its activity lose about 30%.

Meat products: Myoglobin in muscles contains Fe²⁺, which has little effect on Nisin activity if not oxidized to Fe³⁺ by sodium nitrite. However, added NaCl (>5%) during pickling may cause slight conformational changes in Nisin due to increased osmotic pressure, with activity decreasing by about 10%-15%.

Acidic beverages: Nisin has the best stability in juice containing citric acid (0.5%), with over 80% activity remaining after storage at room temperature for 6 months. However, carbonated beverages (such as cola, pH 2.8-3.2) may accelerate the oxidative inactivation of Nisin due to bubble disturbance caused by carbon dioxide dissolution, requiring sealed storage.

Optimization Strategies for Processing Technology

In fermented foods (such as yogurt), Nisin should be added in the late fermentation stage (after the acidity meets the standard) to avoid inhibiting the lactic acid bacteria fermentation process (although Nisin has self-resistance to Lactococcus lactis, it may have a slight inhibitory effect on other lactic acid bacteria).

In baked foods (such as bread), Nisin should be added during the dough kneading stage, using starch in flour to encapsulate and protect it, reducing activity loss caused by high-temperature baking (180-220°C, 15-30 minutes) (usually a loss of about 40%-50%, but the remaining activity can still inhibit spore-forming bacteria).

VI. Stability Monitoring and Activity Maintenance Measures

Activity Detection Methods

Monitor Nisin activity through the bacteriostatic ring test (Oxford cup method) or turbidity method. For example, add the Nisin solution to an agar plate containing indicator bacteria (such as Staphylococcus aureus ATCC 25923) and measure the diameter of the bacteriostatic ring. If the diameter is reduced by more than 20% compared to the initial value, it indicates a significant decline in activity.

Means to Improve Stability

pH regulation: Control the pH of the food system below 4.5, and preferentially use organic acids (such as lactic acid, citric acid) for regulation to avoid strong bases.

Metal ion control: Use deionized water to dissolve Nisin, avoid contact with metal containers, and add 0.01%-0.05% EDTA to complex trace metal ions.

Embedding technology: Use sodium alginate-calcium ion microcapsules to 包埋 (embed) Nisin, which can extend its half-life in neutral foods from 1 day to 7 days.

Low-temperature storage: Unopened Nisin raw materials need to be refrigerated at 4°C. After opening, prepare it into a 1% acidic aqueous solution (pH 3-4) and store it in the dark at 4°C, with a validity period of about 1 month.

The stability of Nisin in food processing is affected by multiple factors synergistically. The core is to maintain its structural integrity through acidic environment control, metal ion chelation, avoiding proteases, and optimizing processing timing. In practical applications, the addition method and dosage of Nisin should be adjusted according to the characteristics of the food matrix (such as pH, composition) and process conditions (such as sterilization temperature, storage cycle). When necessary, combine with compound bacteriostatic agents (such as organic acids, EDTA) for synergistic enhancement to ensure the long-term exertion of its bacteriostatic activity.