The role of Nisin in fruit juice beverages

I. Antibacterial Mechanism of Nisin: The "Targeted Bactericidal" 特性 of Natural Peptides

Nisin is a natural antibacterial peptide secreted by Lactococcus lactis, whose bacteriostatic effect relies on specific disruption of bacterial cell membranes:

Action Target: Cell membranes of Gram-positive bacteria (e.g., Listeria, Staphylococcus aureus, bacilli). Nisin recognizes Lipid II (a peptidoglycan synthesis precursor) on the membrane, inserts into the membrane structure to form pores, causing leakage of intracellular ions (e.g., K+) and ATP, ultimately inhibiting bacterial reproduction.

Broad-Spectrum Limitations and Synergistic Enhancement: It has weak 单独 action against Gram-negative bacteria (e.g., E. coli) due to the lipopolysaccharide barrier of the outer membrane blocking Nisin penetration. However, under acidic conditions (juice pH often 2.5–4.5), the outer membrane permeability of Gram-negative bacteria increases. Nisin can be compounded with citric acid, ascorbic acid, etc., to enhance bacteriostatic effect by damaging the inner membrane integrity.

II. Antibacterial Efficacy in Juices: Influences of Concentration, Microbial Types, and Environmental Factors

Effective Bacteriostatic Concentration Range

The common addition amount of Nisin in juices is 50–200 mg/L, specifically determined by microbial contamination degree and juice components. For example, the minimum inhibitory concentration (MIC) against Staphylococcus aureus in fresh apple juice is approximately 80 mg/L, while 150–200 mg/L is required for heat-resistant bacilli (e.g., Bacillus stearothermophilus).

Synergistic Bacteriostatic Effect: When combined with tea polyphenols (50–100 mg/L), the bacteriostatic concentration can be reduced by 30%–50%, and the inhibitory effect on yeasts (e.g., Saccharomyces cerevisiae) is significantly enhanced (Nisin alone is ineffective against fungi).

Inhibition Effects on Typical Contaminant Bacteria

Control of Bacilli: Nisin inhibits spore germination and growth in juices, especially in acidic environments (pH<4.0), which damages the osmotic barrier of the spore cortex to prevent activation. For example, adding 100 mg/L Nisin to citrus juice can reduce the viable count of Bacillus stearothermophilus by 4 log levels.

Pathogen Prevention and Control: For common foodborne pathogens in juices (e.g., Listeria monocytogenes), Nisin shows the strongest bacteriostatic activity at pH 3.5–4.0, reducing the bacterial count from 10⁵ CFU/mL to below 10² CFU/mL within 24 hours.

III. Stability of Nisin in Juices: The "Double-Edged Sword" Effect of Environmental Factors

Nisin stability is significantly influenced by juice components (e.g., sugar, organic acids, polyphenols) and processing conditions (e.g., temperature, pH, storage time):

1. pH Value: The Most Critical Stability Factor

Acidic Environment (pH 2.0–4.5): Amide bonds and disulfide bonds in Nisin structure are less prone to hydrolysis, with stability significantly better than neutral or alkaline conditions. For example, in lemon juice at pH 3.0, Nisin retains over 90% activity after heating at 60℃ for 30 minutes; in carrot juice at pH 6.0, the same conditions cause 50% activity loss.

Mechanism Explanation: Nisin carries a positive charge under acidic conditions, forming stable complexes with anionic components in juices (e.g., pectin, citrate), reducing heat-induced structural denaturation.

2. Influence of Thermal Processing and Sterilization Processes

Compatibility with Pasteurization: Nisin maintains good stability in low-temperature short-time (LTLT, e.g., 70℃/15 s) or high-temperature short-time (HTST, e.g., 90℃/10 s) sterilization, with activity loss usually <20%; however, under ultra-high temperature sterilization (UHT, 135℃/3 s), the activity retention rate drops to 60%–70%, requiring appropriate increase in addition amount.

Residual Activity After Sterilization: Nisin continues to exert bacteriostatic effect in sterilized juices. For example, in apple juice stored at 4℃ for 30 days with 150 mg/L Nisin added, the inhibition rate of thermophile regrowth reaches 85%.

3. Synergistic and Antagonistic Effects of Juice Components

Components Promoting Stability:

Sucrose (concentration >10%) reduces thermal degradation of Nisin by lowering water activity (aw). For instance, in a 15% sucrose solution, Nisin retains 25% more activity after 80℃/20 min than in the absence of sucrose.

Ascorbic acid (vitamin C) has antioxidant effects, inhibiting oxidation of methionine residues in Nisin molecules and delaying activity decay.

Components Inhibiting Stability:

Polyphenols (e.g., tannins, anthocyanins) can bind to Nisin through hydrophobic interactions, forming insoluble complexes and reducing its effective concentration. For example, adding 200 mg/L Nisin to grape juice with tannin content >500 mg/L decreases bacteriostatic effect by about 30%.

Metal ions (e.g., Fe³⁺, Cu²⁺) catalyze oxidative degradation of Nisin, requiring addition of EDTA (0.05%–0.1%) to chelate metal ions and improve stability.

4. Influence of Storage Conditions

Temperature and Time: Nisin activity loss is slow during refrigeration at 4℃ (about 5% per month), while at 25℃ room temperature, activity loss can reach 30%–40% within 30 days, requiring cold chain transportation to maintain efficacy.

Oxygen and Light: Oxygen promotes Nisin oxidation, and ultraviolet irradiation damages its peptide bond structure. Therefore, juice packaging should use light-proof and oxygen-barrier materials (e.g., brown glass bottles or aluminum foil bags).

IV. Challenges and Solutions in Practical Applications

Adaptability to Juice Processing Techniques

Problem: Some juices (e.g., peach juice, pear juice) release proteases (e.g., papain) during blanching or homogenization, degrading the peptide chain structure of Nisin.

Solution: Adjust the addition timing of Nisin, such as adding after sterilization and cooling to below 40℃ to avoid direct contact with proteases; or select Nisin variants with stronger protease resistance (e.g., Nisin Z).

Balance Between Cost and Safety

Nisin has a high production cost (about 500–800 yuan/kg), and high-dose addition significantly increases beverage costs. A compounding scheme of "Nisin + natural preservative" (e.g., 100 mg/L Nisin + 80 mg/L tea polyphenols) can maintain bacteriostatic effect while reducing usage.

In terms of safety, Nisin has been approved as GRAS (Generally Recognized As Safe) by institutions such as FDA and EFSA, with an acceptable daily intake (ADI) of 0–33,000 IU/kg body weight. Calculated based on an addition amount of 200 mg/L in juice, adult daily consumption of 1 L is far below the safety threshold.

V. Research Trends and Prospects

Molecular Modification to Enhance Performance

Site-directed mutagenesis to modify the Nisin structure (e.g., replacing methionine residues) enhances its resistance to acid proteases and thermal stability, achieving over 50% increase in activity retention rate in laboratories.

Application of Nano-Drug Delivery Systems

Encapsulating Nisin in chitosan-alginate nanospheres delays its release rate, maintaining long-acting bacteriostasis during juice storage while reducing binding loss by polyphenols.

Optimization of Green Extraction Processes

Using membrane separation technologies (e.g., ultrafiltration, reverse osmosis) instead of traditional precipitation methods to extract Nisin can increase purity from 80% to over 95%, reducing the impact of impurities on juice flavor.

Conclusion

As a natural and efficient biological preservative, the application of Nisin in fruit juices requires combining its antibacterial mechanism and stability characteristics. Through process optimization and compounding technologies, a balance between preservation effect and product quality can be achieved, providing a new path for low-additive and high-safety juice processing.