Many food companies and brands are developing bio-preservation and fresh-keeping technologies for various products, among which the use of Nisin is very critical. Therefore, some particularly important tips on the use of nisin and instructions on its antibacterial effect, It will be shown in the article in turn, in order to help us in the actual use process

Part One:Properties of nisin

Nisin, produced by Streptococcus lactis, is a polypeptide compound composed of a variety of amino acids

Characteristic:

White to pale yellow free flowing powder, slightly salty (sodium chloride component)

Contains milk protein, the aqueous solution is slightly turbid

Sensitive:

Particularly sensitive to proteolytic enzymes(trypsin, pancreatic enzymes, salivary enzymes, digestive enzymes)

Storage:

Store in a cool, dry and sealed condition at 4-15°C, avoid direct sunlight

Safety:

Absorbed and utilized by the human body as a nutrient, it is a non-toxic natural preservative

No adverse effects on food color, aroma, taste, taste, etc.

Can be decomposed by intestinal digestive enzymes, with high safety

Antibacterial pH value: 6.5-6.8

Part Two:Nisin Solubility

1. It needs to be dissolved in water or liquid when used, and insoluble in non-polar solution

2. Nisin within the allowable usage range can be well miscible with water or other processing liquids

3. The actual use concentration is generally not more than 0.25‰, so the solubility will not be an obstacle to its use in various foods

4. The solubility depends on the pH value, and the solubility in water decreases with the increase of the pH value

❶When PH=2.5, the solubility is 12%

❷When PH=5.0, the solubility is 4%

❸When PH>7 (under neutral or alkaline conditions), it is almost insoluble

5. When Nisin is added for anti-corrosion, the food must be acidic, and it is stable at room temperature and acid during processing and storage

6. Nisin's industrial products contain a certain amount of denatured milk protein, its insolubility will cause the solution to appear misty suspension, but it will not affect its antiseptic effect

7. Nisin is particularly sensitive to proteolytic enzymes (such as pancreatic enzymes, sleeping fluid enzymes and digestive enzymes, etc.), so it can be hydrolyzed into amino acids by proteolytic enzymes in the digestive tract after eating. After inhaling Nisin-containing liquid for 10 minutes, Nisin cannot be detected in saliva

Part Three:Stability of nisin

1. The best antibacterial pH value is 6.5-6.8

2. Nisin has extremely stable activity under acidic conditions. The stability mainly depends on factors such as temperature, pH, and matrix. The activity decreases with the increase of pH

If heated at 121°C for 15 minutes, its activity decreased by 29%, 69%, 86% and 99.7% at pH 4, 5, 6 and 7, respectively.

❶When PH<2.0, it can be sterilized at 116℃ without loss of activity

❷When pH=2, it is still active after high temperature sterilization at 121°C for 30 minutes, and has good heat resistance

❸When pH=3, the maximum activity (97.5%) is still retained after heat sterilization at 121°C for 20 minutes

It has excellent acid and heat resistance, and its activity will decrease if the pH value is lower or higher than this value.

❹When pH>4, heating will speed up its decomposition and reduce its antibacterial activity

❺When PH=5.0, after heating to 115.6℃ for sterilization, 40% of the activity will be lost

❻When PH>5, the heat resistance decreases, and after pasteurization at 85°C for 15 minutes, the activity only loses 15%

❼ In skimmed milk with PH=6.5, 90% of the activity will be lost when heated to 115.6°C

❽When PH>6.8, 90% of the activity will be lost after sterilization

❾It is not suitable to heat for too long under neutral conditions, otherwise the activity will be greatly reduced

Inactivation occurs at room temperature at pH = 7.0

⑩When pH=11, after sterilizing at 63°C for 30 minutes, inactivate immediately

3. In foods such as milk, macromolecular proteins can provide protection for Nisin, making it less decomposed and less likely to lose antibacterial activity when heated.

Part Four: The concentration of nisin

1. In order to unify the standard, WHO regards a sample with a content of 25mg/g as a reference preparation, and defines 0.001mg; this preparation is an "international unit" (IU), and the amount is the same as the "reading unit"

2. 0.001mg of a sample with a content of 25mg/g, which is equivalent to one international unit or reading unit:

That is, 0.025mg of pure nisin is equivalent to 1U or 1 reading unit, or 1mg of pure nisin is equivalent to 401U or 40 reading units

3. In a preparation of 1 IU or 1 reading unit, the concentration of nisin is 0.0251 U

4. Commercial preparations are often expressed in international units (IU), 1IU is equivalent to 0.025mg of pure nisin

Part five: Antibacterial Mechanism of Nisin

Antibacterial mechanism:

By interfering with the normal function of the cell membrane, causing cell membrane penetration, nutrient loss and membrane potential drop, leading to the death of pathogenic bacteria and spoilage bacteria cells

1. Nisin has effects on both vegetative cells and spores of bacteria, and the main point of action on vegetative cells is the cytoplasmic membrane

It can inhibit the biosynthesis of peptidoglycan in the bacterial cell wall, block the synthesis of cell wall plasma membrane and phospholipid compounds, cause the contents of the cell membrane to leak out, and even cause cell lysis

2. In most cases, the mode of action of Nisin on bacterial spores is - antibacterial rather than bactericidal

❶The effect on spores is to inhibit the germination of spores at the initial stage of expansion

❷ When the spores germinate and expand, Nisin acts as a cationic surfactant, affecting the bacterial spore membrane and inhibiting the synthesis of gram-positive bacteria.

❸ When the spores germinate, they are killed because they are sensitive to Nisin, thereby inhibiting the germination process of the spores [Inhibiting the germination of the spores]

3. There are many factors that affect the effect of Nisin. In addition to the type of bacteria, the age of the bacteria, the number of bacteria (the amount of bacteria carried in food), the medium and environmental conditions, and the dissolved state of Nisin all have an impact.

❶ In terms of bacterial age, Nisin has a stronger inhibitory ability to spores than to bacteria

❷Even the bacterial cell membrane destroyed by mechanical means is more sensitive to Nisin than the undamaged one

❸The amount of Nisin used is directly related to the amount of bacteria in the food (number of bacteria)

4. From the perspective of Nisin stability, its antibacterial activity is directly related to heating temperature and heating time, and it should also be seen that Nisin has a more obvious effect on bacteria or spores damaged by heating

❶Normal bacteria are heated at 121.1°C for 3 minutes, and their sensitivity to Nisin is about 10 times that of those without heat

❷For dairy products, generally speaking, if the number of bacteria increases by 10 times, the amount of Nisin should be doubled

❸The dissolution state of Nisin is closely related to the bactericidal effect during use. The best solvent for Nisin is 0.02m/L hydrochloric acid

Nisin and heat treatment can promote each other:

❶ On the one hand, the use of a small amount (0.25-10mg/kg) of Nisin can improve the heat sensitivity of spoilage microorganisms

❷ On the one hand, heat treatment also increases the sensitivity of bacteria to Nisin

❸So heating after adding Nisin can not only improve the effect of Nisin, thereby reducing the concentration, but also greatly improve the effect of heat treatment and reduce the temperature of heat treatment

❹The commercial sterilization of food after adding Nisin has become a means to maintain the nutritional value and improve the sensory quality of canned food and dairy products

❺Nisin and heat treatment can synergize each other, which is of great significance in practical application

For example, dairy dessert products cannot be thoroughly sterilized by heating, so as not to damage the appearance, taste and consistency of the product. After pasteurization, the shelf life is limited, but after adding Nisin and then pasteurized, the shelf life can be significantly extended.

Part Six: Antibacterial Spectrum of Nisin

1. Nisin has a relatively narrow antibacterial spectrum and can only inhibit or kill Gram-positive bacteria, especially bacterial spores

❶Mainly targeting G+ bacteria, such as Bacillus and Clostridium sporogenes, especially can inhibit the reproduction of botulism and the formation of toxins

❷ Against Staphylococcus aureus, Lactobacillus, Clostridium botulinum, Bacillus, Clostridium, Micrococcus, Streptococcus, Hemolytic Streptococcus, Listeria monocytogenes, Clostridium butyricum, fat thermophiles A variety of spoilage and food pathogens such as Bacillus, or other anaerobic spore-forming bacteria, have a strong inhibitory effect—inhibiting their growth and reproduction

2. The inhibitory effect on the germination of Bacillus and Clostridium spores is greater than that on the vegetative cells

3. Generally, bacterial spores cannot be killed, but when the spores germinate and expand, it will increase their sensitivity to milk chains and be killed

4. Nisin has a strong antibacterial effect on Gram-positive bacteria such as Clostridium botulinum, but it has a weak effect on Gram-negative bacteria, mold and yeast, and it needs to be stable under acidic conditions sex

The cell wall of Gram-negative bacteria is relatively complex, and can only allow substances with a molecular weight below 60Da to pass through. Nisin has a molecular weight of 3150Da, so it cannot reach the cell membrane, resulting in no effect on Gram-negative bacteria.

5. The bactericidal spectrum of Nisin is relatively narrow, and it is often used in combination with other antiseptic methods to make up for the shortcoming of its narrow antibacterial spectrum and exert a wide range of antiseptic effects

6. Nisin is particularly sensitive to proteolytic enzymes, and is often used in the preservation of cheese, cream products, canned food, and high-protein products

7. Nisin is successfully used in high-acid food (PH value less than 4.5). In non-acid food, Nisin can reduce the temperature and time of heat treatment, and better maintain the nutritional content and flavor of food

8. Most of the spoilage microorganisms and pathogenic bacteria in dairy products, canned foods and some alcoholic beverages can be killed or inhibited by Nisin

For example: Staphylococcus aureus, Streptococcus hemolyticus, Clostridium botulinum and other pathogenic bacteria in dairy products; Lactobacillus and Leuconostococcus in beer; Clostridium niger, Clostridium botulinum, Clostridium sporogenes, Pasteurella, Lactobacillus, Bacillus coagulans, Bacillus polymyxa, and Bacillus softening are all sensitive to Nisin, generally 10mg/kg is effective

An important role of Nisin in food:

Inhibition of the growth of Clostridium botulinum in meat products and low-acid canned foods--Supplementation of nitrite in meat curing to inhibit the growth of Clostridium difficile

For example, in fish and meat products, Nisin can significantly reduce the amount of nitrate used without affecting the color and antiseptic effect of meat, so as to effectively prevent the formation of Clostridium botulinum toxin