Application of all-natural plant preservative grass extract in research and development of food fresh-keeping process

Abstract

The invention discloses a research and development application of an all-natural plant preservative grass extract in a food preservation process. The research and development application comprises the following steps: step 1, selecting plant preservative grass; 2, determining an extraction method; step 3, purifying the extract; and step 4, application of the food fresh-keeping technology. The method has the advantages that the risk of chemical substance residues in food is reduced through the all-natural plant preservative grass extract in the food preservation process, safer and healthier food choices are provided for consumers, growth and reproduction of microorganisms such as bacteria, molds and saccharomycetes in the food can be effectively inhibited, and the food preservation effect is improved. The fresh-keeping agent has the advantages that the quality guarantee period of food is prolonged, excessive mining and damage to the environment are avoided, the requirement of sustainable development is met, meanwhile, the good fresh-keeping effect can prolong the sales cycle and shelf life of the food, the economic benefits of food enterprises are increased, the requirements of consumers for high-quality and safe food are met, and the market competitiveness of the enterprises is improved.

Description

Technical Field

[0001] The invention relates to the technical field of all-natural plant food preservatives, and in particular to the research and development and application of an all-natural plant antiseptic grass extract in food preservation technology. Background Art

[0002] In the food industry, freshness preservation is a vital issue. In recent years, food safety issues have become increasingly severe and have become a focus of great concern from all walks of life. In the field of food processing, the hazards caused by excessive or unsafe chemical preservatives have become increasingly prominent, causing widespread consumer concerns. Consumers' requirements for food safety and quality are constantly increasing, and consumers are paying more and more attention to food safety and health, and are eager to buy healthy and safe food without chemical additives.

[0003] Compared with traditional chemical preservatives, all-natural plant food preservatives have significant advantages. First, they are biodegradable and can decompose quickly in the natural environment without causing long-term pollution to the environment. Secondly, all-natural plant food preservatives have minimal impact on the human body and the environment. They are derived from natural plants, do not contain harmful chemicals, and will not cause harm to human health. At the same time, during the production and use process, no waste harmful to the environment will be generated.

[0004] Therefore, the development and application of Landel's all-natural plant food preservatives is of great significance. It is an important way to achieve the green and sustainable development of the food industry. By promoting the use of all-natural plant food preservatives, the use of chemical preservatives can be reduced, reducing the risks to human health and the environment. At the same time, it can also promote technological innovation and industrial upgrading in the food industry, and promote the development of the food industry in a green, healthy and sustainable direction. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a kind of all-natural plant antiseptic grass extract which is simple and reliable to use, easy to clean, strong in anti-breakage ability and low in cost and can be applied in the research and development of food preservation technology.

[0006] In order to solve the above technical problems, the present invention is solved by the following technical solution: a method for developing and applying a natural plant antiseptic grass extract in food preservation technology comprises the following steps:

[0007] Step 1. Selection of plant antiseptic grass: determine the plant species with antiseptic properties, select specific antiseptic grass varieties after extensive screening and research, conduct origin surveys on the selected antiseptic grass to ensure that its growth environment is pollution-free, free of pesticide residues and excessive heavy metals, establish strict procurement standards, and only select antiseptic grass with good growth and moderate maturity for subsequent processing;

[0008] Step 2, determination of the extraction method: try different organic solvents, after determining the best solvent, optimize the extraction temperature, time and solvent concentration and other parameters, conduct gradient experiments within a certain temperature range, determine the most suitable extraction temperature, observe the changes in the extract yield and preservative activity by changing the extraction time, adjust the solvent concentration, find the best ratio that can both ensure extraction efficiency and reduce costs, filter and concentrate the extracted solution, remove impurities and excess solvents, obtain preliminary extracts, select supercritical fluids such as carbon dioxide as extractants, study the effects of different pressures, temperatures and flow rates on the extraction effect, optimize the extraction process parameters, ensure the purity and preservative activity of the extract, gradually increase the pressure, and observe the changes in the extract yield and quality; adjust the temperature to find the most suitable extraction temperature range, control the flow rate to make the extraction process more efficient, collect the extracted extracts, and conduct subsequent processing and analysis;

[0009] Step 3, purification of the extract: purify the extract by column chromatography, select a suitable adsorbent, separate the components with antiseptic activity by adjusting the composition and flow rate of the eluent, test the purified extract to ensure that its purity meets the requirements, and use high performance liquid chromatography (HPLC) and other analytical methods to detect the content and purity of the active ingredients in the extract;

[0010] Step 4, application of food preservation technology: determine the appropriate addition method for different types of food, conduct experiments to study the effects of different addition amounts on food preservation, conduct storage experiments on foods treated with different addition amounts of extracts, observe their microbial growth, color changes, texture changes and other indicators, comprehensively consider preservation effects and cost factors, and determine the best addition amount. Generally speaking, too little addition may not achieve the ideal preservation effect, while too much addition may affect the taste and quality of the food, and also increase costs. Combined with traditional preservation methods such as low-temperature storage, vacuum packaging, and modified atmosphere packaging, further improve the preservation effect of food, study the synergistic effect of different preservation measures, optimize preservation process parameters, determine the best storage temperature and humidity conditions, as well as suitable packaging materials and packaging methods;

[0011] Step 5. Effect evaluation and optimization: Detect microbial indicators in food to ensure that the food meets hygiene standards and evaluate the sensory quality of the food. This can be done by a professional sensory evaluation team, or by using instrumental analysis methods to monitor changes in physical and chemical indicators of food during storage. If the preservation effect is found to be unsatisfactory, the extraction process parameters can be adjusted to improve the purity and preservative activity of the extract, or the preservation process can be optimized, continuously improved and perfected to meet the preservation needs of different foods and enhance the market competitiveness of products.

[0012] Preferably, the organic solvent includes ethanol, acetone, ethyl acetate, etc., and the adsorbent includes silica gel, alumina, etc.

[0013] Preferably, the adding method includes spraying or dipping for fruits and vegetables to evenly cover the extract on the food surface; adding an appropriate amount of extract during the processing of meat and aquatic products, or using a vacuum packaging combined with extract treatment method; for dairy products and beverages, the extract can be directly added to the product, or the extract can be slowly released into the food through dipping treatment of the packaging material; the microbial indicators include total bacteria count, mold and yeast count, etc.

[0014] Preferably, the sensory qualities of the food include color, smell, taste and texture, etc., the instrumental analysis method includes a colorimeter, a texture analyzer, etc., the physical and chemical indicators include pH value, water content, vitamin content, etc., and the preservation process includes the addition method, addition amount, and combination with other preservation measures, etc.

[0015] Preferably, the 50 main compounds predicted by the antiseptic herbal materials include isoanisole, kalinol, anisole, 5-methoxypsoralen, isoflavone, Da-2-methylpiperidine, amino acids, angelica root root, dihydrocyperquinone, piceatannol, kaempferol, betaine, angelica root root lactone, isofraxin, hypoxanthine, toxicostearin, 3-hydroxycoumarin, ferulic acid, dihydrocarbuncle, Alcohol, pursenaol, cyperus quinone, cnidium monnieri, zephyranthes lactone, diosmin, flavonoid glycosides, isoeugenol, garcinol, anemonin, chlorogenic acid, isochlorogenic acid, neochlorogenic acid, eugenol, camphor, isopulegone, L-pineneketone, myrtenol, menthone, norarecaquinone, isonepetalactone, artemisinic acid, cordycepin, danshensu, bergamotol, linolenic acid, costus alcohol, caltropone, pursenaside and coumarin.

[0016] Preferably, the antiseptic grass extract is obtained through a specific extraction process, which includes but is not limited to one or more combinations of solvent extraction, supercritical fluid extraction, and ultrasonic-assisted extraction. The antiseptic grass extract can be used to preserve various foods, including but not limited to fruits, vegetables, meat, dairy products, pastries, etc. The extraction technology includes ultrasonic extraction, microwave extraction, supercritical fluid extraction, etc.

[0017] Preferably, the antiseptic grass extract is added to food at a specific concentration and in a specific manner to achieve an effective preservation effect. The specific concentration and addition method are adjusted according to different types of food. During the food preservation process, the antiseptic grass extract can effectively inhibit the growth and reproduction of microorganisms such as bacteria, molds and yeasts, extend the shelf life of the food, and at the same time maintain the original color, flavor and nutritional components of the food.

[0018] Preferably, the raw material plant of the antiseptic grass extract is a natural plant resource that can be grown sustainably, and its extraction and application process is environmentally friendly and will not cause pollution. The process of using the antiseptic grass extract for food preservation includes the steps of extraction, purification, and adding to food. Each step is carried out under specific conditions to ensure the activity and preservation effect of the extract.

[0019] Preferably, the application of the antiseptic grass extract in the food preservation process can be used alone or in combination with other natural preservation methods to further improve the preservation effect of food. The natural plant antiseptic grass extract is obtained by a specific physical extraction method, including but not limited to pressing, distillation, and cold soaking.

[0020] Preferably, in the process of preserving vegetables, the antiseptic grass extract can be used in combination with specific packaging materials to enhance the preservation effect, reduce water loss, and extend the shelf life of vegetables. In the preservation of meat products, the antiseptic grass extract can inhibit bacterial growth, reduce odor generation, and maintain the fresh and tender texture of meat. The amount of the antiseptic grass extract used in the preservation of pastries should be controlled within a specific range to avoid affecting the taste and quality of the pastries.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] 1. The all-natural plant antiseptic grass extract of the present invention is used in the research and development of food preservation technology. The plant antiseptic grass extract is completely derived from natural plants and does not contain any artificially synthesized chemical preservatives, which greatly reduces the risk of chemical residues in food and provides consumers with safer and healthier food choices. At the same time, compared with traditional chemical preservatives, the natural plant antiseptic grass extract has no toxic side effects on the human body and will not burden the liver, kidneys and other organs of the human body. It is especially suitable for the preservation of food consumed by special groups such as children, pregnant women and the elderly;

[0023] 2. The all-natural plant antiseptic grass extract of the present invention is used in the research and development of food preservation technology. After research and development and experimental verification, it is found that the extract has a significant antiseptic effect, can effectively inhibit the growth and reproduction of microorganisms such as bacteria, molds and yeasts in food, and prolong the shelf life of food. While preserving food, it can better maintain the original color, flavor and nutritional components of the food, and will not have an adverse effect on the taste and quality of the food. For example, in the preservation of fruits, it can prevent the fruit from changing color, softening and losing flavor. In the preservation of meat, it can reduce the generation of odor and keep the meat fresh and tender;

[0024] 3. The all-natural plant antiseptic grass extract of the present invention is used in the research and development of food preservation technology. The raw materials of the plant antiseptic grass extract come from natural plants, grow rapidly and are renewable, and will not cause excessive exploitation and damage to the environment, which meets the requirements of sustainable development. In addition, when the extract is used in the food preservation process, even if there is a small amount of residue, it can be quickly degraded in the natural environment, and will not cause pollution to the soil, water source and other environments;

[0025] 4. The all-natural plant antiseptic grass extract of the present invention is used in the research and development of food preservation technology. Although a certain amount of investment may be required in the early stage of research and development, in the long run, the cost of natural plant antiseptic grass extract is relatively low. On the one hand, natural plant resources are abundant and the acquisition cost is relatively low. On the other hand, the loss caused by food spoilage is reduced and the overall production cost is reduced. At the same time, the good preservation effect can extend the sales cycle and shelf life of food, increase the economic benefits of food companies, meet consumers' demand for high-quality and safe food, and help enhance the market competitiveness of enterprises. DETAILED DESCRIPTION

[0026] Unless the context clearly dictates otherwise, unmodified nouns and nouns modified by "the" include singular and plural referents.

[0027] As used in the specification and claims, the terms "comprises," "comprising," "having," "may," "containing," and variations thereof as used herein refer to open transitional phrases, terms, or words that require the presence of specified ingredients / steps and allow for the presence of other ingredients / steps. However, such descriptions should be interpreted as also describing compositions or methods as "consisting of" and "consisting essentially of" the recited ingredients / steps, which allows for the presence of only the specified ingredients / steps and any unavoidable impurities that may result therefrom, and excludes other ingredients / steps.

[0028] Numerical values ​​in the specification and claims of this application should be understood to include the same numerical values ​​when reduced to the same number of significant figures and numerical values ​​that differ from the stated value by less than the experimental error of ordinary measurement techniques of the type described in this application for determining the stated value.

[0029] All ranges disclosed herein are inclusive of the indicated endpoints and are independently combinable (eg, the range of "2 grams to 10 grams" includes the endpoints 2 grams and 10 grams, and all intermediate values).

[0030] The terms "about" and "approximately" can be used to include any numerical value that can be varied without changing the basic function of the value. When used with a range, "about" and "approximately" also disclose the range defined by the absolute values ​​of the two endpoints, for example, "about 2 to about 4" also discloses a range of "2 to 4". Generally, the terms "about" and "approximately" can refer to ±10% of the indicated number. However, for temperature, the term "approximately" refers to ±1°C.

[0031] Unless expressly stated otherwise, the percentages of elements are to be considered as percentages by weight of the alloy in question.

[0032] The present disclosure may refer to the temperature of certain method steps. It should be noted that these specifications generally refer to the temperature set by the heat source (such as a furnace), and not necessarily the temperature that the heated material must reach.

[0033] The following description is used to disclose the present invention so that those skilled in the art can implement the present invention. The preferred embodiments described below are only examples, and those skilled in the art can think of other obvious variations. The basic principles of the present invention defined in the following description can be used for other embodiments, variations, improvements, equivalents, and other technical solutions that do not deviate from the spirit and scope of the present invention.

[0034] Example 1: Research and development of an all-natural plant antiseptic grass extract for food preservation technology, acute toxicity test of antiseptic grass:

[0035] Acute toxicity refers to a test method in which a large dose of a test substance is administered to test animals at one time to observe its toxic effects on test animals within a short period of time (generally 24 hours to 2 weeks), including general behavior and appearance changes, gross morphological changes, and death effects. Acute toxicity testing is the first step in understanding and studying the toxic effects of exogenous chemicals on the body. It usually uses oral, inhalation, or percutaneous exposure to observe the acute poisoning symptoms of rats and mice, and determine the median lethal dose (LD5o), thereby determining the acute toxicity of the test substance.

[0036] This experiment studied the acute toxicity of antiseptic grass. Based on the poisoning symptoms, toxicity intensity and mortality of mice, this experiment preliminarily evaluated the toxic effect characteristics, target organs, and dose-effect relationship of antiseptic grass on the body, and provided a preliminary reference for the dose selection in subsequent repeated dose toxicity tests.

[0037] 1. Materials:

[0038] Preservative grass (sample is extracted concentrated powder), preservative grass emulsion prepared by dry gel method (colostrum composition: oil: water: Tween = 2:2:1). Tween-80 (pharmacopoeia grade, CAS No.: 9005-65-6), distilled water.

[0039] Experimental animals: Sexually mature mice were selected. Feeding conditions: 12h light / 12h dark, 25±3℃, humidity 40%-60%, ventilation, free access to food and water. The experiment started after one week of adaptation.

[0040] 2. Methods:

[0041] 2.1 Preliminary test

[0042] Healthy mice were randomly divided into 3 groups, 4 mice in each group, half male and half female. The experimental mice were gavaged at 8:00 in the morning, and the administration volume was 0.2mL / 10g. Preliminary tests were repeated to determine the LDn (minimum lethal dose) and LDm (maximum lethal dose).

[0043] 2.2 Formal test

[0044] Referring to the acute toxicity test of toxicology, 50 healthy mice weighing 18-22g were randomly divided into 5 groups, 10 mice in each group, half male and half female, and fed in separate cages. Based on the preliminary test, 5 toxic dose groups were set up: 2970, 3375, 3850, 4385 and 5000 mg / kg. Each dose group was given a single oral gavage. After the administration, the feeding, drinking, breathing, activity and death of the mice were observed and recorded. The observation was continued for 14 days, and the LD50 and 95% confidence interval of the antiseptic grass were calculated by the modified Koch method. The mice that died during the test and were killed at the end of the observation period were autopsied, and pathological histological microscopy was performed if abnormal tissues or organs were found.

[0045] 3. Test results:

[0046] 3.1 Preliminary test results

[0047] In the preliminary experiment, the LDn value of the antiseptic grass was measured to be 2783 mg / kg, and the LDm value was 5000 mg / kg. According to the formula: The difference between the logarithms of adjacent doses was calculated to be 0.055, that is, the dose interval of the formal test was 1:0.83.3.2 Results of the formal test After administration, the activity of mice decreased. In severe cases, symptoms such as shortness of breath or difficulty breathing, mental depression, prone position, arched back, and loss of appetite appeared, and they died within 2h-8d. In mild cases, symptoms such as slow reaction, lack of energy, and decreased appetite appeared, and they returned to normal after 3-5d. The intensity of toxic reactions of animals in each dose group was positively correlated with the dose. Distension of the stomach and colon of dead mice was found in autopsy, and no lesions were visible to the naked eye in other organs. The mortality is shown in Table 1.

[0048] Table 1 Results of acute oral test of antiseptic grass on mice

[0049]

[0050]

[0051] Calculate LD5o according to the formula:

[0052] LD5o=4535.22mg / kg;

[0053] The 95% confidence limit of LD50 is 3565.3-4927.3rng / kg.

[0054] 4. Analysis and discussion:

[0055] Acute toxicity test is the first step in the toxicological safety evaluation of drugs, and it plays a role in understanding the acute toxicity of the test substance. LD5o is a classic indicator for detecting the acute toxicity of a substance, with good stability and high sensitivity. The larger the LD5o value, the weaker and smaller the acute toxicity of the test substance, and vice versa, the stronger and larger the acute toxicity. In this experiment, mice were gavaged with preservative grass, and the organs of the mice that died of acute poisoning were dissected without obvious lesions. The LD5o was 4535.22mg / kg, and the 95% confidence limit was 3565.3-4927.3rng / kg.

[0056] In the acute toxicity test of mice gavage with the preservative grass emulsion, the mice in each test group showed clinical symptoms such as mental depression, reduced activity and gradual paralysis of limbs. Its LD50 value is large, indicating that its toxicity is extremely low. According to the WTO acute toxicity classification standard for exogenous chemicals and the acute toxicity classification method of food toxicology in my country, preservative grass is a very slightly toxic substance.

[0057] Example 2: Research and development of an all-natural plant antiseptic grass extract for food preservation technology, and the cumulative toxicity test of the antiseptic grass on mice:

[0058] Cumulative toxicity refers to the functional or structural damage caused by repeated exposure or poisoning of exogenous chemicals at doses below the acute poisoning range, when the amount absorbed is greater than the amount excreted or the toxic effects are accumulated multiple times. The accumulation of exogenous chemicals in the body is the basis for causing subchronic toxicity and chronic toxicity. The cumulative toxicity test is a test method for evaluating the cumulative toxicity of foreign compounds. Commonly used methods include the increasing dose accumulation coefficient method and the fixed dose accumulation toxicity coefficient method.

[0059] This study used the fixed-dose cumulative toxicity coefficient method to conduct cumulative toxicity tests, studied the accumulation of antiseptic grass in mice, calculated the accumulation coefficient, determined the effects of antiseptic grass on the clinical manifestations, feed utilization and organ coefficients of mice, judged the strength of its cumulative toxicity, and provided a reference for dose selection in subacute toxicity and subchronic toxicity tests.

[0060] 1. Materials:

[0061] Preservative grass (sample is extracted concentrated powder), preservative grass emulsion prepared by dry gel method (colostrum composition: oil: water: Tween = 2:2:1). Tween-80 (pharmacopoeia grade, CAS No.: 9005-65-6), distilled water.

[0062] Experimental animals: Sexually mature mice were selected. Feeding conditions: 12h light / 12h dark, 25±3℃, humidity 40%-60%, ventilation, free access to food and water. The experiment started after one week of adaptation.

[0063] 2. Methods:

[0064] The fixed-dose cumulative toxicity coefficient method was used to evaluate the cumulative toxicity of antiseptic grass to mice. 40 mice were randomly divided into two groups: the test group and the negative control group, with 20 mice in each group, half male and half female. The mice in the test group were given a Tween-80 emulsion containing 907.4 mg / kg (1 / 5 LD5o) of antiseptic grass by oral gavage every day, and the mice in the negative control group were given the same dose of 2% Tween-80 solution. The test can be terminated when half of the mice in the test group have died. At this time, the cumulative total exposure dose LD5o(n) is calculated, and the cumulative coefficient Kcum value is calculated according to the formula: K=LD5o(n)LD5o for evaluation. If the cumulative exposure dose reaches 5 LD5o doses and the death of the mice in the test group has not reached half, the test can be terminated, and the conclusion of Kcum>5 is drawn at this time.

[0065] During the administration period, the spirit, appetite, drinking water, death and abnormal reactions of the mice were observed and recorded. The feed consumption of the mice was weighed every week and the feed utilization rate was calculated (feed utilization rate = total increment / total food intake x 100). After the experiment, all surviving animals were killed, and the heart, liver, spleen, kidney, lung, brain, etc. were collected and weighed to calculate the organ coefficient (organ coefficient = weight of each organ / body weight x 100).

[0066] 3. Results:

[0067] 3.1 Clinical manifestations and calculation of accumulation coefficient

[0068] After 28 days of administration, there was no death in the 20 mice in the control group and the test group. During the test, the mice had dry, soft and dirt-free fur; there was no abnormality in the eyes, the pupils were normal, there was no obvious secretion in the corners of the eyes, and there was no turbidity in the cornea and iris; the feces were black or light black in color, and round or oval in shape. After oral administration, there was no restlessness, no spasm, tremor, paralysis, convulsion or forced action, and no excitability; appetite, movement, fur finish and color, breathing, demeanor, etc. were all normal, and no obvious symptoms of poisoning were observed. No abnormal changes were found in the mice of each group by eye observation and pathological autopsy, and the accumulation coefficient Kcum>5 was determined. According to the accumulation toxicity evaluation standard, the accumulation toxicity of antiseptic grass is extremely mild accumulation or no accumulation.

[0069] 3.2 Impact on feed utilization

[0070] In the four weeks after the mice were treated with the antiseptic herbs, the feed utilization rate of the female mice in the test group was significantly lower than that of the female mice in the control group in the 2nd and 4th weeks (P<0.01), and the feed utilization rate of the male mice in the test group was significantly lower than that of the male mice in the control group in the 4th week (P<0.01). In the other time periods, the feed utilization rate of the test mice treated with the antiseptic herbs was not significantly different from that of the control group (P>0.05). This result shows that the administration of 907.4mg / kg (15LD5o) dose of antiseptic herbs to mice within 28 days has little effect on the feed utilization rate of mice.

[0071] 3.3 Impact on organ coefficients

[0072] The experiment on the effect of antiseptic grass on the organ coefficients of mice found that the organ coefficients of the heart, liver, spleen, lung, kidney, and brain of the experimental group of mice were not significantly different from those of the control group (P>0.05), which indicates that giving mice antiseptic grass within a dosage range of 907.4 mg / kg (1 / 5LD5o) and below within 28 days has no effect on the organ coefficients of mice.

[0073] 4. Analysis and discussion:

[0074] After exogenous chemicals enter the body, they are metabolically transformed and then excreted as metabolites or untransformed parent chemicals. However, when chemicals are used for repeated disinfection, and the speed or total amount of chemicals entering the body exceeds the speed of metabolic transformation and the speed or total amount of chemicals excreted from the body, the chemicals or their metabolites may gradually increase in the body and be stored in certain parts. This phenomenon is called the accumulation of chemicals. Most accumulations will produce cumulative toxicity. So far, there are almost no reports on the accumulation toxicity test of antiseptic grass at home and abroad. This test uses the fixed dose accumulation toxicity coefficient method to determine that its accumulation coefficient Kcum is greater than 5. According to the accumulation coefficient classification standard, antiseptic grass is a substance with basically no accumulation toxicity. This result can preliminarily determine the safety of the use of antiseptic grass, and provide a certain scientific basis for the selection of doses for subacute toxicity and subchronic toxicity tests and the development and utilization of products in the future.

[0075] Feed utilization rate refers to the number of grams of body weight gained by an animal for every 100 g of feed ingested. Feed consumption is a key parameter for determining the exposure dose during exposure to the contaminated feed, and it is also an important indicator of the toxic effect of chemical substances. Generally, drugs will affect the water intake and food intake of animals. When analyzing and evaluating the impact of drugs on animals' food intake, feed utilization rate is usually used for comparison. If preservative grass affects the animal's appetite, the daily food intake will decrease, and the body weight gain will be affected, but the feed utilization rate may not necessarily change. If preservative grass interferes with the absorption or metabolism of food by animals, although it may not necessarily affect appetite, the body weight gain will be very slow, and thus the feed utilization rate will change. There was no significant difference in the feed utilization rate between the experimental group and the control group measured in the cumulative toxicity test, indicating that within 28 days, giving mice preservative grass at a dose of 907.4 mg / kg (1 / 5 LD50) and below had no adverse effect on the feed utilization rate of mice (P>0.05).

[0076] The organ coefficient, also known as the organ / body ratio, refers to the mass of a certain organ wet weight in 100 g of body weight, and is generally used for parenchymal organs such as the heart, liver, spleen, lungs, and kidneys. After excluding the influence of factors such as water loss before weighing, age, gender, and malnutrition, if the organ coefficient increases, it indicates changes such as congestion, edema, hyperplasia, and hypertrophy; if the organ coefficient decreases, it indicates that the organ may have atrophy, degeneration, etc. The comparison result of the organ coefficient is regarded as an important indicator in drug toxicity tests, and this test result to a certain extent explains the degree of the impact of drug toxic effects and sometimes can support or confirm the results of pathological examinations. In the cumulative toxicity test of preservative grass, there was no significant difference in the organ coefficient between the experimental group of mice and the negative control group (P>0.05), indicating that within 28 days, giving preservative grass at a dose of 907.4 mg / kg (1 / 5 LD50) and below had no adverse effect on the organs of mice (P>0.05). The analysis and comparison of the feed utilization rate and organ coefficient of each group of mice verified to a certain extent that the determination of preservative grass as a low-toxic substance by the cumulative coefficient measured by the fixed-dose cumulative toxicity coefficient method was correct.

[0077] Example 3: R & D and application of a fully natural plant preservative grass extract in food preservation technology, subacute oral toxicity test of preservative grass on mice:

[0078] Subacute oral toxicity refers to the harmful effects on the health of the body caused by daily oral contact with the test substance in experimental animals within 14-28 days. The subacute toxicity test of antiseptic grass was carried out in accordance with the subacute test method of the Organization for Economic Cooperation and Development (OECD). This experiment intends to use mice as experimental subjects; according to the clinical medication period of antiseptic grass emulsion of 7 days, its subacute toxicity test period is determined to be 30 days; by observing the clinical manifestations, body weight, growth rate of the animals in the drug administration group, as well as the organ coefficient, blood physiological and biochemical indexes after sacrifice, and gross autopsy, histological structure and other observations, compared with the blank control group animals, in order to determine the toxic and side effects of antiseptic grass emulsion on mice, and provide a theoretical basis for further clinical drug trials.

[0079] 1. Materials:

[0080] Preservative grass (sample is extracted concentrated powder), preservative grass emulsion prepared by dry gel method (colostrum composition: oil: water: Tween = 2:2:1). Tween-80 (pharmacopoeia grade, CAS No.: 9005-65-6), distilled water.

[0081] Experimental animals: Select sexually mature mice. Feeding conditions: 12h light / 12h dark, 25±3℃, humidity 40%-60%, ventilation, free access to food and water. The experiment started after one week of adaptation.

[0082] 2. Methods:

[0083] 2.1 Trial Grouping

[0084] 80 healthy sexually mature mice were selected and randomly divided into 4 groups, including 3 dosage groups and 1 solvent control group, with 20 mice in each group.

[0085] 2.2 Dosage and route of administration

[0086] The subacute test method of the Organization for Economic Cooperation and Development (OECD) was used as reference. The test groups included a high-dose group (1 / 5LD5o, 907.4 mg / kg), a medium-dose group (1 / 10LD5o, 453.7 mg / kg), and a low-dose group (1 / 20LD5o, 226.8 mg / kg). The vehicle control group was given 2% Tween-80. The daily dosage of each medication group was calculated according to the average body weight of each group of animals. The gavage needle was disinfected by boiling water every day. Gavage was given once in the morning every day for 30 consecutive days, and the animals were allowed to eat and drink freely.

[0087] 2.3 Observation of clinical manifestations

[0088] During the experiment, the time, degree and duration of changes in the mice's fur, eyes, mucous membranes, respiratory system, nervous system, limb activity, behavior, etc. were observed and recorded every morning and evening. Dead animals or endangered animals were promptly autopsied to reduce cannibalism between animals and postmortem autolysis of tissues.

[0089] 2.4 Body weight and food intake

[0090] Feed or water consumption was measured daily, and body weight was measured weekly.

[0091] 2.5 Routine blood test

[0092] All mice were killed on the second day after the gavage period. 0.5 mL of blood was collected from the tail of the mouse with a syringe and injected into a vacuum sodium heparin anticoagulant tube for the determination of blood physiological indicators, including hemoglobin (HGB), red blood cells (RBC), platelets (PLT), white blood cells (WBC) and differential counts of neutrophils (GR.A), lymphocytes (LYM) and monocytes (MID).

[0093] 2.6 Blood biochemical test

[0094] After the tail blood was collected, the eyeballs of the mice were enucleated and 2.0 mL of blood was collected. The blood was placed in a clean centrifuge tube and allowed to stand at room temperature to precipitate the serum. The serum was centrifuged at 2000 r / min for 5 min and the serum was collected for the determination of blood biochemical indicators. The examination indicators included albumin (ALB), alkaline phosphatase (ALP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), urea nitrogen (BUN), total cholesterol (CHO), creatinine (CRE), blood glucose (GLU), total bilirubin (TBIL), triglycerides (TG) and total protein (TP) to understand the effects of antiseptic grass on the functions of organs such as liver and kidney.

[0095] 2.7 Organ coefficient determination

[0096] All animals were dissected to examine the body surface, openings of the body cavity, the skull, chest, abdominal cavity and their contents; the heart, liver, spleen, lungs, kidneys and other organs were weighed as soon as possible after separation to prevent moisture loss. The absolute weight of the organs was recorded and the relative weight was calculated (organ coefficient = organ weight / body weight × 100%).

[0097] 2.8 Histopathological examination

[0098] Animals that died during the experiment and animals that were killed after the experiment were autopsied and the relevant tissues and organs (heart, liver, spleen, lung, kidney, stomach, intestine, ovary) were fixed in pre-cooled 4% paraformaldehyde fixative. Tissue sections were made according to the routine paraffin section making procedure, and the pathological histological changes were observed under an optical microscope after hematoxylin-eosin (HE) staining.

[0099] 3. Results:

[0100] 3.1 Clinical manifestations

[0101] During the experiment, no mice in the control group and the experimental group died, and the weight gain of mice in each dose group was normal; the changes in the skin, fur, eyes, mucous membranes, and the respiratory system, circulatory system, nervous system, limb activities, and behavior of mice in the low- and medium-dose groups were normal, and no obvious clinical symptoms were shown; the mice in the high-dose group showed rough fur after 3.5 weeks of medication.

[0102] 3.2 Body weight and food intake

[0103] During the experiment, the mice in each group ate and drank water normally, and their weight continued to increase. Analysis showed that there was no significant difference in weight gain between the male and female mice in the antiseptic grass group and the control group (P>0.05).

[0104] Table 1 Effect of antiseptic grass on the body weight of mice in subacute toxicity test

[0105]

[0106] 3.3 Routine blood test

[0107] After subacute oral exposure to antiseptic grass, the blood routine test results of mice showed that compared with the control group, except for lymphocytes and platelets in each dose group, other hematological indicators were higher than the control group, but the differences were not significant (P>0.05) and had no statistical significance.

[0108] Table 2 Results of routine blood test in subacute toxicity test

[0109]

[0110] ANOVA analysis of variance showed that there was no significant difference in the blood routine parameters between the experimental group and the control group (P>0.05).

[0111] 3.4 Blood biochemical test

[0112] After subacute oral poisoning with antiseptic grass, blood biochemical tests of mice showed that compared with the control group, the serum blood glucose of mice in the high-dose group was lower than that of the control group, and the other liver and kidney indicators were higher than those of the control group; the albumin, total protein and blood glucose of the medium and low-dose groups were lower than those of the control group, and the other indicators were higher than those of the control group. However, statistical analysis showed that except for the total bilirubin of mice in the high and medium-dose groups, which was significantly different from that in the control group (P<0.01), there were no significant differences in other blood biochemical indicators of the other dose groups compared with the control group (P>0.05).

[0113] 3.5 Organ coefficient

[0114] During the experiment, the experimental mice were killed, and no obvious abnormal lesions and injuries were found in the heart, liver, spleen and other organs. Compared with the control group, the organ coefficients of each dose group were not significantly different (P>0.05); among them, the liver and kidney organ coefficients of each dose group increased to varying degrees compared with the control group, but the differences were not significant (P>0.05), which was not statistically significant.

[0115] Table 3 Results of organ coefficient determination in subacute toxicity test

[0116]

[0117] ANOVA analysis of variance showed that there was no significant difference in the coefficients of various organs of mice in each dose group compared with those in the control group (P>0.05).

[0118] 3.6 Histopathological examination

[0119] The heart, liver, spleen, lung, kidney, duodenum, testis and ovary of mice in each dose group and control group sampled on the 30th day were examined by pathological histology. The results are as follows:

[0120] Liver: The liver lobules of the control group had normal structures, and the liver cells were arranged in a cord-like structure. The central veins of the livers of the mice in the low, medium, and high dose groups of the experimental group were congested, the liver cells were swollen and rounded, and different degrees of granular and vesicular degeneration occurred.

[0121] Spleen: The control group had normal spleen tissue structure, and the white pulp, red pulp and spleen trabeculae were clearly discernible. The high, medium and low dose groups of the experimental group all showed red pulp congestion, with a large number of macrophages and Langerhans cells infiltrating.

[0122] The staining results of kidney sections of mice in each dose group of the experimental group showed varying degrees of congestion in the glomerular capillaries and tubular interstitial capillaries, varying degrees of granular degeneration of the tubular epithelial cells, resulting in narrowing of the tubular lumen, and the presence of red filamentous substances in the lumen of some tubules.

[0123] Testis: The control group had normal testicular tissue structure, normal testicular interstitium, seminiferous tubules and spermatogenic cells at all levels, and a large number of mature sperm in the tubular lumen; Experimental group: Each dose group showed relatively normal spermatogenic cells at all levels in the seminiferous tubules, with a large number of mature sperm.

[0124] Histopathological examination of the heart, lungs, duodenum, and ovaries showed no obvious pathological damage.

[0125] 4. Analysis and discussion:

[0126] Routine blood test is one of the most basic and important laboratory tests in clinical practice. When pathological changes occur in the blood, it often affects tissues and organs throughout the body. By observing the changes in the number and morphological distribution of red blood cells, white blood cells and platelets in the blood, as one of the important bases for clinical diagnosis and treatment of diseases, it is possible to judge the infection and damage of exogenous chemicals to the body. The results of this study showed that the effect of subacute poisoning of antiseptic grass on the blood routine indicators of mice was not significantly different from that of the control group. The results of routine blood test showed that antiseptic grass had no effect on the components of the blood system of mice. Alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) in serum are the main indicators reflecting liver function. When liver cells are damaged, ALT and AST escape from the damaged liver cells in large quantities, thereby increasing their content in serum. The results of this study showed that the alanine aminotransferase (ALT), aspartate aminotransferase (AS) and alkaline phosphatase (ALP) of mice in the experimental group were slightly increased compared with those in the control group, indicating that antiseptic grass had very slight damage to the liver and bile duct obstruction in mice. When the liver is damaged by inflammation, necrosis, poisoning, and bile duct disease and hemolytic disease, the serum total bilirubin level will increase significantly. The results of this study showed that compared with the control group, the serum total bilirubin level of mice in the experimental group increased, but there was no dose-effect relationship, indicating that the increase in the serum total bilirubin test result was caused by hemolysis of blood samples due to careless experimental operation.

[0127] The organ index refers to the ratio of a certain organ to unit body weight, also known as the organ / body ratio. The organ index is an important indicator to measure the functional status of animals, and is also a designated test item for drug toxicity testing. An increase in the organ index indicates congestion, edema, or hyperplasia of the organ; a decrease in the organ index indicates organ atrophy and other degenerative changes. The results of this study showed that the liver and kidney organ indexes of mice in the high-dose group were higher than those in the control group, but there was no statistical significance, indicating that the liver and kidneys of mice were very slightly enlarged.

[0128] The results of pathological histological observations on the internal organs of mice in each dosage group showed that the antiseptic grass mainly affected the liver and kidneys. It mainly caused very mild vascular congestion in the liver and kidneys. In addition, antiseptic grass caused the appearance of Langerhans cells and macrophages in the spleen tissue of mice, indicating that antiseptic grass has a certain anti-inflammatory effect and is a protective response of the body to external stimuli. The research results confirmed that it has anti-inflammatory effects.

[0129] Example 4: A natural plant antiseptic grass extract was used in the research and development of food preservation technology. Subchronic oral toxicity test of antiseptic grass on mice:

[0130] Subchronic oral toxicity refers to the health damage effects caused by repeated daily oral exposure of test animals to the test sample during part of their life span (not exceeding 10% of the life span).

[0131] Through repeated oral exposure to the test chemical over a certain period of time, the toxic effects caused by the test sample are understood, and the maximum subchronic oral no-effect dose of the test substance is obtained, providing a basis for determining the dose of chronic toxicity tests and preliminarily calculating the safe level of human exposure.

[0132] 1. Materials:

[0133] Preservative grass (sample is extracted concentrated powder), preservative grass emulsion prepared by dry gel method (colostrum composition: oil: water: Tween = 2:2:1). Tween-80 (pharmacopoeia grade, CAS No.: 9005-65-6), distilled water.

[0134] Experimental animals: Sexually mature mice were selected. Feeding conditions: 12h light / 12h dark, 25±3℃, humidity 40%-60%, ventilation, free access to food and water. The experiment started after one week of adaptation.

[0135] 2. Methods:

[0136] The subchronic test method of the Organization for Economic Cooperation and Development was used. According to the median lethal dose data (LD504535.22mg / kg) obtained from previous laboratory studies, 96 healthy mice were randomly divided into 4 groups, 24 mice in each group, half male and half female. The high, medium and low dose groups were given preservative grass-Tween emulsions containing 226.7, 75.6 and 25.2mg / kg (1 / 20-1 / 180LD50) of preservative grass, respectively, and the control group was given 2% Tween-80. The drug solution was administered at 0.2mL / 10g body weight, once / d for 90 consecutive days.

[0137] Samples were collected on the 30th, 60th and 90th day after administration, and 6 mice were killed in each group, half of them were male and half were female. After the end of the experiment, the observation was resumed for 30 days, and the remaining mice were killed. The observation and measurement indicators were the same as those in the subacute toxicity test. After the end of intragastric administration, the hearts, livers, spleens, lungs, kidneys, testes and ovaries of mice in each group were collected, and routine HE staining was performed. Observation under a light microscope: The livers, kidneys and testes of mice were collected from the control group and the high-dose group. Imm 3 Cut into pieces, pre-fixed in 2.5% glutaraldehyde phosphate buffer and post-fixed in 1% osmium acid, dehydrated in acetone gradient, embedded in epoxy resin 618, sliced ​​into 50 nm thin sections, double stained with uranyl acetate and poly(lead) sulfate, and observed by TEM2000 transmission electron microscope.

[0138] 3. Results:

[0139] 3.1 Effects on mouse body weight

[0140] During the experiment, no obvious abnormal reactions were observed in the animals in each group, and their drinking and eating were basically normal. There were no abnormalities in the animals' fur, feces, and activities. Starting from the 6th week of exposure, the weight gain of male and female mice in each experiment slowed down. The weight of the high-dose group was lower than that of the control group, and the weight of the medium and low-dose groups was higher than that of the control group, but the differences were not significant. When the exposure reached 10 weeks, the weight of the high-dose group was significantly lower than that of the control group. The weight of mice in the medium and low-dose groups was equivalent to or higher than that of the control group at various times, but there was no significant difference in weight between the groups. The 13th to 16th week was a recovery observation period, and the weight of mice in each group remained basically at the same level.

[0141] 3.2 Effects on blood routine tests in mice

[0142] After 90 days of subchronic oral administration of antiseptic grass and 30 days of intragastric administration, there were no significant differences in the test indicators of each dose group compared with the control group (P>0.05). After 60 days of intragastric administration, compared with the control group, the percentage of neutrophils in the high and medium dose groups was higher than that in the control group, and the percentage of lymphocytes was lower than that in the control group, but the difference was not significant (P>0.05), and there were no differences in other indicators. After 90 days of intragastric administration, compared with the control group, the percentage of neutrophils in the high dose group was significantly higher than that in the control group, and the percentage of lymphocytes was lower than that in the control group, and there were significant differences (P<0.05). The percentage of neutrophils and lymphocytes was similar to that of intragastric administration for 60 days, and the difference was not significant; there were no significant differences in other indicators compared with the control group. After 30 days of recovery observation, there were no statistical differences in the indicators of each dose group compared with the control group.

[0143] 3.3 Effects on mouse blood biochemistry

[0144] After subchronic oral administration of antiseptic grass for 90 days, the detection of blood biochemical indexes of mice showed that compared with the control group, the ALB level in the serum of the high-dose group was decreased, and the ALP, ALT, AST, BLIN, CRE, and TBIL were increased 60 days after oral administration, with significant or extremely significant differences (P<0.05 or P<0.01). After 90 days of oral administration, the changes in the levels of serum liver and kidney index enzymes were consistent with those at 60 days.

[0145] There were no significant changes in serum liver and kidney indices of mice in the medium and low dose groups compared with those in the control group during the experimental period. There were no significant changes in serum biochemical indices of mice in the high dose group compared with those in the control group at 30 days.

[0146] 3.4 Histopathological examination

[0147] Subchronic poisoning with antiseptic grass did not cause obvious pathological damage to the heart, spleen, lungs, testicles and ovaries of mice. The pathological examination results of the liver and kidneys were as follows:

[0148] Liver: Under the light microscope, the liver lobule structure of the mice in the control group was intact, the hepatocytes were closely arranged, the nuclear staining was darker, and the structure was normal. Experimental group: The lobule structure of the mice in the low-dose group was intact, and the hepatocyte cords were arranged normally; the lobule structure of the mice in the medium-dose group was intact, the hepatocyte cords were slightly disordered, and the central veins and hepatic sinusoids were slightly congested; the central veins of the hepatic lobules of the mice in the high-dose group were congested, the hepatic sinusoids were dilated and congested, the hepatocytes were slightly granular and vacuolar, and the infiltration of inflammatory cells such as lymphocytes and neutrophils was visible.

[0149] Kidney: Under light microscopy, the kidneys of mice in the control group had normal nephron structure, and the glomeruli, proximal tubules and distal tubules were clearly discernible. Experimental groups: In the low-dose group, the tubular interstitial capillaries were hemolyzed, and the tubular epithelial cell granules were very slightly degenerated; in the medium-dose group, the glomerular capillaries and tubular interstitial capillaries were congested, and the glomerular epithelial cells were slightly swollen; in the high-dose group, rod cells and neutrophil infiltration were observed in the glomeruli.

[0150] 3.5 Ultrastructural observation of target organs

[0151] Under electron microscopy, the effects of antiseptic grass on the ultrastructure of mouse liver cells and renal tubular epithelial cells were as follows:

[0152] Liver: Under electron microscopy, in the control group, the organelle structure of liver cells was normal, the nuclear membrane was intact, and the nuclear chromatin was evenly distributed. In the experimental group, the mitochondria were extremely slightly deformed; the rough endoplasmic reticulum expanded and became cystic, proliferated, fractured and degranulated; the nucleus was abnormal in morphology, and the chromatin in the nucleus was condensed.

[0153] Kidney: Under electron microscopy, the mitochondria in the cytoplasm of renal tubular epithelial cells in the control group were abundant, with normal size and structure, and were clearly lamellar; the ultrastructure of the rough endoplasmic reticulum and the scattered ribosomes was normal; the nucleus was round, the nuclear membrane was flat, and the chromatin in the nucleus was uniform. In the experimental group, the mitochondria were extremely slightly swollen, containing myelin-like inclusions, with a slightly blurred structure, and a very small part was vacuolar degeneration.

[0154] 4. Analysis and discussion:

[0155] The results of the effect of subchronic poisoning with antiseptic grass on the weight of mice showed that after a certain dose of antiseptic grass was used to subchronically poison mice, the weight growth of mice was significantly inhibited, while the weight of mice in the medium and low dose groups was equivalent to or even higher than that of the control group at each time period. The results of this study showed that lower doses of antiseptic grass had no effect on the weight growth of mice or promoted the growth of mice. As the dose increased, the drug showed an inhibitory effect on the weight growth of mice. The possible reason is that high doses of preservatives affect the digestion and absorption of food.

[0156] Routine blood test is one of the most basic and important laboratory tests in clinical practice, which can determine the infection and damage of exogenous chemicals to the body. The results of this study showed that compared with the control group, the percentage of neutrophils in the blood of mice in the preservative grass dosage group increased, the lymphocytes decreased relatively, and the total number of white blood cells did not increase significantly compared with the control group. This study results show that the experimental mice have mild acute infection or tissue damage.

[0157] The detection of blood biochemical indexes is one of the important detection indicators of new drug safety evaluation test, which is of great significance for determining the dose and target organ of drug toxicity. The blood biochemical indexes of healthy mice are usually relatively stable. According to the changes in blood biochemical indexes and the relationship between the degree of change and the dose, the nature and degree of the test substance on the animal's pathological changes can be reflected. The dose groups were significantly higher than the control group, indicating that high doses of antiseptic grass can cause very slight damage to the renal function of mice, resulting in a decrease in glomerular filtration capacity. The lowest dose of the experimental mice at which the damage was observed was 75.6 mg / kg, and the dose at which no harmful effects were observed, that is, the maximum no-effect dose, was 25.2 mg / kg.

[0158] Example 5: Research and development of a natural plant antiseptic grass extract for food preservation technology, and study on the mutagenicity of antiseptic grass on mice:

[0159] Mutagenicity study is a necessary item in toxicological evaluation. Through mutagenicity test, mutagens of somatic cells and germ cells can be identified, potential carcinogens can be predicted, and environmental mutagens can be monitored and evaluated. Mouse bone marrow micronucleus test and sperm abnormality test are both in vivo mutagenicity tests, which detect the effects of foreign compounds on chromosome damage and mutagenicity on germ cells respectively. They are easy to operate, economical, practical and rapid, and are important test contents for drug safety evaluation. This experiment used mouse bone marrow polychromatic erythrocyte micronucleus test and sperm abnormality test to explore the mutagenic effect of antiseptic grass on experimental mice, providing experimental data for the full development and safe use of antiseptic grass.

[0160] 1. Materials:

[0161] Preservative grass (sample is extracted concentrated powder), preservative grass emulsion prepared by dry gel method (colostrum composition: oil: water: Tween = 2:2:1). Tween-80 (pharmacopoeia grade, CAS No.: 9005-65-6), distilled water.

[0162] Experimental animals: Sexually mature mice were selected. Feeding conditions: 12h light / 12h dark, 25±3℃, humidity 40%-60%, ventilation, free access to food and water. The experiment started after one week of adaptation.

[0163] method:

[0164] 2.1 Mouse bone marrow polychromatic erythrocyte micronucleus test

[0165] Animal grouping and administration: 40 mice aged 7-12 weeks were randomly divided into 5 groups, 8 mice in each group. The test group was gavaged with 1 / 5, 1 / 10, 1 / 20 LD50 preservative grass-Tween 80 emulsion, the positive control group was intraperitoneally injected with 40 mg (kgd) of cyclophosphamide, and the negative control group was orally gavaged with 2% Tween-80. The oral gavage was used for 30 hours, that is, the test substance was given twice within 24 hours, and the mice were killed by dislocation of the neck 6 hours after the second administration. The femurs on both sides of the hind legs of the mice were washed out with calf serum to remove the bone marrow fluid, and then smeared, fixed with methanol, and stained with Giemsa stain. Observed under a low-power microscope, selected the area with complete cells, uniform dispersion, and appropriate coloring, and then observed and counted under an oil microscope.

[0166] 2.2 Mouse sperm abnormality test

[0167] Experimental animal grouping and administration: 40 male mice were randomly divided into 5 groups, 8 mice in each group. The dosage of each group was the same as that of the micronucleus test. The mice in the experimental group 1 / 5, 1 / 10, 1 / 20 LD5o group and the negative control group were gavaged once a day, and the positive control group was intraperitoneally injected with cyclophosphamide once a day for 5 consecutive days. On the 35th day after the first day of administration, the mice were killed by cervical dislocation, and the bilateral testicles and epididymis were removed, and their wet weights were weighed and recorded with a precision electronic balance, and the organ coefficient (organ coefficient = organ wet weight / body weight X100) was calculated, and epididymal sperm smears were performed, and the deformity rate was statistically examined under a microscope.

[0168] Sperm deformity rate: Place the epididymis in a centrifuge tube containing 1 mL of saline, cut the epididymis longitudinally 3-5 times with ophthalmic scissors, let it stand for 5-10 minutes, shake it gently two or three times, filter it with a 300-mesh nylon filter membrane according to the national standard GB15193.7-2003, absorb the filtrate smear, stain it with 1% eosin Y for 1 hour, rinse it gently with running water, and dry it naturally. Under a low-power microscope, find an area with a clear background and few sperm overlaps, use an oil immersion lens to sequentially check the sperm morphology, and count the sperm with complete structure and deformity. Record the number of sperm deformities (fat head, no hook, banana type, double head, double tail mixed, tail folded, amorphous, small head, double body, broken tail, tail folded, etc.), and count 1000 complete sperm for each mouse.

[0169] The sperm deformity rate is calculated as follows:

[0170] Sperm abnormality rate (%) = total number of sperm abnormalities / total number of sperm examined × 100

[0171] 3. Results:

[0172] 3.1 Using the Gimsa staining method, polychromatic erythrocytes (PCE) appear gray-blue, and orthochromatic erythrocytes (NCE) appear orange-yellow. Typical micronuclei are mostly single, round, with smooth and neat edges, and the color is consistent with the nucleoplasm, appearing purple-red or blue-purple.

[0173] Microscopic observation and counting showed that the micronucleus rates of the high, medium and low dose groups of antiseptic grass, the negative control group and the positive control group were 4.12%, 3.37%, 3.21%, 3.17% and 28.5%, respectively. The difference in micronucleus rates between the positive control group and the high, medium and low dose groups was extremely significant (P<0.01), which was statistically significant; the difference in micronucleus rates between the negative control group and the high, medium and low dose groups was not statistically significant (P>0.05), indicating that antiseptic grass had no significant effect on the incidence of micronuclei in polychromatic erythrocytes in the bone marrow of mice.

[0174] 3.2 Effects of antiseptic grass on testis, attachment mass and organ coefficient of mice

[0175] Compared with the negative control group, the epididymis and testis organ mass indexes in the positive control group were significantly reduced (P<0.05); while there were no significant differences in the epididymis organ mass index and testis organ mass index between the three doses of antiseptic grass and the negative control group (P>0.05).

[0176] 3.3 Effect of antiseptic grass on sperm morphology (deformity rate) in mice

[0177] Sperm deformities in mice include no hook, banana shape, fat head, amorphous, tail folded, double head, double tail, etc. Sperm morphology, normal sperm and common abnormal sperm morphology were observed under a microscope. Microscopic observation and counting showed that the sperm deformity rates in the high, medium and low dose groups of antiseptic grass, the negative control group and the positive control group were 3.29%, 3.22%, 3.30%, 2.93% and 18.1%, respectively. There was no significant difference in the sperm deformity rate induced by each dose group of antiseptic grass compared with the negative control group (P>0.05); while the sperm deformity rate induced by cyclophosphamide was significantly increased, and the difference was significant compared with the negative control group (P<0.01). The positive control group and the high, medium and low dose groups of antiseptic grass had no significant effect on mouse sperm deformity.

[0178] 4. Analysis and discussion:

[0179] The formation of micronuclei is a genetic endpoint manifestation of cells after being acted upon by genetic toxins. Changes in the micronucleus rate can reflect the mutagenic toxicity of the tested drugs on experimental animals. It has become one of the rapid and sensitive conventional methods for screening the genetic toxicity of chemicals, and is also a necessary test for evaluating the toxicological safety of compounds such as pesticides, new drugs, and food additives. In this study, after gavage of three doses of antiseptic grass to experimental mice, there was no significant difference in the micronucleus rate of polychromatic erythrocytes in the bone marrow of mice in each dose group compared with the negative control group, and they were all significantly lower than the micronucleus rate of polychromatic erythrocytes in the bone marrow induced by the positive control cyclophosphamide (P<0.05). This result shows that antiseptic grass has no significant effect on the micronucleus rate of polychromatic erythrocytes in the bone marrow of mice.

[0180] Bone marrow is an important hematopoietic organ in the body. All blood cells originate from the bone marrow after birth. The multipotent stem cells in the bone marrow first differentiate into myeloid stem cells, and further differentiate into erythroid cells, monocytic cells, and granulocytes. Under normal microenvironmental factors, the erythroid system evolves into erythrocytes after proerythrocytes*proerythrocytes*intererythrocytes*late erythrocytes*denucleation; all cells undergoing division, such as under the action of chromosome clastogens, the acentric fragments of chromatids or chromosomes or the entire chromosome lost due to spindle injury, remain in the cytoplasm in the late stage of division, or the material protrudes outward after the nuclear membrane is damaged, forming one or several regular round or oval bodies, with similar chromatin to the cell nucleus and smaller micronuclei than the main nucleus. Therefore, detecting the changes in the micronucleus rate of bone marrow polychromatophilic erythrocytes (PCE) can detect whether the test substance has the ability to induce mutagenesis; the results of this experimental study showed that antiseptic grass had no obvious effect on the micronucleus rate of mouse bone marrow polychromatophilic erythrocytes, indicating that antiseptic grass does not have the effect of inducing cell chromosomes.

[0181] The organ coefficient is a sensitive indicator of the toxicity of chemical substances to the body0091, and the reproductive system is very sensitive to the effects of chemical poisons. The reproductive system may have been damaged before other systems have become toxic. Therefore, the level of the epididymis organ coefficient and the testis organ coefficient can reflect the degree of damage to the reproductive system by chemical poisons. The results of this study showed that there was no significant difference in the epididymis organ coefficient and the testis organ coefficient of mice in each dose group of antiseptic grass compared with the negative control group, and they were significantly lower than the positive control group, indicating that antiseptic grass did not have a damaging effect on the reproductive system at the organ level.

[0182] Sperm formation is controlled by polygenic inheritance. When a compound causes a related gene to mutate, sperm morphology will become abnormal, leading to an increase in sperm deformity. The level of sperm deformity can reflect the reproductive toxicity of the chemical poison and its potential mutagenicity to germ cells. The results of this study showed that there was no significant difference in the sperm deformity rate of mice in each dose group of antiseptic grass compared with the control group, while the deformity rate was significantly lower than that of the positive control cyclophosphamide group, indicating that antiseptic grass had no mutagenic induction effect on the germ cells of male adult mice at the dose (1 / 5-1 / 20LD5o) given in the experiment.

[0183] Specific conclusions:

[0184] 1. The results of acute toxicity tests showed that the oral median lethal dose LD50 of antiseptic grass for mice (both male and female) was 4535.22 mg / kg, and the 95% confidence limit was 3565.3-4927.3 rng / kg. According to the acute toxicity classification standards for exogenous chemicals, antiseptic grass is an extremely low toxic substance.

[0185] 2. The results of the cumulative toxicity test show that the accumulation coefficient Kcum of antiseptic grass is greater than 5. According to the accumulation coefficient classification standard, antiseptic grass is judged to be a substance with basically no cumulative toxicity.

[0186] 3. Subacute toxicity test Through the correlation results of blood biochemical indicators and organ coefficients, it can be preliminarily determined that the subacute toxic organs of antiseptic grass to mice are mainly liver and kidney.

[0187] 4. The pathological damage caused by subchronic poisoning of antiseptic grass is mainly cell granule degeneration and water

[0188] The saturation is reversible. The changes in the ultrastructure mainly include mitochondrial vacuolization and endoplasmic reticulum fragmentation accompanied by ribosome shedding. Combining the results of pathological histological examination and ultrastructural analysis, it was concluded that the lowest dose of antiseptic grass to observe harmful effects on experimental mice was 75.6 mg / kg; the dose without observed harmful effects, that is, the maximum no-effect dose, was 25.2 mg / kg.

[0189] Through the results of the micronucleus test on polychromatic erythrocytes in mouse bone marrow and the sperm deformity test, it was concluded that antiseptic grass has no genetic toxicity to male mice and it is safe to use antiseptic grass within the prescribed dosage range.

[0190] Study on the antibacterial activity of antiseptic grass

[0191] The antibacterial activity of the samples submitted for inspection was studied as follows:

[0192] 1. Materials:

[0193] Antiseptic grass (sample is extracted concentrated powder), Staphylococcus aureus, Escherichia coli, Streptococcus agalactiae, Staphylococcus epidermidis. TSB broth, McFarland turbidimeter tube, LB plate culture medium, MRS broth, L-shaped wave rod, Oxford cup, sterile ceramic cover, sterile test tube, micropipette, sealed fermentation tube, phosphate buffer, etc.

[0194] 2. Test methods:

[0195] 2.1 Preparation of test samples: purified antiseptic grass, penicillin, and crude extract of antiseptic grass: Dissolve purified antiseptic grass, penicillin, and crude extract of antiseptic grass in distilled water respectively, make up to volume, and prepare relevant solutions separately for use.

[0196] 2.2 Preparation of culture medium: The composition of the culture medium is: peptone, yeast extract, agar powder, distilled water, adjust to about 400 μl, heat and boil to fully dissolve the agar, divide into portions, sterilize under high pressure at 121°C, and keep in the refrigerator for later use.

[0197] 2.3 Preparation of culture plates: In a clean bench, place the sterilized nutrient agar medium in an electric oven to make it liquid. Pour the medium into sterilized culture dishes while it is still hot. Pour the medium into each culture dish and wait for it to solidify before use. Sterilize the materials to be used.

[0198] 2.4 Minimum inhibitory concentration test, using the two-fold dilution method, the purified preservative grass, penicillin, and crude extract of preservative grass were diluted to a series of concentrations, and the concentrations were measured and placed in nutrient broth test tubes, sterilized and set aside. Then add the diluted bacterial suspension μ and culture in a 37°C constant temperature oscillating box. Since the crude extract of the medicinal materials used is dark in color and the observation results are inaccurate, the test tubes that are relatively clear and preliminarily judged as having no bacterial growth will be marked. The liquid is absorbed and spread on the plate culture medium, and inverted in a 37°C constant temperature incubator to observe the growth of the colonies. The minimum inhibitory concentration is the concentration of the drug solution corresponding to the culture plate with no colony growth.

[0199] 2.5 Inhibition zone test: The antibacterial activity of the purified antiseptic grass, penicillin solution and crude antiseptic grass extract on different bacteria can be judged by the size of the inhibition zone. The antibacterial activity was determined by the Oxford cup method. Take the prepared culture plate, absorb the μ bacterial suspension and add it into it, use a spreader to evenly apply various bacterial solutions to make a bacterial plate, use sterile tweezers to clamp the Oxford cup that has been sterilized by high-pressure steam and dried, and gently place it on the surface of the culture plate coated with the bacterial solution. Each culture plate is divided into three parts and an Oxford cup is placed. Use a sterile pipette to transfer 200μ samples of the purified antiseptic grass, penicillin solution, and crude antiseptic grass extract into the Oxford cup. Make three parallel plates for each solution, and slowly move the culture plate into a constant temperature incubator and culture at 37°C. Take out the cultured plates and observe the growth of the colonies in each plate, that is, whether there is a transparent circle near the Oxford cup. If there is, use a vernier caliper to measure the diameter of the inhibition zone and record the data. An inhibition zone diameter greater than 300 indicates that the bacteria are highly sensitive to the drug; an inhibition zone diameter of 500 indicates that the bacteria are moderately sensitive; an inhibition zone diameter less than 600 indicates low sensitivity; and no inhibition zone indicates insensitivity.

[0200] Activation of strains: Before activating the strains, first turn on the ultraviolet lamp to sterilize the clean bench. Take various strains stored on the slant, pick the strains with a bamboo stick and draw snake-shaped lines on the culture medium on the slant of the test tube, and culture the inoculated strains in a 37℃ constant temperature incubator. Rejuvenate two strains of each strain to restore the vitality of the strains.

[0201] 2.6 Preparation of bacterial solution: Pick two loops of each activated strain and add them to sterilized distilled water to make a bacterial suspension. Dilute the prepared bacterial suspension with sterilized distilled water in turn to make the concentration of the strain reach. Each type of bacteria is prepared in this way and labeled for use.

[0202] 3. Results:

[0203] 3.1 Inhibition zone test

[0204] Table 1 Inhibition zone test results

[0205] Medicinal material samples Staphylococcus aureus Escherichia coli Streptococcus agalactiae Staphylococcus epidermidis Purified antiseptic grass 17 25 19 14 Penicillin solution 25 9 22 27 Crude extract of antiseptic grass 7 12 10 7

[0206] The antibacterial effect of the purified antiseptic grass, penicillin solution and crude antiseptic grass extract on different bacteria can be judged by the size of the inhibition zone, as shown in the table. We can see that the purified antiseptic grass has a certain inhibitory effect on the four bacteria, especially the most significant inhibitory effect on Escherichia coli. In addition, compared with the inhibitory effect of the crude antiseptic grass extract on the four bacteria, the antibacterial activity of the purified antiseptic grass is significantly improved. Compared with the citrus penicillin solution, the antibacterial effect of the purified antiseptic grass is slightly worse, but the diameter of the inhibition zone of the penicillin solution is not much different, and the inhibitory effect on the four bacteria can reach about the same as the antibacterial effect of the penicillin solution.

[0207] 3.2 Minimum inhibitory concentration test

[0208] Medicinal material samples Staphylococcus aureus Escherichia coli Streptococcus agalactiae Staphylococcus epidermidis Purified antiseptic grass 23 12.8 25 95 Penicillin solution 1.09 6.7 0.82 0.46 Crude extract of antiseptic grass 49 26 52 98

[0209] The results of the minimum inhibitory concentration test of the purified antiseptic grass, penicillin solution and crude antiseptic grass extract on four bacteria, Staphylococcus aureus, Escherichia coli, Streptococcus agalactiae and Staphylococcus epidermidis, are shown in the table. The purified antiseptic grass has the most obvious inhibitory effect on Escherichia coli, with a minimum inhibitory concentration of. Compared with the minimum inhibitory concentration of the crude antiseptic grass extract, the purified antiseptic grass is much lower and can show a good antibacterial effect. Compared with the minimum inhibitory concentration of penicillin solution, the minimum concentration of the purified antiseptic grass is much higher, but in the previous inhibition zone test, the size of the inhibition zone is not much different, indicating that the purified antiseptic grass has a certain antibacterial effect.

[0210] Test results:

[0211]

[0212] The 50 main compounds predicted by HPLC-MS / MS analysis are as follows:

[0213]

[0214]

[0215]

[0216]

[0217] The specific steps are as follows: the staff first determines the plant species with antiseptic properties, and after a lot of screening and research, selects a specific antiseptic grass variety, conducts a place of origin survey on the selected antiseptic grass to ensure that its growth environment is pollution-free, free of pesticide residues and excessive heavy metals, and at the same time establishes strict procurement standards, and only selects antiseptic grass with good growth and moderate maturity for subsequent processing. The staff also tries different organic solvents, such as ethanol, acetone, ethyl acetate, etc., to compare the extraction effects. After determining the best solvent, optimize the extraction temperature, time, solvent concentration and other parameters. For example, conduct a gradient experiment within a certain temperature range to determine the optimal extraction temperature; observe the changes in extract yield and antiseptic activity by changing the extraction time; adjust the solvent concentration to find a solvent that can both ensure extraction efficiency and reduce The optimal ratio for low cost, and finally the extracted solution is filtered and concentrated to remove impurities and excess solvent to obtain a preliminary extract. Next, supercritical fluids such as carbon dioxide are selected as extractants to study the effects of different pressures, temperatures and flow rates on the extraction effect, and optimize the extraction process parameters to ensure the purity and preservative activity of the extract. For example, gradually increase the pressure and observe the changes in extract yield and quality; adjust the temperature to find the optimal extraction temperature range; control the flow rate to make the extraction process more efficient. Finally, the staff collects the extracted extract for subsequent processing and analysis, and uses column chromatography to purify the extract. Suitable adsorbents, such as silica gel, alumina, etc. are selected, and the components with preservative activity are separated by adjusting the composition and flow rate of the eluent., and then test the purified extract to ensure that its purity meets the requirements. For example, use analytical methods such as high performance liquid chromatography (HPLC) to detect the content and purity of the active ingredients in the extract. At the same time, the staff determines the appropriate addition method for different types of food. For fruits and vegetables, spraying, dipping, etc. can be used to evenly cover the extract on the surface of the food. For meat and aquatic products, an appropriate amount of extract can be added during the processing process, or vacuum packaging combined with extract treatment can be used. For dairy products and beverages, the extract can be added directly to the product, or the extract can be slowly released into the food through dipping of the packaging material. Then, the effect of different addition amounts on the preservation effect of food is studied experimentally. For example, storage experiments are carried out on foods treated with different amounts of extracts to observe indicators such as microbial growth, color changes, and texture changes. Finally, the staff comprehensively considers the preservation effect and cost factors to determine the optimal addition amount. Generally speaking, too little addition may not achieve the ideal preservation effect, while too much addition may affect the taste and quality of the food, and also increase the cost. At the same time, the staff will combine it with traditional preservation methods such as low-temperature storage, vacuum packaging, and modified atmosphere packaging to further improve the preservation effect of food, and study the synergistic effect of different preservation measures, optimize the preservation process parameters, for example, determine the best storage temperature and humidity conditions, as well as suitable packaging materials and packaging methods, and conduct a comprehensive quality assessment of foods treated with extracts, including testing microbial indicators in food, such as total bacteria, mold and yeast counts, etc., to ensure that the food meets hygiene standards, and evaluate the sensory quality of food, including color, smell, taste and texture. It can be evaluated by a professional sensory evaluation team, or by instrumental analysis methods, such as colorimeter, texture analyzer, etc., and monitor the changes in physical and chemical indicators of food during storage, such as pH value, moisture content, vitamin content, etc. Finally, if the staff finds that the preservation effect is not ideal, they can adjust the extraction process parameters to improve the purity and preservative activity of the extract; or optimize the preservation process, such as changing the addition method and addition amount, combining other preservation measures, etc., and will continue to improve and perfect the preservation process to meet the preservation needs of different foods and improve the market competitiveness of products.

[0218] Those skilled in the art should understand that the embodiments of the present invention described above and shown in the accompanying drawings are only examples and do not limit the present invention. The purpose of the present invention has been fully and effectively achieved. The functions and structural principles of the present invention have been demonstrated and explained in the embodiments. Without departing from the principles, the embodiments of the present invention may have any deformation or modification. Although the embodiments of the present invention have been shown and described, it is understood by those of ordinary skill in the art that various changes, modifications, substitutions and deformations can be made to these embodiments without departing from the principles and spirit of the present invention. The scope of the present invention is defined by the attached claims and their equivalents. This specific embodiment is only an explanation of the present application, and it is not a limitation of the present application. After reading this specification, those skilled in the art can make modifications to the present embodiment without creative contribution as needed, but as long as they are within the scope of the claims of the present application, they are protected by the patent law.

Claims

1. A fully natural plant antiseptic grass extract for use in food preservation technology research and development, characterized in that: The following steps are involved: Step 1. Selection of plant antiseptic grass: determine the plant species with antiseptic properties, select specific antiseptic grass varieties after extensive screening and research, conduct origin surveys on the selected antiseptic grass to ensure that its growth environment is pollution-free, free of pesticide residues and excessive heavy metals, establish strict procurement standards, and only select antiseptic grass with good growth and moderate maturity for subsequent processing; Step 2, determination of the extraction method: try different organic solvents, after determining the best solvent, optimize the extraction temperature, time and solvent concentration and other parameters, conduct gradient experiments within a certain temperature range, determine the most suitable extraction temperature, observe the changes in the extract yield and preservative activity by changing the extraction time, adjust the solvent concentration, find the best ratio that can both ensure extraction efficiency and reduce costs, filter and concentrate the extracted solution, remove impurities and excess solvents, obtain preliminary extracts, select supercritical fluids such as carbon dioxide as extractants, study the effects of different pressures, temperatures and flow rates on the extraction effect, optimize the extraction process parameters, ensure the purity and preservative activity of the extract, gradually increase the pressure, and observe the changes in the extract yield and quality; adjust the temperature to find the most suitable extraction temperature range, control the flow rate to make the extraction process more efficient, collect the extracted extracts, and conduct subsequent processing and analysis; Step 3, purification of the extract: purify the extract by column chromatography, select a suitable adsorbent, separate the components with antiseptic activity by adjusting the composition and flow rate of the eluent, test the purified extract to ensure that its purity meets the requirements, and use high performance liquid chromatography (HPLC) and other analytical methods to detect the content and purity of the active ingredients in the extract; Step 4, application of food preservation technology: determine the appropriate addition method for different types of food, conduct experiments to study the effects of different addition amounts on food preservation, conduct storage experiments on foods treated with different addition amounts of extracts, observe their microbial growth, color changes, texture changes and other indicators, comprehensively consider preservation effects and cost factors, and determine the best addition amount. Generally speaking, too little addition may not achieve the ideal preservation effect, while too much addition may affect the taste and quality of the food, and also increase costs. Combined with traditional preservation methods such as low-temperature storage, vacuum packaging, and modified atmosphere packaging, further improve the preservation effect of food, study the synergistic effect of different preservation measures, optimize preservation process parameters, determine the best storage temperature and humidity conditions, as well as suitable packaging materials and packaging methods; Step 5. Effect evaluation and optimization: Detect microbial indicators in food to ensure that the food meets hygiene standards and evaluate the sensory quality of the food. This can be done by a professional sensory evaluation team, or by using instrumental analysis methods to monitor changes in physical and chemical indicators of food during storage. If the preservation effect is found to be unsatisfactory, the extraction process parameters can be adjusted to improve the purity and preservative activity of the extract, or the preservation process can be optimized, continuously improved and perfected to meet the preservation needs of different foods and enhance the market competitiveness of products.

2. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The organic solvent includes ethanol, acetone, ethyl acetate, etc., and the adsorbent includes silica gel, alumina, etc.

3. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The adding methods include spraying or dipping for fruits and vegetables to evenly cover the extract on the food surface; adding an appropriate amount of extract during the processing of meat and aquatic products, or using a vacuum packaging combined with extract treatment method; and adding the extract directly to dairy products and beverages, or slowly releasing the extract into the food through dipping of the packaging material. The microbial indicators include total bacteria count, mold and yeast count, etc.

4. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The sensory qualities of the food include color, smell, taste and texture, etc., the instrumental analysis method includes a colorimeter, a texture analyzer, etc., the physical and chemical indicators include pH value, water content, vitamin content, etc., and the preservation process includes the addition method, addition amount, and combination with other preservation measures, etc.

5. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The 50 main compounds predicted by the antiseptic herbal materials include isoanisole, kalinol, anisole, 5-methoxypsoralen, isofragrutin, Da-2-methylpiperidine, amino acids, angelica root root, dihydrosperquinone, piceatannol, kaempferol, betaine, angelica root root lactone, isofraxin, hypoxanthine, toxicostearin, 3-hydroxycoumarin, ferulic acid, dihydroeucalyptus alcohol, Primarol, cyperquinone, cnidium monnieri, cyperus lactone, diosmin, flavonoid glycoside, isoeugenol, garcinol, anemonin, chlorogenic acid, isochlorogenic acid, neochlorogenic acid, eugenol, camphor, isopulegone, L-pineneketone, myrtenol, menthone, norarecoline, isonepetalactone, artemisinic acid, cordycepin, danshensu, bergamotol, linolenic acid, costus alcohol, caltropone, purpurogenol and coumarin.

6. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The antiseptic grass extract is obtained through a specific extraction process, which includes but is not limited to one or more combinations of solvent extraction, supercritical fluid extraction, and ultrasonic-assisted extraction. The antiseptic grass extract can be used to preserve various foods, including but not limited to fruits, vegetables, meat, dairy products, pastries, etc. The extraction technology includes ultrasonic extraction, microwave extraction, supercritical fluid extraction, etc.

7. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The antiseptic grass extract is added to food in a specific concentration and manner to achieve an effective preservation effect. The specific concentration and addition method are adjusted according to different types of food. During the food preservation process, the antiseptic grass extract can effectively inhibit the growth and reproduction of microorganisms such as bacteria, molds and yeasts, extend the shelf life of the food, and at the same time maintain the original color, flavor and nutritional components of the food.

8. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The raw material plants of the antiseptic grass extract are natural plant resources that can be grown sustainably. The extraction and application process is environmentally friendly and will not cause pollution. The process of using the antiseptic grass extract for food preservation includes the steps of extraction, purification, and addition to food. Each step is carried out under specific conditions to ensure the activity and preservation effect of the extract.

9. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: The application of the antiseptic grass extract in the food preservation process can be used alone or in combination with other natural preservation methods to further improve the preservation effect of food. The natural plant antiseptic grass extract is obtained by a specific physical extraction method, including but not limited to pressing, distillation, and cold soaking.

10. The all-natural plant antiseptic grass extract according to claim 1 is used in the research and development of food preservation technology, characterized in that: In the process of preserving vegetables, the antiseptic grass extract can be used in combination with specific packaging materials to enhance the preservation effect, reduce water loss, and extend the shelf life of vegetables. In the preservation of meat products, the antiseptic grass extract can inhibit bacterial growth, reduce odor generation, and maintain the fresh and tender texture of meat. The amount of the antiseptic grass extract used in the preservation of pastries should be controlled within a specific range to avoid affecting the taste and quality of the pastries.

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