CN114128792A - Method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil - Google Patents
Method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil Download PDFInfo
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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- A23K30/00—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs
- A23K30/10—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder
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Abstract
The invention discloses a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil, which sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, and (3) fumigating. Has the advantages that: the invention provides a method for inhibiting high-moisture alfalfa storage mildewing by fumigating plant essential oil, which achieves the effects of preventing and controlling the mildews and inhibiting the growth of the mildews, ensures the stable quality of the hay bales in the storage time, avoids deterioration, has good safety and does not bring the problem of environment; does not affect the health of livestock, has higher safety and simple and convenient operation.
Description
The technical field is as follows:
the invention relates to the technical field of hay mildew prevention, in particular to a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil.
Background art:
alfalfa, as a forage grass with global cultivation, wide adaptability and excellent quality, has become an important part for the development of the grass industry and the animal husbandry in China. In the process of harvesting the alfalfa, high moisture is bundled to a certain degree, so that the drying time can be shortened, the number of leaves is saved to the maximum extent, the damage of rainfall to the quality of the hay is reduced, and the method has practical significance for solving the contradiction that the alfalfa hay is prepared in rainy seasons. However, high moisture content baling inevitably leads to mold growth during hay storage, and higher alfalfa hay moisture content is more conducive to fungal growth and faster bale mildew growth. Therefore, the development of the alfalfa high-moisture bale mildew preventive is a main technical means for reducing the activity of harmful microorganisms and reducing the loss of nutrient substances.
Starting in the 80 s of the last century, the focus of domestic and foreign research on hay harvest was to establish preservation specifications by means of chemical treatment methods. At present, the mildew preventive mainly comprises organic acids, ammonium compounds, urea, biological agents and the like, such as sodium diacetate, propionic acid, ammonium propionate, urea, ammonia water and the like. The main function of these fungicides is to inhibit the fungi, rather than kill them, so that a sufficient concentration of the fungicide must be maintained throughout storage. And the products have large irritation and strong corrosivity, the residue is not easy to solve, the wide use can cause environmental pollution, and the toxic action on animals and human bodies can be caused to a certain extent.
The invention content is as follows:
the invention aims to provide a method for inhibiting high-moisture alfalfa storage mildews by fumigating with plant essential oil, which has the advantages of good mildewing inhibition effect, no pollution and simple and convenient operation.
The purpose of the invention is implemented by the following technical scheme: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1-4:5-1 to obtain a biological mildew preventive;
and (2) airing and bundling: naturally airing the harvested alfalfa until the water content is 20-25%, and bundling the hay by using mechanical equipment, wherein the bundling density is 120-3Obtaining dry straw bales;
fumigating in step (3): placing the dry grass bales in the step (2) into a closed room, then placing the biological mildew preventive in the step (1) around the dry grass bales, and then volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.8-1L/m3And fumigating the dry grass bales for 72-96 h.
Further, the step (1) can also be: mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1-4:5-1, adding the mixture into gelatin solution at 35-40 ℃, stirring for 30-40min, then placing the mixture into a refrigerator at 4-6 ℃ for cooling and settling for 24-30h, pouring out supernatant, placing the remainder into a freeze dryer at-50 ℃ to-60 ℃ for freeze drying, and finally obtaining the solid biological mildew preventive.
Further, the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is preferably 4: 1.
Further, the method for preparing the gelatin solution comprises the following steps:
(a) dissolving gelatin: adding 100-200g of gelatin into 1-2L of distilled water, wherein the temperature of the distilled water is 60-70 ℃, and then stirring until the gelatin is completely dissolved to obtain a gelatin hot solution;
(b) and (3) cooling: and naturally cooling the hot gelatin solution to 35-40 ℃ to obtain the gelatin solution.
Further, in the step (3), 1-2 bundles of the dry grass bundles are placed in each cubic meter of the closed room.
Further, the environmental humidity in the room in the step (3) is controlled to be below 40%, and if the humidity is greater than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the hay bales is influenced.
The invention has the advantages that:
1. the invention provides a method for inhibiting high-moisture alfalfa storage mildew by utilizing plant essential oil fumigation, which utilizes the extracted mixture of natural litsea cubeba essential oil and cinnamon essential oil to fumigate hay bales, achieves the effects of preventing and controlling mildew and inhibiting the growth of the mildew, ensures the stable quality of the hay bales in the storage time, avoids deterioration, has strong volatility and no residue, and has good safety without bringing the problem of environment on the premise of controlling the dosage; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
2. The invention adopts the fumigation technology to promote the biological mildew preventive to act on the hay bales in a form of fast and uniform dispersion in a closed room, effectively inhibits the hay bales from mildewing, and can control the quality reduction of the hay bales in the storage process to a certain extent.
3. The treatment method has higher safety and simple and convenient operation.
Description of the drawings:
FIG. 1 is a photograph showing the growth of Fusarium in different proportions of Litsea cubeba essential oil and Cinnamomi corex essential oil, wherein a is a photograph showing the growth of Fusarium in a control group; b is a photograph of the growth of fusarium with cinnamon essential oil added; c is a picture of the growth condition of fusarium with the litsea cubeba essential oil only; d is a picture of the growth condition of fusarium when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 1: 1; e, taking a picture of the growth condition of fusarium when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 1: 3; f is a photograph of the growth condition of fusarium when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 1: 4.
FIG. 2 is a cell structure diagram of Fusarium, wherein a is a cell structure diagram of Fusarium treated with biological mildew preventive, and b is a cell structure diagram of Fusarium in control group.
FIG. 3 is a photograph showing the growth of Aspergillus flavus in different proportions of Litsea cubeba essential oil and Cinnamomi corex essential oil; wherein a is a photo of the growth condition of the aspergillus flavus in the control group, and b is a photo of the growth condition of the aspergillus flavus when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 4: 1.
The specific implementation mode is as follows:
the present invention will be described in further detail by way of examples.
Example 1: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 2:3, adding the mixture into a gelatin solution at 37 ℃, stirring for 35min, then placing the mixture into a refrigerator at the temperature of 5 ℃ for cooling and settling for 27h, pouring out supernatant, placing the remainder in an environment at the temperature of-55 ℃, and freeze-drying to obtain a solid biological mildew preventive; the solid biological mildew preventive has long preservation time; wherein the method of preparing the gelatin solution comprises the steps of:
(a) dissolving gelatin: 150g of gelatin was added to 1.5L of distilled water at a temperature of 65 ℃, and then stirred until the gelatin was completely dissolved, to obtain a hot gelatin solution.
(b) And (3) cooling: and naturally cooling the hot gelatin solution to 37 ℃ to obtain the gelatin solution.
The hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
And (2) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 23%, bundling the hay with mechanical equipment, wherein the bundling density is 135kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the bales of hay from step (2) in a closed room, wherein 2 bales of hay are placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.9L/m3Fumigating the dry bales for 84 h; the environmental humidity in the room is controlled to be below 40%, and if the humidity is higher than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry bales is influenced; by adopting the fumigation technology, the biological mildew preventive acts on the hay bales in a form of rapidly and uniformly dispersing in a closed room, so that the hay bales are effectively inhibited from mildewing, and the quality of the hay bales in the storage process can be controlled to be reduced to a certain extent.
The treatment method has higher safety and simple and convenient operation.
Example 2: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1:5, adding the mixture into a gelatin solution at 40 ℃, stirring the mixture for 40min, then placing the mixture into a refrigerator at the temperature of 6 ℃ for cooling and settling for 30h, pouring out supernatant, placing the remainder in an environment at the temperature of-60 ℃ for freeze drying, and finally obtaining a solid biological mildew preventive; the solid biological mildew preventive has long preservation time; wherein the method of preparing the gelatin solution comprises the steps of:
(a) dissolving gelatin: 200g of gelatin was added to 2L of distilled water at a temperature of 70 ℃ and then stirred until the gelatin was completely dissolved, to obtain a hot gelatin solution.
(b) And (3) cooling: and naturally cooling the hot gelatin solution to 40 ℃ to obtain the gelatin solution.
The hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
And (2) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 25%, bundling the hay with mechanical equipment, wherein the bundling density is 150kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the bales of hay from step (2) in a closed room, wherein 2 bales of hay are placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 1L/m3Fumigating the dry grass bales for 96 h; controlling the humidity of the room to be below 40%, and controlling the moisture in the air to affect the essential oil if the humidity is higher than 40%The volatilization effect is good, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry straw bales is influenced; by adopting the fumigation technology, the biological mildew preventive acts on the hay bales in a form of rapidly and uniformly dispersing in a closed room, so that the hay bales are effectively inhibited from mildewing, and the quality of the hay bales in the storage process can be controlled to be reduced to a certain extent.
The treatment method has higher safety and simple and convenient operation.
Example 3: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to a volume ratio of 4:1, adding the mixture into a gelatin solution at 35 ℃, stirring for 30min, then placing the mixture into a refrigerator at 4 ℃ for cooling and settling for 24h, pouring out supernatant, placing the remainder in an environment at-50 ℃ for freeze drying, and finally obtaining a solid biological mildew preventive; the solid biological mildew preventive has long preservation time; wherein the method of preparing the gelatin solution comprises the steps of:
(a) dissolving gelatin: 100g of gelatin was added to 1L of distilled water at a temperature of 60 ℃ and then stirred until the gelatin was completely dissolved, to obtain a hot gelatin solution.
(b) And (3) cooling: and naturally cooling the hot gelatin solution to 35 ℃ to obtain the gelatin solution.
The hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
Step (2) airingAnd (3) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 20%, bundling the hay with mechanical equipment, wherein the bundling density is 120kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the hay bundles of step (2) in a closed room, wherein 1 bale of the hay bundle is placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.8L/m3Fumigating the dry grass bales for 72 h; the environmental humidity in the room is controlled to be below 40%, and if the humidity is higher than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry bales is influenced; by adopting the fumigation technology, the biological mildew preventive acts on the hay bales in a form of rapidly and uniformly dispersing in a closed room, so that the hay bales are effectively inhibited from mildewing, and the quality of the hay bales in the storage process can be controlled to be reduced to a certain extent.
The treatment method has higher safety and simple and convenient operation.
Example 4: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 2:3 to obtain a biological mildew preventive; the hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
And (2) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 23%, bundling the hay with mechanical equipment, wherein the bundling density is 135kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the bales of hay from step (2) in a closed room, wherein 2 bales of hay are placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.9L/m3Fumigating the dry bales for 84 h; the environmental humidity in the room is controlled to be below 40%, and if the humidity is higher than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry bales is influenced; by adopting the fumigation technology, the biological mildew preventive acts on the hay bales in a form of rapidly and uniformly dispersing in a closed room, so that the hay bales are effectively inhibited from mildewing, and the quality of the hay bales in the storage process can be controlled to be reduced to a certain extent.
The treatment method has higher safety and simple and convenient operation.
Example 5: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1:5 to obtain a biological mildew preventive; the hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
And (2) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 25%, bundling the hay with mechanical equipment, wherein the bundling density is 150kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the bales of hay from step (2) in a closed room, wherein 2 bales of hay are placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 1L/m3Fumigating the dry grass bales for 96 h; the environmental humidity in the room is controlled to be below 40%, and if the humidity is higher than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry bales is influenced; by adopting the fumigation technology, the biological mildew preventive acts on the hay bales in a form of rapidly and uniformly dispersing in a closed room, so that the hay bales are effectively inhibited from mildewing, and the quality of the hay bales in the storage process can be controlled to be reduced to a certain extent.
The treatment method has higher safety and simple and convenient operation.
Example 6: a method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil sequentially comprises the following steps: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to a volume ratio of 4:1, mixing to obtain the biological mildew preventive; the hay bundle is fumigated by the extracted mixture of the natural litsea cubeba essential oil and the cinnamon essential oil, so that the effects of preventing and controlling mold and inhibiting the growth of the mold are achieved, the quality stability of the hay bundle in the storage time is ensured, the deterioration is avoided, meanwhile, the mixture of the natural litsea cubeba essential oil and the cinnamon essential oil has strong volatility and no residue, and under the premise of controlling the dosage, the safety is good, and the problem of environment is not caused; meanwhile, the litsea cubeba essential oil and the cinnamon essential oil belong to plant essential oil, so that the litsea cubeba essential oil has no side effect after being used by livestock, and the health of the livestock cannot be influenced.
And (2) airing and bundling: naturally drying the harvested alfalfa grass until the water content is 20%, bundling the hay with mechanical equipment, wherein the bundling density is 120kg/m3If the bundling density is too loose, air enters the interior of the dry straw bundle to provide a proper propagation environment for the mold in the storage process, the mold is easy to occur, the bundling is too dense, the biological mildew preventive cannot fumigate the interior of the dry straw bundle, and the mildew-proof effect is poor;
fumigating in step (3): placing the hay bundles of step (2) in a closed room, wherein 1 bale of the hay bundle is placed in each cubic meter of the closed room; then placing the biological mildew preventive in the step (1) around the dry grass bales, and volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.8L/m3Fumigating the dry grass bales for 72 h; the environmental humidity in the room is controlled to be below 40%, and if the humidity is higher than 40%, the moisture in the air influences the volatilization effect of the essential oil, so that the volatilization effect of the biological mildew preventive is poor, and the fumigation effect on the dry bales is influenced; by adopting the fumigation technology, the biological mildew preventive is acted on the hay bundle in a form of promoting the biological mildew preventive to be rapidly and uniformly dispersed in a closed room, so that the hay bundle is effectively inhibited from mildewingAnd the quality of the hay bales can be controlled to be reduced to a certain extent in the storage process.
The treatment method has higher safety and simple and convenient operation.
Experiment 1: mildew inhibiting effect of mixed essential oil
Separating different moulds such as fusarium, aspergillus flavus, rhizopus, penicillium and mucor from alfalfa, selecting 2 strains of toxin-producing strains fusarium and aspergillus flavus to perform an essential oil mildew inhibition test, taking purified bacteria to cultivate on a potato-glucose agar medium (PDA) for 120h at 25 ℃, repeatedly burning a puncher on flame for three times under an aseptic condition, respectively punching holes at the edge of a single bacterial colony of a prepared bacterium-containing flat plate after cooling, transferring the bacterial colony to a potato-glucose liquid medium (PDB), cultivating for 120h at 25 ℃, taking out and shaking for 2h, filtering by double-layer gauze to obtain a spore suspension, calculating the concentration of the spore suspension by using a plate counting method, then diluting to about 106cfu/mL, preparing the bacterium-containing flat plate by using a pouring method, respectively dipping essential oil, bacillus subtilis and mucor with different concentrations on a cover of a culture dish, The biological mildew preventive and a filter paper sheet without dipping any object are cultured for 120 hours at 25 ℃ with the diameter of 1cm, the growth of the mold and the size of a bacteriostatic circle are observed, and the filter paper sheet without dipping any object is used as a control group. And taking the minimum essential oil concentration with the inhibition rate of 100% within 72h as the Minimum Inhibitory Concentration (MIC), and continuing culturing for 48h, wherein the essential oil concentration without growing bacteria is the minimum inhibitory concentration (MFC).
1.1 investigation of the fungicidal Effect of Individual essential oils
The MIC results of litsea cubeba essential oil against fusarium and aspergillus flavus are shown in table 1: the MICs of Litsea cubeba essential oil to Fusarium and Aspergillus are respectively 0.15L/m3And 1.25L/m3The MIC results of cinnamon essential oil to fusarium and aspergillus are shown in table 2: MIC is 0.08L/m respectively3And 0.9L/m3。
TABLE 1 minimum inhibitory concentration of Litsea cubeba essential oil
In Table 1, "-" indicates sterile growth and negative, "+" indicates growth and positive, and "+" indicates positive.
TABLE 2 minimum inhibitory concentration of cinnamon essential oil
In Table 2, "-" indicates sterile growth and negative, "+" indicates growth and positive, and "+" indicates positive.
1.2 study of the mold-inhibiting effect of the biological mildew inhibitor
The MIC results of the mixed litsea cubeba essential oil and cinnamon essential oil biological mildewcide on fusarium and aspergillus flavus are shown in table 3: the MICs of the biological mildew inhibitor mixed by the litsea cubeba essential oil and the cinnamon essential oil to fusarium and aspergillus are respectively 0.04L/m3And 0.8L/m3。
Table 3 minimum inhibitory concentrations of Complex essential oils
In Table 5, "-" indicates sterile growth and negative, "+" indicates growth and positive, and "+" indicates positive.
The addition amount of the biological mildew inhibitor mixed by the litsea cubeba essential oil and the cinnamon essential oil is 0.02L/m3In the meantime, the inhibition effect of the litsea cubeba essential oil and cinnamon essential oil in different ratios on fusarium is expressed by the diameter of the inhibition zone, and the results are shown in table 4 and fig. 1.
TABLE 4 inhibition effect of biological mildew preventive on Fusarium
As can be seen from table 4, with the increase of the ratio of the cinnamon essential oil, the inhibition effect of the biological mildew inhibitor on fusarium tends to increase, and when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil increases by 1:5, the inhibition effect of the biological mildew inhibitor on fusarium tends to decrease, so that when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 1:4, the inhibition effect of the biological mildew inhibitor on fusarium is most obvious and is significantly higher than the single action effect of the cinnamon essential oil, which indicates that the cinnamon essential oil has a dominant inhibition effect on fusarium, and the increase of the litsea cubeba essential oil can increase the inhibition effect of the cinnamon essential oil on fusarium, and the two essential oils have a synergistic effect when mixed.
The clear part in fig. 1 is a bacteriostatic circle, wherein fig. 1b has a significant bacteriostatic circle compared with fig. 1c, so that the cinnamon essential oil has a more significant inhibitory effect on fusarium than the litsea cubeba essential oil; however, compared with fig. 1a, fig. 1c has a significant inhibition zone, so the litsea cubeba essential oil also has an inhibition effect on fusarium; as can be seen from fig. 1d to fig. 1f, as the proportion of the cinnamon essential oil increases, the inhibition zone is larger, so that as the proportion of the cinnamon essential oil increases, the inhibition effect of the biological mildew inhibitor on fusarium tends to increase, and when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 1:4, the inhibition effect of the biological mildew inhibitor on fusarium is most obvious.
The addition amount of the biological mildew inhibitor mixed by the litsea cubeba essential oil and the cinnamon essential oil is 0.02L/m3When the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 4:1, the damage of the biological mildew inhibitor to fusarium cells is shown in detail in figure 2a, and the fusarium cell of a control group is shown in figure 2 b.
As can be seen from FIG. 2, the morphological structure of Fusarium in the control group was normal under the electron microscope; the nucleus of the cell is irregular polygon, the chromatin is distributed evenly, the euchromatin is the main, and the nuclear membrane is clear and complete; the cytoplasm has uniform electron density, can see organelles such as mitochondria, ribosome and the like, has complete and clear structure, and has more lipid drops in the cytoplasm; the cell wall and the cell membrane are complete in structure; the fusarium after the biological mildew preventive is adopted is suspected to be necrotic, the chromatin of cell nuclei is lost, the cytoplasmic contents are lost, and only a small amount of residual swelling mitochondria and a small amount of ribosomes are seen; cell wall and cell membrane dissolution disappears under an electron microscope, the cell wall and the cell membrane become discontinuous, and the thalli are subjected to plasmolysis; therefore, when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is 4:1, the fusarium can be killed.
The addition amount of the biological mildew inhibitor mixed by the litsea cubeba essential oil and the cinnamon essential oil is 0.4L/m3In the meantime, the inhibition effect of the litsea cubeba essential oil and cinnamon essential oil on aspergillus flavus is shown in table 5 and fig. 3.
TABLE 5 inhibitory Effect of biological mildewcides on Aspergillus flavus
As can be seen from Table 5, when the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is changed from 1: 0 to 1:4, the bacteriostatic effect of the litsea cubeba essential oil slightly increases with the increase of the volume ratio of the cinnamon essential oil, but the increase difference is not obvious; continuously changing from 5:1 to 0:1, wherein when the ratio of 5:1 to 4:1 is changed, the antibacterial effect of the composition obviously increases along with the decrease of the volume ratio of the litsea cubeba essential oil, but when the ratio of 4:1 to 0:1 is changed, the antibacterial effect of the composition obviously decreases along with the decrease of the volume ratio of the litsea cubeba essential oil, so that the volume ratio of the litsea cubeba essential oil and the cinnamon essential oil with the best antibacterial effect is 4:1, which is obviously higher than that of the essential oil after being treated independently, and the synergistic effect of the litsea cubeba essential oil and the cinnamon essential oil can obviously increase the inhibition effect on the aspergillus flavus; the amount of the biological mildew preventive required for inhibiting the fusarium is small, so that the aspergillus flavus is mainly considered, and the amount of the biological mildew preventive required for inhibiting the aspergillus flavus can inhibit and kill the fusarium.
It can be seen from fig. 3a that the aspergillus flavus of the control group without the added essential oil diffuses to the four sides, and it can be seen from fig. 3b that when the volume ratio of the litsea cubeba essential oil and the cinnamon essential oil in the added biological mildew preventive is 4:1, the aspergillus flavus does not diffuse to the four sides, so that the litsea cubeba essential oil and the cinnamon essential oil have an inhibiting effect on the aspergillus flavus.
Experiment 2: application of mildew-inhibiting effect on alfalfa hay bundles
The alfalfa variety used in the test is Zhongmu No. 2, and the Saerqin base alfalfa test field of the grassland research institute of Chinese agricultural academy of sciences is planted in 3 months in 2017. Mechanically mowing 7-16 days in 2021, then processing by the method in the step 2 in the embodiment 3 of the invention to obtain 10 bundles of hay bundles, averagely dividing the hay bundles into two parts which are respectively an experimental group and a control group, placing the control group in a No. 1 hay storage warehouse for 3 months, then taking out and measuring the mould number, dry matter, crude protein and relative feeding value of each bundle of hay bundles, and calculating to obtain an average value; the experimental group is continuously treated by the method in the step 3 in the embodiment 3 of the invention, and is taken out from a closed room after the treatment is finished, then is placed in the No. 2 grass storage with the storage environment parameters consistent with those of the No. 1 grass storage for 3 months, is taken out, measures the mould quantity, dry matters, crude proteins and relative feeding values of each bundle of dry grass, and calculates to obtain an average value; the results are shown in Table 6.
TABLE 6 quality of stored bales of hay and changes in mold thereon
As can be seen from Table 6, the number of the mold in the experimental group is significantly lower than that in the control group, and the mold growth rate is reduced by 99.7%; in terms of quality, the loss of the dry matter of the experimental group is reduced by 3.2% compared with that of the control group, the loss of the relative feeding value of the experimental group is reduced by 4.5% compared with that of the control group, and the change difference of other quality indexes is not obvious, so that the dry straw bale is acted on the dry straw bale in a form of promoting the biological mildew preventive to be rapidly and uniformly dispersed in a closed room by adopting a fumigation technology, the mildew of the dry straw bale is effectively inhibited, and the quality reduction of the dry straw bale in the storage process can be controlled to a certain extent.
The foregoing is a preferred embodiment of the present invention, and it will be apparent to those skilled in the art that various modifications and adaptations can be made without departing from the principles of the invention, and such modifications and adaptations are intended to be included within the scope of the invention.
Claims (6)
1. A method for inhibiting high-moisture alfalfa storage mildew by fumigating plant essential oil is characterized by sequentially comprising the following steps of: (1) preparing a biological mildew preventive, (2) airing and bundling, (3) fumigating;
wherein:
preparing a biological mildew preventive in step (1): mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1-4:5-1 to obtain a biological mildew preventive;
and (2) airing and bundling: naturally airing the harvested alfalfa until the water content is 20-25%, and bundling the hay by using mechanical equipment, wherein the bundling density is 120-3Obtaining dry straw bales;
fumigating in step (3): placing the dry grass bales in the step (2) into a closed room, then placing the biological mildew preventive in the step (1) around the dry grass bales, and then volatilizing the biological mildew preventive at normal temperature to enable the concentration of the biological mildew preventive in the air in the room to reach 0.8-1L/m3And fumigating the dry grass bales for 72-96 h.
2. The method for inhibiting high-moisture alfalfa storage mold by fumigating with plant essential oil according to claim 1, wherein the step (1) can be further comprising: mixing litsea cubeba essential oil and cinnamon essential oil according to the volume ratio of 1-4:5-1, adding the mixture into gelatin solution at 35-40 ℃, stirring for 30-40min, then placing the mixture into a refrigerator at 4-6 ℃ for cooling and settling for 24-30h, pouring out supernatant, placing the remainder into a freeze dryer at-50 ℃ to-60 ℃ for freeze drying, and finally obtaining the solid biological mildew preventive.
3. The method for inhibiting high-moisture alfalfa storage mold by fumigating with plant essential oil according to claim 1 or 2, wherein the volume ratio of the litsea cubeba essential oil to the cinnamon essential oil is preferably 4: 1.
4. The method for inhibiting high-moisture alfalfa storage mold by fumigating with plant essential oil according to claim 2, wherein the method for preparing the gelatin solution comprises the following steps:
(a) dissolving gelatin: adding 100-200g of gelatin into 1-2L of distilled water, wherein the temperature of the distilled water is 60-70 ℃, and then stirring until the gelatin is completely dissolved to obtain a gelatin hot solution;
(b) and (3) cooling: and naturally cooling the hot gelatin solution to 35-40 ℃ to obtain the gelatin solution.
5. The method for inhibiting high-moisture alfalfa storage mold by fumigating with plant essential oil according to claim 1, characterized in that in step (3), 1-2 bales of the dry grass are placed in each cubic meter of the closed room.
6. The method for inhibiting high-moisture alfalfa storage mold by fumigating with plant essential oil as claimed in claim 1, wherein the ambient humidity in the room in step (3) is controlled to be below 40%.
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