CN111838140A - Antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection - Google Patents
Antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection Download PDFInfo
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Abstract
The invention relates to the technical field of cultural relic detection, and discloses an antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection, which comprises the following steps of 1) preparing chitosan-acetic acid solution; 2) adjusting pH, performing ultrasonic treatment, adding Tween-80, heating, and stirring; 3) adding tea tree essential oil solution and TPP solution to obtain suspension; 4) ultracentrifugation at low temperature, suspending the lower layer precipitate with normal saline, and freeze drying; 5) mixing the microcapsule with quicklime; 6) placing the product obtained in the step 5) in a non-woven fabric bag; 7) cleaning mulberry leaves and mulberry twigs; 8) packaging folium Mori and ramulus Mori into storage bag, adding non-woven bag, and vacuum sealing. The invention solves the storage problem before the treatment of mulberry leaves and mulberry twigs in the producing area through the vacuum pumping and the moisture absorption and antibiosis functions, can eliminate the influence of microorganisms on samples, reduce the natural fractionation of carbon and nitrogen elements, reduce the loss of isotope information in the producing area and further ensure the accuracy of carbon and nitrogen isotope detection.
Description
Technical Field
The invention relates to the technical field of cultural relic detection, in particular to an antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection.
Background
China is the earliest country planting mulberry, silkworm breeding, silk reeling and silk weaving in the world, but the vast literature data can only witness the unique leadership of silk production and transmission in ancient manufacturing industry and luxury trade; while mythological statements about the origin and spread of silk are not convincing. Under the circumstances, a stable, effective and scientific method for tracing silk needs to be established, and isotope tracing technology is proved to be a very effective tracing technology at present.
The isotope tracing technology is mainly related to the production place, can distinguish biological products of different types and sources, and is a method for judging the regional sources more directly and effectively. Isotopes refer to a series of atoms in the nucleus of which the number of protons is the same, but the number of neutrons is different. The carbon-nitrogen stable isotope, which is the most commonly used light stable isotope, can be determined by isotopic mass spectrometry. The sample to be tested by the isotope mass spectrometry needs to be subjected to a series of pre-treatments so as to prepare a sample meeting the test conditions. The storage quality of the sample can be improved, and the content and proportion test result of the light stable isotope can be seriously influenced. After a fresh mulberry leaf and mulberry twig sample is collected in the field, considering that various microorganisms (aerobic bacteria, anaerobic bacteria and the like) in the nature can use the mulberry leaf and mulberry twig containing carbon and nitrogen elements as a carbon source and a nitrogen source required by growth and reproduction of the mulberry leaf and mulberry twig, under the general condition, the mulberry leaf and mulberry twig sample is difficult to test immediately after being collected, and even can be treated in a centralized way after being placed for many days, and the fresh mulberry leaf and mulberry twig can be easily corrupted and deteriorated due to microbial attack and the like, so that the accuracy of isotope data is influenced. Therefore, after the samples are collected, the samples need to be treated promptly to prevent microorganisms (aerobic bacteria, anaerobic bacteria and the like) from corroding the samples, influence the values of carbon and nitrogen isotopes of the samples to cause errors of isotope source tracing data, reduce natural fractionation of the carbon and nitrogen isotopes, and keep production place information as much as possible. Therefore, prior to detection of carbon and nitrogen stable isotopes, it is necessary to address the problem of sample storage prior to processing.
Disclosure of Invention
In order to solve the technical problems, the invention provides an antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection, which solves the storage problem before treatment of the mulberry leaves and the mulberry twigs in a production place through vacuumizing and the moisture absorption and antibacterial effects of tea tree essential oil chitosan microcapsules wrapped by quick lime, can eliminate the influence of microorganisms on a sample, reduce natural fractionation of carbon and nitrogen elements, reduce loss of isotope information in the production place and further ensure the accuracy of carbon and nitrogen isotope detection.
The specific technical scheme of the invention is as follows:
an antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection comprises the following steps in g and mL:
1) weighing 0.1-0.2g chitosan powder, adding 98-196mL deionized water and 2-4mL acetic acid solution with concentration of 0.5-1.5 wt%, stirring to obtain clear solution, and filtering with microporous membrane to obtain chitosan-acetic acid solution.
2) Adjusting the pH value of the chitosan-acetic acid solution to 4-5, carrying out ultrasonic treatment at room temperature, adding 0.8-1.2 wt% of Tween-80, wherein the volume ratio of the Tween-80 to the chitosan-acetic acid solution is 1: 8-12, and heating and stirring at the constant temperature of 55-65 ℃ for 30-40min to obtain a uniform and stable solution.
3) Dropwise adding 2.5-5 wt% of tea tree essential oil solution into the solution, wherein the volume ratio of the solution obtained in the step 2) to the tea tree essential oil solution is 8-12: 1, stirring at room temperature, dropwise adding 1-2mg/mL TPP solution at the speed of 2-3 drops/s, and stirring 1 to obtain white emulsion, namely tea tree essential oil chitosan microcapsule suspension.
4) And (3) carrying out low-temperature ultracentrifugation on the obtained tea tree essential oil chitosan microcapsule suspension, suspending the lower-layer precipitate by using 0.8-1.0 wt% of normal saline, and freeze-drying and storing by using 2-4 wt% of mannitol as a freeze-drying protective agent to obtain the tea tree essential oil chitosan microcapsule.
5) And (3) physically mixing the tea tree essential oil chitosan microcapsule obtained in the step (4) with quick lime under the action of mechanical force, wherein the mass ratio of the tea tree essential oil chitosan microcapsule to the quick lime is 1: 2-2.5, and obtaining the tea tree essential oil chitosan microcapsule wrapped by the quick lime.
The tea tree essential oil chitosan microcapsule wrapped by the quick lime prepared by the invention has a small amount of water absorption function and sterilization effect (chitosan). Wrapping with non-woven fabric, and placing in storage bag. When trace moisture is generated from mulberry leaves and mulberry branches in the vacuum sealed bag, the moisture is rapidly absorbed by calcium oxide because the calcium oxide has excellent hygroscopicity. Furthermore, calcium oxide releases heat after absorbing water, so that the surface of the material is heated, volatilization of tea tree essential oil in tea tree essential oil chitosan microcapsules wrapped in the material is promoted, the overall sterilization effect of the material is improved (microorganisms are easy to breed in a humid environment, and therefore the sterilization effect needs to be enhanced), and after moisture is absorbed, the calcium oxide reaction is finished, heat is not released any more, tea tree essential oil is not released at an accelerated speed, and the long-acting sterilization function is achieved.
6) Placing the product obtained in the step 5) into a polypropylene non-woven fabric bag, and sealing by heat sealing.
7) Fresh mulberry leaves and mulberry twigs which are collected from a self-producing area and have full color, no worm damage and no disease spots are selected, and the surfaces of the mulberry leaves and the mulberry twigs are respectively cleaned.
The mulberry leaves and the mulberry branches which are fresh, full in color, free of worm damage and disease spots are selected, under the same condition, the mulberry leaves and the mulberry branches which are damaged by worms and disease spots can be stored for a longer time, and exogenous influences, such as isotope influence of the damaged mulberry leaves and the mulberry branches and isotope change of the mulberry branches caused by disease spots, can be eliminated when carbon and nitrogen isotopes of the mulberry leaves and the mulberry branches are detected
8) Filling the cleaned mulberry leaves and mulberry twigs into a storage bag, adding the polypropylene non-woven fabric bag obtained in the step 6), and then carrying out vacuum sealing treatment on the storage bag by using a vacuum sealing machine.
Compared with the method of storing and collecting samples by a self-sealing bag, the invention can isolate the samples from air after vacuum sealing by a vacuum sealing machine, thereby eliminating the influence of aerobic microorganisms and being beneficial to the storage of the samples. Under the vacuum state, the storage time of the sample is prolonged, and the mulberry leaves and mulberry branches inside are not easy to deteriorate to cause the change of isotopes, thereby influencing the detection of carbon and nitrogen stable isotopes.
Preferably, in the step 1), the stirring time is 4-6h, and the pore diameter of the micro-porous filtering membrane is 0.4-0.5 μm.
Preferably, in the step 2), the reagent for adjusting the pH is 0.8-1.2mol/L NaOH solution, and the ultrasonic time is 10-20 min.
Preferably, in the step 3), the mass fraction of the tea tree essential oil solution is 2.5-5 wt%, the tea tree essential oil solution is added and stirred for 10-15min, and the TPP solution is added and stirred for 15-25 min. The volume ratio of the solution in the step 2) to the tea tree essential oil solution is 8-12: 1
Preferably, in the step 4), the parameters of the low-temperature ultracentrifugation are set to be 3-5 ℃, 14000-; the freeze drying temperature is-20 to-10 ℃.
Preferably, in step 7), the material for cleaning treatment is dust-free paper, and the solvent for cleaning treatment is 75% ethanol.
The method is characterized in that the method comprises the steps of removing impurities by using dust-free paper, wiping along the texture of mulberry leaves, removing easily-removed impurities such as dust and the like adhered to the surfaces of the mulberry leaves, enabling the dust-free paper to be soft, not damaging a sample, not leaving impurities such as particles and threads on the mulberry leaves, and having a good water retention effect, wherein 75% of ethanol solution is medical sterile alcohol, and wiping by using the medical sterile alcohol can remove part of microorganisms and some tiny dust on the surfaces of the mulberry leaves.
Preferably, in step 8), the vacuum sealing machine comprises a base and an upper flip cover which is connected with the base in a flip manner.
The top surface of the base is provided with a raised central console, a water storage tank, a lower sealing ring and a heating strip; the central console is integrated with an integrated chip, and a vacuumizing mechanism, a temperature control panel, a temperature display screen, an electric quantity display screen, a vacuum degree detection sensor and an operation button which are connected with the integrated chip; a battery is integrated on the integrated chip; the water storage tank is detachably arranged on the base and positioned on the outer side of the central console, and lower sealing rings are arranged on the peripheral sides of the water storage tank; the air exhaust port of the vacuumizing mechanism is arranged on the inner side edge of the lower sealing ring; the heating strip is arranged on the base and positioned outside the water storage tank, and the heating strip is connected with the integrated chip.
The side of the base is provided with a charging port connected with the integrated chip and a temperature adjusting knob connected with the temperature control panel.
The inner side of the upper flip cover is provided with a hollow structure corresponding to the position of the central console, an upper sealing ring matched with the lower sealing ring is arranged corresponding to the position of the lower sealing ring, and a heat-resistant pressing strip is arranged corresponding to the position of the heating strip.
At present, a portable vacuum sealing machine specially suitable for outdoor use does not exist, the existing vacuum sealing machine needs to be connected with a power supply for electrifying use, and the heat sealing temperature of the vacuum sealing machine can not be set, so that the vacuum sealing machine can only be matched with a vacuum sealing bag made of a specific material. This is extremely inconvenient for researchers to use for perishable sample collection in the field. There is therefore a need for a vacuum sealer that is portable, suitable for outdoor use, and does not require a special sealing bag.
The working principle of the vacuum sealing machine is as follows: the upper turnover cover is turned up, the unsealed side opening of the storage bag is placed in the range of the sealing ring, the upper turnover cover is closed after the open side of the storage bag is completely arranged in the sealing ring (the raised center console can play a limiting role), and the upper sealing ring and the lower sealing ring are contacted. Adjusting a temperature adjusting knob according to the material of the storage bag, clicking an operation button, starting vacuumizing, and heating and sealing the temperature adjusting knob by a heating strip after vacuumizing is finished.
The purpose of the reservoir is that when a small amount of liquid is present in the sample, a small amount of liquid may be drawn during the vacuum, and the reservoir may be used to contain the drawn liquid. The water storage tank is in a detachable installation mode and is convenient to assemble and disassemble.
Preferably, the vacuumizing mechanism comprises a micro vacuum pump, and an exhaust pipe which are connected with the micro vacuum pump; the exhaust pipe is communicated with the outside of the base; the air exhaust pipe is communicated with the air exhaust port.
The micro vacuum pump is pumped into a commercial product, and the vacuumizing principle is as follows: the circular motion of the motor makes the diaphragm inside the pump do reciprocating motion through a mechanical device, so that air in the pump cavity with fixed volume is compressed and stretched to form vacuum (negative pressure), pressure difference is generated between the pump suction port and the external atmospheric pressure, and gas pressure (suction) is sucked into the pump cavity under the action of the pressure difference and then is discharged from the exhaust port.
Preferably, the inner side of the upper flip cover and the top surface of the base are respectively provided with a buckle and a clamping groove which are matched with each other. Two sides of the base are provided with a lock catch. The bottom surface of the base is provided with a heat dissipation port.
Preferably, anti-skid rubber pads are arranged at four corners of the bottom of the base. The upper sealing ring and the lower sealing ring are made of sponge materials. The heat-resistant pressing strip is made of rubber. The upper flip cover and the base are made of environment-friendly engineering ABS waterproof materials; the electric quantity display screen and the temperature display screen are waterproof screens.
Preferably, the temperature adjusting knob is provided with four-gear temperature adjustment.
Preferably, the movable angle of the upper flip cover is designed to be 75 °.
Compared with the prior art, the invention has the beneficial effects that:
(1) the invention stores the sample in the vacuumized storage bag, can isolate the air, enables aerobic microorganisms not to influence the mulberry leaves and the mulberry branches, and can better preserve the sample. When the isotope detection is carried out, the method is particularly effective for samples needing reduced substance fractionation, and related isotope information can be kept as much as possible.
(2) The tea tree essential oil chitosan microcapsule wrapped by the quick lime prepared by the invention has a small amount of water absorption function and sterilization effect (chitosan). Wrapping with non-woven fabric, and placing in storage bag. When trace moisture is generated from mulberry leaves and mulberry branches in the vacuum sealed bag, the moisture is rapidly absorbed by calcium oxide because the calcium oxide has excellent hygroscopicity. Furthermore, calcium oxide releases heat after absorbing water, so that the surface of the material is heated, volatilization of tea tree essential oil in tea tree essential oil chitosan microcapsules wrapped in the material is promoted, the overall sterilization effect of the material is improved (microorganisms are easy to breed in a humid environment, and therefore the sterilization effect needs to be enhanced), and after moisture is absorbed, the calcium oxide reaction is finished, heat is not released any more, tea tree essential oil is not released at an accelerated speed, and the long-acting sterilization function is achieved.
(3) When the collected mulberry leaf and mulberry twig sample is cleaned by using the dust-free paper, the sample is slightly damaged, no residual particles and thread ends exist, the test result of the sample is not influenced, part of microorganisms can be removed by wiping the sample by using 75% alcohol solution, and the 75% alcohol solution is volatile and does not remain on the surface of the sample.
(4) The vacuum sealing machine disclosed by the invention is compact in structure, light and portable, can be used for both power plug and battery power supply, is convenient for outdoor use, and can meet the requirement of vacuum packaging when outdoor collection scientific research workers collect some special scientific research samples. In addition, an electric quantity display screen is arranged, so that information interaction is facilitated. The temperature of the storage bag is adjustable, the sealing requirements of the storage bags made of different materials can be met, special sealing bags are not needed, and the outdoor collection worker can conveniently select the sealing bags made of different materials and suitable for the sizes of the samples when collecting the samples.
Drawings
FIG. 1 is a schematic view of a vacuum sealer of the present invention in a configuration (with the upper lid open);
FIG. 2 is a schematic view of the outer structure of the vacuum sealing machine of the present invention (with the upper cover closed);
FIG. 3 is a schematic view of the internal structure of a base of the vacuum sealing machine of the present invention;
FIG. 4 is a side view of the vacuum sealer of the present invention;
fig. 5 is a bottom view of the vacuum sealer of the present invention.
The reference signs are: the device comprises a base 1, an upper flip cover 2, a center console 3, a water storage tank 4, a lower sealing ring 5, a heating strip 6, an integrated chip 7, a temperature control plate 8, a temperature display screen 9, an electric quantity display screen 10, a vacuum degree detection sensor 11, an operating button 12, an air pumping port 13, a charging port 14, a temperature adjusting knob 15, an upper sealing ring 16, a heat-resisting pressing strip 17, a micro vacuum pump 18, an air pumping pipe 19, an exhaust pipe 20, a buckle 21, a clamping groove 22, a lock catch 23, an anti-skidding rubber pad 24 and a heat dissipation port 25.
Detailed Description
The present invention will be further described with reference to the following examples.
Example 1
1) Weighing 0.1g of chitosan powder, adding 98mL of deionized water and 2mL of acetic acid solution with the mass fraction of 1%, placing on a magnetic stirrer, stirring for 6h to obtain a clear solution, and filtering by using a 0.4-micrometer microporous filter membrane to obtain the chitosan-acetic acid solution.
2) Adjusting the pH value of the chitosan solution to 4.5 by using 1mol/L NaOH solution, carrying out ultrasonic treatment for 10min at room temperature, adding 20ml of Tween-80 with the mass fraction of 1% into 200ml of chitosan-acetic acid solution, and heating and stirring at the constant temperature of 60 ℃ for 30min to obtain uniform and stable solution.
3) Adding 22mL of 2.5 wt% tea tree essential oil solution dropwise into the solution, stirring at room temperature for 10min on a magnetic stirrer, adding 120mL of 1.5mg/mL TPP solution dropwise at the speed of 2 drops/s, and stirring for 20min to obtain white emulsion, namely tea tree essential oil chitosan microcapsule suspension.
4) After the obtained tea tree essential oil chitosan microcapsule suspension is subjected to ultracentrifugation at low temperature (4 ℃, 15000r/min, 30min), suspending the lower-layer precipitate by using normal saline with the mass fraction of 0.9%, and performing freeze drying at the temperature of-20 ℃ by using 3 wt% of mannitol as a freeze-drying protective agent for storage to obtain the tea tree essential oil chitosan microcapsule.
5) And 4) physically mixing the tea tree essential oil chitosan microcapsule obtained in the step 4) with quick lime, wherein the mass ratio of the tea tree essential oil chitosan microcapsule to the quick lime is 1: 2, and taking 10g of the quick lime and 5g of the tea tree essential oil chitosan microcapsule to obtain the quick lime-coated tea tree essential oil chitosan microcapsule.
6) Taking a non-woven fabric made of polypropylene with the side length of 5cm, folding the non-woven fabric into a bag shape capable of containing objects, placing the material in the step 5), and performing heat sealing by using the heating function of a vacuumizing machine to enable the material to become a material which can release heat after absorbing water and trigger an antibacterial effect.
7) Selecting fresh mulberry leaves and mulberry twigs which are collected from a self-production place and have full color, no worm damage and no disease spots, and respectively cleaning the surfaces of the mulberry leaves and the mulberry twigs.
8) Selecting 20 fresh mulberry leaves which are full in color, free of worm damage and disease spots, folding mulberry twigs into a plurality of sections with proper size, selecting bags with proper size, adding the materials prepared in the step 5), and then carrying out vacuum sealing treatment by using a vacuum sealing machine.
Example 2
1) Weighing 0.2g of chitosan powder, adding 98mL of deionized water and 2mL of acetic acid solution with the mass fraction of 1%, placing on a magnetic stirrer, stirring for 6h to obtain a clear solution, and filtering by using a 0.5-micrometer microporous filter membrane to obtain the chitosan-acetic acid solution.
2) Adjusting the pH value of the chitosan solution to 4.5 by using 1mol/L NaOH solution, carrying out ultrasonic treatment for 20min at room temperature, adding 20ml of Tween-80 with the mass fraction of 1% into 200ml of chitosan-acetic acid solution, and heating and stirring at the constant temperature of 60 ℃ for 40min to obtain uniform and stable solution.
3) 22mL of a 3 wt% tea tree essential oil solution was added dropwise to the above solution, stirred on a magnetic stirrer at room temperature for 10min, 150mL of the required 1.5mg/mL TPP solution was added dropwise at a rate of 2 drops/s, and stirred for 25min to obtain a white emulsion, i.e. a suspension of tea tree essential oil chitosan microcapsules.
4) After the obtained tea tree essential oil chitosan microcapsule suspension is subjected to ultracentrifugation at low temperature (4 ℃, 15000r/min, 30min), suspending the lower-layer precipitate by using normal saline with the mass fraction of 0.9%, and performing freeze drying at the temperature of-20 ℃ by using 3% mannitol as a freeze-drying protective agent for storage to obtain the tea tree essential oil chitosan microcapsule.
5) And 4) physically mixing the tea tree essential oil chitosan microcapsule obtained in the step 4) with quick lime in a mass ratio of 1: 2.5, taking 4g of tea tree essential oil chitosan microcapsule and 10g of quick lime to obtain the quick lime-coated tea tree essential oil chitosan microcapsule.
6) Taking a non-woven fabric made of polypropylene with the side length of 4cm, folding the non-woven fabric into a bag shape capable of containing objects, placing the material in the step 5), and performing heat sealing by using the heating function of a vacuumizing machine to enable the material to become a material which can release heat after absorbing water and trigger an antibacterial effect.
7) Selecting fresh mulberry leaves and mulberry twigs which are full in color, free of worm damage and disease spots and collected from a self-production place, and respectively cleaning the surfaces of the mulberry leaves and the mulberry twigs;
8) selecting 15 fresh mulberry leaves which are full in color, free of worm damage and disease spots, folding mulberry twigs into a plurality of sections with proper size, selecting bags with proper size, adding the materials prepared in the step 5), and then carrying out vacuum sealing treatment by using a vacuum sealing machine.
Comparative example 1
1) Fresh mulberry leaves which are full in color, free of worm damage and disease spots and collected from a self-producing area are selected, and the surfaces of the mulberry leaves are respectively cleaned.
2) Drying picked mulberry leaves immediately, grinding into powder, storing in a sample bottle after treatment, and sealing the joint of a bottle cap and a bottle mouth with a sealing film to isolate air.
Comparative example 2
1) Fresh mulberry leaves which are full in color, free of worm damage and disease spots and collected from a self-producing area are selected, and the surfaces of the mulberry leaves are respectively cleaned.
2) Selecting 15 fresh mulberry leaves which are full in color, free of worm damage and disease spots, selecting bags with proper sizes, and carrying out vacuum sealing treatment by using a vacuum sealing machine, wherein the vacuum degree reaches-0.15 MPa.
Comparative example 3
1) Weighing 0.1g of chitosan powder, adding 98mL of deionized water and 2mL of acetic acid solution with the mass fraction of 1%, placing on a magnetic stirrer, stirring for 6h to obtain a clear solution, and filtering by using a 0.4-micrometer microporous filter membrane to obtain the chitosan-acetic acid solution.
2) Adjusting the pH value of the chitosan solution to 4.5 by using 1mol/L NaOH solution, carrying out ultrasonic treatment for 10min at room temperature, adding 20ml of Tween-80 with the mass fraction of 1% into 200ml of chitosan-acetic acid solution, and heating and stirring at the constant temperature of 60 ℃ for 30min to obtain uniform and stable solution.
3) Adding 22mL of 5 wt% tea tree essential oil solution dropwise into the above solution, stirring at room temperature for 15min on a magnetic stirrer, adding 120mL of 2mg/mL TPP solution dropwise at the speed of 2 drops/s, and stirring for 20min to obtain white emulsion, i.e. tea tree essential oil chitosan microcapsule suspension.
4) After the obtained tea tree essential oil chitosan microcapsule suspension is subjected to ultracentrifugation at low temperature (4 ℃, 15000r/min, 30min), suspending the lower-layer precipitate by using normal saline with the mass fraction of 0.9%, and performing freeze drying at the temperature of-20 ℃ by using 3% mannitol as a freeze-drying protective agent for storage to obtain the tea tree essential oil chitosan microcapsule.
5) And 4) physically mixing the tea tree essential oil chitosan microcapsule obtained in the step 4) with quick lime, wherein the mass ratio of the tea tree essential oil chitosan microcapsule to the quick lime is 1: 2, and taking 10g of the quick lime and 5g of the tea tree essential oil chitosan microcapsule to obtain the quick lime-coated tea tree essential oil chitosan microcapsule.
6) Taking a non-woven fabric made of polypropylene with the side length of 5cm, folding the non-woven fabric into a bag shape capable of containing objects, placing the material in the step 5), and performing heat sealing by using the heating function of a vacuumizing machine to enable the material to become a material which can release heat after absorbing water and trigger an antibacterial effect.
7) Selecting fresh mulberry leaves and mulberry twigs which are full in color, free of worm damage and disease spots and collected from a self-production place, and respectively cleaning the surfaces of the mulberry leaves and the mulberry twigs;
8) selecting 20 fresh mulberry leaves which are full in color, free of worm damage and disease spots, folding the mulberry twigs into a plurality of sections with proper size, selecting bags with proper size, placing the materials in the step 5), and then storing the materials by using common self-sealing bags.
Comparative example 4
1) Selecting fresh mulberry leaves which are full in color, free of worm damage and disease spots and collected from a self-production place, and respectively cleaning the surfaces of the mulberry leaves;
2) selecting 20 fresh mulberry leaves which are full in color, free of worm damage and disease spots, selecting bags with proper sizes, and storing the bags by using common self-sealing bags.
The above examples or comparative examples employ a dual-purpose temperature-adjustable vacuum sealer for the storage of test specimens for text, as shown in FIG. 1: comprises a base 1 and an upper flip cover 2 which is connected with the base in a flip way.
As shown in fig. 1, a raised center console 3, a water storage tank 4, a lower sealing ring 5 and a heating strip 6 are arranged on the top surface of the base. As shown in fig. 1 and 3, the console is integrated with an integrated chip 7, and a vacuum pumping mechanism, a temperature control board 8, a temperature display screen 9, an electric quantity display screen 10, a vacuum degree detection sensor 11 and an operation button 12 which are connected with the integrated chip. A battery is integrated on the integrated chip; the water storage tank is detachably arranged on the base and positioned on the outer side of the central console, and lower sealing rings are arranged on the peripheral sides of the water storage tank; the air exhaust port 13 of the vacuumizing mechanism is arranged on the inner side edge of the lower sealing ring; the heating strip is arranged on the base and positioned outside the water storage tank, and the heating strip is connected with the integrated chip. The vacuumizing mechanism comprises a micro vacuum pump 18, an air exhaust pipe 19 and an exhaust pipe 20, wherein the air exhaust pipe 19 and the exhaust pipe 20 are connected with the micro vacuum pump; the exhaust pipe is communicated with the outside of the base; the air exhaust pipe is communicated with the air exhaust port.
As shown in fig. 4-5, a charging port 14 connected with the ic and a temperature adjusting knob 15 connected with the temperature control board are provided at the side of the base (four-step temperature adjustment). The two sides of the base are also provided with a lock 23. The bottom four corners of base are equipped with anti-skidding rubber pad 24, and the bottom surface of base still is equipped with thermovent 25.
As shown in fig. 1-3, the inner side of the upper flip cover corresponding to the center console is a hollow structure, an upper sealing ring 16 matching with the lower sealing ring is disposed corresponding to the lower sealing ring, and a heat-resistant pressing strip 17 is disposed corresponding to the heating strip. The inner side of the upper flip cover and the top surface of the base are respectively provided with a buckle 21 and a clamping groove 22 which are matched with each other.
Wherein, go up sealing washer and lower sealing washer are the sponge material. The heat-resistant pressing strip is made of rubber.
The working principle of the embodiment is as follows: the upper turnover cover is turned up, the unsealed side opening of the storage bag is placed in the range of the sealing ring, the upper turnover cover is closed after the open side of the storage bag is completely arranged in the sealing ring (the raised center console can play a limiting role), and the upper sealing ring and the lower sealing ring are contacted. Adjusting a temperature adjusting knob according to the material of the storage bag, clicking an operation button, starting vacuumizing, and heating and sealing the temperature adjusting knob by a heating strip after vacuumizing is finished.
It should be noted that the integrated chip, the vacuum pumping mechanism, the temperature control board, the temperature display screen, the control knob, the power display screen, the vacuum degree detection sensor, the operation button and other components and the circuit connections therebetween in this embodiment are all common components and circuit connections in the prior art.
The purpose of the reservoir is that when a small amount of liquid is present in the sample, a small amount of liquid may be drawn during the vacuum, and the reservoir may be used to contain the drawn liquid. The water storage tank is in a detachable installation mode and is convenient to assemble and disassemble.
The related technical parameters of the vacuum sealing machine of the embodiment are described as follows: the rated voltage is 220V, the rated frequency is 50Hz, the rated power is 220W, the vacuum strength is-50 KPa, the heating wire is 4mm, the temperature application range can be from common plastic bags to kraft bags, the packaging size is 300mm x 120mm x 80mm, the sealing length is up to 20 cm, the voltage of the battery is 12V, the capacity is 5000mAh, the size is 95mm x 60mm x 18mm, the stable working current is 8A, the maximum working current is 10A, and the overall weight is 1.32 kg.
The mulberry leaves in all the above examples and comparative examples were collected from the same mulberry (aged almost, 8 th leaf from top of mulberry branch). The sample of comparative example 1 was sent for testing on a portion of the day after encapsulation; the stored mulberry leaf samples of example 1, example 2, comparative example 3 and comparative example 4 were all subjected to the same operations of drying and grinding into powder as in comparative example 1 after being placed in a room for 7 days under natural conditions (5 months in the south of the Yangtze river), and then sent to a unified test for C, N stable isotopes together with another part of the sample of comparative example 1 placed for the same days under the same conditions, each sample was tested 5 times, and finally averaged. The data measured for the sample with the 7-day standing of comparative example 1 is represented by comparative example 5.
The following are the data for carbon and nitrogen isotopes for the examples and comparative examples (with the data measured for the 7-day-old sample of comparative example 1 being shown as comparative example 5):
| species of | δ13C | δ15N |
| Example 1 | -29.727±0.295 | 4.220±1.219 |
| Example 2 | -29.731±0.314 | 4.218±1.197 |
| Comparative example 1 | -29.734±0.371 | 4.213±1.211 |
| Comparative example 2 | -29.115±0.697 | 3.186±3.201 |
| Comparative example 3 | -28.672±0.812 | 2.735±4.113 |
| Comparative example 4 | -27.706±0.923 | 2.528±3.897 |
| Comparative example 5 | -29.732±0.382 | 4.217±1.205 |
Comparative example 1 is the data tested on the same day, the obtained isotope data is standard numerical values, the local geographic information can be reflected best, and the complete isotope information is reserved, so that the isotope data can be used as a comparison between other several examples and comparative examples. From the above table, we can see that of examples 1, 2 and comparative example 513C value and15the error of the N value is within 0.007% compared with that of the comparative example 1, and the examples 1 and 2 adopt vacuum storage and use of antibacterial materials, and the difference is that the concentrations of the antibacterial materials are different, which shows that the effect of the invention is remarkable, and the maximum degree of the effect can be realizedThe isotope information of the sample producing area is reserved. Comparative example 2 was a sample that was stored under vacuum but not with an antimicrobial material, while comparative example 3 was a sample that was stored with a pouch and was visible13The C value tends to be enriched in the comparative examples 2 and 3, the error between the comparative example 2 and the standard value reaches about 0.6 per thousand, and the error between the comparative example 3 and the standard value reaches about 1 per thousand. While 15The N value is continuously depleted in the comparative examples 2 and 3, and the errors of the comparative examples 2 and 3 respectively reach 1.1 per thousand and 1.5 per thousand. The marked fractionation effect of carbon and nitrogen isotopes of the sample occurs in the preservation process, and the isotope information of the producing area carried by the sample is lost. In contrast, comparative example 4 neither employed vacuum storage nor used an antibacterial material. Comparative example 4 sample of13C value and15compared with the standard value, the error of the N value reaches 2 per mill and 1.7 per mill respectively, which belongs to huge errors in isotope analysis, and indicates that the loss of isotope information in a production place is serious, thereby having huge influence on scientific research and analysis.
The raw materials and equipment used in the invention are common raw materials and equipment in the field if not specified; the methods used in the present invention are conventional in the art unless otherwise specified.
The above description is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and all simple modifications, alterations and equivalents of the above embodiments according to the technical spirit of the present invention are still within the protection scope of the technical solution of the present invention.
Claims (10)
1. An antibacterial vacuum storage method of mulberry leaves and mulberry twigs for carbon and nitrogen isotope detection is characterized by comprising the following steps of:
1) Weighing 0.1-0.2 g of chitosan powder, adding 98-196 mL of deionized water and 2-4 mL of 0.5-1.5wt% acetic acid solution, stirring to obtain a clear solution, and filtering with a microporous filter membrane to obtain a chitosan-acetic acid solution;
2) adjusting the pH value of the chitosan-acetic acid solution to 4-5, carrying out ultrasonic treatment at room temperature, adding 0.8-1.2wt% of Tween-80, wherein the volume ratio of the Tween-80 to the chitosan-acetic acid solution is 1:8-12, and heating and stirring at the constant temperature of 55-65 ℃ for 30-40 min to obtain a uniform and stable solution;
3) dropwise adding 2.5-5wt% of tea tree essential oil solution into the solution, wherein the volume ratio of the solution obtained in the step 2) to the tea tree essential oil solution is 8-12:1, stirring at room temperature, dropwise adding 1-2 mg/mL TPP solution at the speed of 2-3 drops/s, and stirring to obtain white emulsion, namely tea tree essential oil chitosan microcapsule suspension;
4) after the obtained tea tree essential oil chitosan microcapsule suspension is subjected to ultracentrifugation at low temperature, suspending the lower-layer precipitate by using 0.8-1.0wt% of normal saline, and freeze-drying and storing by using 2-4wt% of mannitol as a freeze-drying protective agent to obtain tea tree essential oil chitosan microcapsules;
5) physically mixing the tea tree essential oil chitosan microcapsule obtained in the step 4) with quicklime under the action of mechanical force, wherein the mass ratio of the tea tree essential oil chitosan microcapsule to the quicklime =1 (2-2.5) to obtain the quicklime-coated tea tree essential oil chitosan microcapsule;
6) Placing the product obtained in the step 5) in a polypropylene non-woven fabric bag, and sealing by heat sealing;
7) selecting fresh mulberry leaves and mulberry twigs which are full in color, free of worm damage and disease spots and collected from a self-production place, and respectively cleaning the surfaces of the mulberry leaves and the mulberry twigs;
8) filling the cleaned mulberry leaves and mulberry twigs into a storage bag, adding the polypropylene non-woven fabric bag obtained in the step 6), and then carrying out vacuum sealing treatment on the storage bag by using a vacuum sealing machine.
2. The method of claim 1, wherein in the step 1), the stirring time is 4 to 6 hours, and the pore size of the micro-filtration membrane is 0.4 to 0.5 μm.
3. The method of claim 1, wherein in step 2), the reagent for adjusting the pH is 0.8-1.2 mol/L NaOH solution, and the ultrasonic time is 10-20 min.
4. The method of claim 1, wherein in step 3), the tea tree essential oil solution is added and stirred for 10-15min, and the TPP solution is added and stirred for 15-25 min.
5. The method as claimed in claim 1, wherein in step 4), the parameters of the low-temperature ultracentrifugation are set to 3-5 ℃, 14000-; the temperature of freeze drying is-20 to-10 ℃.
6. The method as claimed in claim 1, wherein in step 7), the material for the cleaning treatment is a dust-free paper, and the solvent for the cleaning treatment is 75% ethanol.
7. The method of claim 1, wherein the vacuum after evacuation of the storage bag in step 8) is from-0.15 to 0 MPa.
8. The method of claim 1, wherein in step 8), the vacuum sealing machine comprises a base (1) and an upper flip cover (2) which is connected with the base in a flip manner;
the top surface of the base is provided with a raised central console (3), a water storage tank (4), a lower sealing ring (5) and a heating strip (6); the central console is integrated with an integrated chip (7), a vacuumizing mechanism connected with the integrated chip, a temperature control panel (8), a temperature display screen (9), an electric quantity display screen (10), a vacuum degree detection sensor (11) and an operation button (12); a battery is integrated on the integrated chip; the water storage tank is detachably arranged on the base and positioned on the outer side of the central console, and lower sealing rings are arranged on the peripheral sides of the water storage tank; an air exhaust port (13) of the vacuumizing mechanism is arranged on the inner side edge of the lower sealing ring; the heating strip is arranged on the base and positioned outside the water storage tank, and the heating strip is connected with the integrated chip; the vacuum pumping mechanism comprises a micro vacuum pump (18), an air pumping pipe (19) and an exhaust pipe (20), wherein the air pumping pipe and the exhaust pipe are connected with the micro vacuum pump; the exhaust pipe is communicated with the outside of the base; the air exhaust pipe is communicated with the air exhaust port;
A charging port (14) connected with the integrated chip and a temperature adjusting knob (15) connected with the temperature control panel are arranged on the side of the base;
the inner side of the upper flip cover corresponding to the center console is of a hollow structure, an upper sealing ring (16) matched with the lower sealing ring is arranged at the position corresponding to the lower sealing ring, and a heat-resisting pressing strip (17) is arranged at the position corresponding to the heating strip.
9. The method of claim 8, wherein the inner side of the upper lid and the top surface of the base are provided with a snap (21) and a slot (22) respectively; two sides of the base are provided with lock catches (23); anti-skid rubber pads (24) are arranged at four corners of the bottom of the base; the bottom surface of the base is provided with a heat dissipation port (25).
10. The method of claim 8, wherein the temperature adjustment knob is provided with a four-step temperature adjustment.
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