CN117801965B - Method for preparing biological sponge for adsorbing sewage by using cordyceps militaris and application of biological sponge - Google Patents
Method for preparing biological sponge for adsorbing sewage by using cordyceps militaris and application of biological sponge Download PDFInfo
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Abstract
The invention belongs to the technical field of biological materials, and discloses a method for preparing biological sponge for adsorbing sewage by using cordyceps militaris and application thereof, wherein the method comprises the following steps: inoculating Cordyceps militaris spore suspension into potato liquid culture medium, culturing at 25deg.C and 120rpm in dark for 72 h, and winding mycelia into uniform size after Cordyceps militaris spores germinate in shake flask; and (3) putting the obtained fungus balls into a glassware, freezing, and transferring into freeze drying to obtain the cordyceps militaris sponge. The invention uses the cordyceps militaris mycelium as the raw material, the developed sewage treatment material is environment-friendly and low in cost, the raw material is renewable, and the developed cordyceps militaris sponge has high-efficiency adsorption capacity. Cordyceps militaris sponge contains a large amount of chitosan with main structure in fungus cell walls. The Cordyceps militaris sponge prepared from the Cordyceps militaris ball has good adsorption capacity for various heavy metal ions and organic dyes in sewage.
Description
Technical Field
The invention belongs to the technical field of biology and materials, and in particular relates to a method for preparing biological sponge for adsorbing sewage by using cordyceps militaris and application thereof.
Background
There is an increasing research in developing methods and materials for sewage treatment, and there is an increasing market demand for materials for sewage treatment. To date, various sewage treatment materials have achieved tremendous efforts in sewage treatment, such as activated carbon for sewage adsorption, and flocculation precipitation of sewage with chemical agents. However, these methods have the disadvantages of high cost, low efficiency, incompatibility with natural environments, and the like. Therefore, development of sewage treatment materials using biological materials has also been greatly progressed in recent years. The microbial material in the biomass material occupies a place, is rich in cellulose, starch, chitin and the like, and can grow and reproduce rapidly, so that the biomass material is a good renewable material, wherein fungi are easy to cultivate, grow rapidly, have low resource requirements, and are a sustainable source of multifunctional and easily-degradable biological composite materials. In recent years, researchers have begun to develop materials and methods for fungus-based sewage treatment, with the aim of making use of environmental resources for environmental protection and development.
Fungal hyphae are natural, renewable, valuable structural polymeric materials consisting primarily of dextran, proteins, chitin and other natural polymers. Unlike bacteria and yeasts, fungi as filamentous species can produce single-tubular hyphae in a suitable medium, forming a large-area fibrous hyphae network. The material can be used for packaging materials, building materials, environment-friendly repair materials, wear-resistant materials and the like. Wherein the main component of the fungal hypha cell wall is chitin. Chitin has rich hydroxyl and other active groups. The active groups have strong adsorption capacity to metal cations and organic dyes in sewage.
The advantages and disadvantages of the conventional sewage treatment mode are shown in table 1, all the conventional sewage treatment modes have high treatment cost, difficult regeneration conditions and reduced adsorption performance after regeneration, so that the development of an adsorption material which is environment-friendly, simple in regeneration and unaffected in adsorption performance after regeneration is very important.
TABLE 1 present treatment method for removing various heavy metal ions
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a method for preparing biological sponge for adsorbing sewage by using cordyceps militaris and application thereof.
The technical scheme adopted for solving the technical problems is as follows:
A method for preparing biological sponge for adsorbing sewage by using cordyceps militaris, which comprises the following steps:
(1) Inoculating Cordyceps militaris spore suspension into potato liquid culture medium, culturing at 25deg.C and 120rpm in dark for 72 h, germinating Cordyceps militaris spores in shake flask, and winding mycelium into uniform-sized fungus balls; removing the fungus balls from the shake flask, and cleaning the residual culture medium and broken hyphae by using sterile water;
(2) And (3) putting the fungus balls obtained in the step (1) into a glassware, freezing for 2-4 hours at the temperature of minus 80 ℃, and transferring into freeze drying to obtain the Cordyceps militaris sponge.
Further, the Cordyceps militaris is Cordyceps militaris ACCC50632, purchased from Xinyang city, and stored in Nanjing university food and pharmaceutical engineering college.
Further, the mycelia of the cordyceps militaris are white, thick, uniform and fast in growth rate, pigments are easy to secrete after the cordyceps militaris is exposed to light, and the aerial mycelia on the surface of the culture medium are thicker.
Further, the cordyceps militaris sponge has the capabilities of dye adsorption and heavy metal ion adsorption.
Further, the cordyceps militaris sponge has an organic dye adsorption capacity.
Further, the cordyceps militaris sponge is subjected to Fourier infrared spectroscopy, an X-ray photoelectron spectroscopy technology and X-ray diffraction to determine the performance of adsorbing heavy metal ions.
The biological sponge prepared by the method is applied to sewage adsorption.
The biological sponge prepared by the method is applied to the adsorption of heavy metal ions Cd and/or Pb in sewage.
The beneficial effects obtained by the invention are as follows:
1. The invention uses the cordyceps militaris mycelium as the raw material, the developed sewage treatment material is environment-friendly and low in cost, the raw material is renewable, and the developed cordyceps militaris sponge has high-efficiency adsorption capacity. Hypha grows out and is wound into balls in the growth and culture process of the cordyceps militaris spores, the cordyceps militaris hypha presents a thick and branched form, and the hypha has the characteristic of high specific surface area, so that the cordyceps militaris sponge contains a large amount of chitosan with a main structure in fungus cell walls. The Cordyceps militaris sponge prepared from the Cordyceps militaris ball has good adsorption capacity for various heavy metal ions and organic dyes in sewage.
2. The invention aims at the development of renewable sewage treatment materials, and prepares a sewage treatment material by using cordyceps militaris, and the cordyceps militaris mycelium pellets in a liquid culture medium are collected, washed and filtered, and then placed into a glassware for vacuum freeze drying. The prepared cordyceps militaris sponge realizes the development and utilization of renewable materials, and has the effects of efficiently purifying sewage and adsorbing heavy metal ions in the sewage.
3. The fungus selected by the invention is Cordyceps militaris. In the submerged culture of Cordyceps militaris, mycelia are wound into balls during the culture process. The spherical hyphae are convenient to collect. Therefore, the cordyceps militaris mycelia are collected and subjected to vacuum freeze drying, and other expensive and complex treatment methods are not needed, so that the cordyceps militaris sponge with super-strong water absorption capacity is prepared, and the cordyceps militaris sponge has adsorption capacity on metal ions and organic dyes due to the characteristics of fungi, so that the cordyceps militaris sponge is hopeful to become a novel sewage adsorption material, and environmental resource protection is promoted.
4. The biological sponge is prepared from cordyceps militaris mycelia in fungi, and the cordyceps militaris sponge is an excellent sewage adsorption material and has the following advantages:
1) The production cost is low, the cordyceps militaris can grow rapidly and propagate simply as a microorganism, the place required by the culture is small, and the space three-dimensional culture can be adopted. 2) The Cordyceps militaris mycelium can be cultured again to the original volume after being placed in the culture medium for 7 days even if only 1/10 of the mycelium remains. 3) The cordyceps militaris sponge is environment-friendly, is prepared from natural cordyceps militaris mycelia, and does not introduce other substances in the preparation process, so that the cordyceps militaris sponge can be degraded and utilized in a natural environment.
Drawings
FIG. 1 is a digital image of a Cordyceps militaris sponge material in example 1 of the invention;
FIG. 2 is a graph showing adsorption kinetics of Cordyceps militaris sponge in different ion solutions in example 2 of the invention;
FIG. 3 is an electron microscopic image of the Cordyceps militaris sponge of example 3 of the invention after adsorbing the heavy metal solution and freeze-drying;
Fig. 4 is an infrared spectrum image (EDS image) of the cordyceps militaris sponge in example 4 before and after adsorption of heavy metal ions in the present invention;
FIG. 5 is a graph showing the comparison of adsorption effects of Cordyceps militaris sponge and commercial sponge in deionized water in comparative example 1 of the present invention;
FIG. 6 is a graph showing the adsorption effect of Cordyceps militaris sponge and commercial sponge in rose bengal aqueous solution in comparative example 2;
FIG. 7 is a graph showing the adsorption effect of Cordyceps militaris sponge and commercial sponge in methylene blue ethanol solution in comparative example 3.
Detailed Description
The present invention will be further described in detail with reference to examples, but the scope of the present invention is not limited to the examples.
The raw materials used in the invention are conventional commercial products unless specified otherwise, the methods used in the invention are conventional methods in the art unless specified otherwise, and the mass of each substance used in the invention is conventional.
A method for preparing biological sponge for adsorbing sewage by using cordyceps militaris, which comprises the following steps:
(1) Inoculating Cordyceps militaris spore suspension into potato liquid culture medium, culturing at 25deg.C and 120rpm in dark for 72 h, germinating Cordyceps militaris spores in shake flask, and winding mycelium into uniform-sized fungus balls; removing the fungus balls from the shake flask, and cleaning the residual culture medium and broken hyphae by using sterile water;
(2) And (3) putting the fungus balls obtained in the step (1) into a glassware, freezing for 2-4 hours at the temperature of minus 80 ℃, and transferring into freeze drying to obtain the Cordyceps militaris sponge.
Preferably, the Cordyceps militaris is Cordyceps militaris ACCC50632, purchased from Xinyang city, and deposited at Nanjing university food and pharmaceutical engineering college.
Preferably, the mycelia of the cordyceps militaris are white, thick, uniform and high in growth rate, pigments are easy to secrete after the cordyceps militaris is exposed to light, and the aerial mycelia on the surface of the culture medium are thicker.
Preferably, the cordyceps militaris sponge has the capabilities of dye adsorption and heavy metal ion adsorption.
Preferably, the cordyceps militaris sponge has an organic dye adsorption capacity.
Preferably, the cordyceps militaris sponge is subjected to Fourier infrared spectroscopy, an X-ray photoelectron spectroscopy technology and X-ray diffraction to determine the performance of adsorbing heavy metal ions.
The biological sponge prepared by the method is applied to sewage adsorption.
The biological sponge prepared by the method is applied to the adsorption of heavy metal ions Cd and/or Pb in sewage.
Specifically, the preparation and detection of the correlation are as follows:
Preparing a potato solid medium: weighing 200g of peeled potatoes, cutting up, putting into pure water, boiling for 30 minutes, filtering potato dregs with double-layer gauze to obtain filtrate, adding 20g of glucose and 15g of agar powder, fixing the volume to 1 liter without regulating the pH, pouring into a blue-cover reagent bottle after stirring uniformly, putting into a sterilizing pot at 115 ℃, and sterilizing for 20 minutes.
Preparing potato liquid culture medium: weighing 200g of peeled potatoes, cutting up, putting into pure water, boiling for 30 minutes, filtering potato dregs with double-layer gauze to obtain filtrate, adding 20g of glucose, fixing the volume to 1 liter without regulating the pH, pouring into a blue-cover reagent bottle after stirring uniformly, putting into a sterilizing pot at 115 ℃, and sterilizing for 20 minutes.
Preparation of potato dextrose water culture medium: 6g of potato extract powder and 20g of glucose, and sterilizing for 20 minutes in a sterilizing pot at 115 ℃ after the volume is fixed to 1 liter.
Example 1
A method for preparing biological sponge for adsorbing sewage by using cordyceps militaris, which comprises the following steps:
the Cordyceps militaris strain ACCC50632 is activated on a potato solid culture medium, placed in a 28 ℃ incubator for culturing for 7 days, and the culture medium is washed by pure water to obtain Cordyceps militaris spore suspension. Preparing potato liquid culture medium, counting with a blood cell counting plate, inoculating Cordyceps militaris spores with an inoculum size of 1x 10≡8 per 100 mL potato liquid culture medium, placing into a 250 mL conical flask, and culturing in a shaking table at 25deg.C and 120rpm for 3 days. Preliminary experiments on the culture temperature at the early stage show that the bacterial balls are better formed under the condition of about 25 ℃. The temperature is too low, and spores are not easy to grow and germinate; the temperature is too high, the spore germination is slow, and the number of the fungus balls is small. The control experiments were therefore carried out at 25℃and 120rpm in shaker conditions. Filtering bacterial liquid with double-layer gauze, cleaning for several times to obtain bacterial balls, placing the bacterial balls in a glass culture dish, freezing at-80deg.C for 5h, and lyophilizing with vacuum freeze dryer for 24: 24 h to obtain Cordyceps militaris sponge, wherein the prepared Cordyceps militaris sponge has light form and can be supported by flowers as shown in figure 1.
Example 2
The Cordyceps militaris sponge in example 1 was placed in 25 mg/mL of heavy metal ion (Cd, pb) solution. And then carrying out adsorption experiments in an oscillation box at the rotating speed of 80 rpm, separating out supernatant after adsorption is finished, filtering the supernatant through a filter head of 0.22 mu m, and finally measuring the content of Cd and Pb in the solution by using a flame atomic absorption spectrophotometer, repeating each experiment for 3 times, and taking an average value as a final result. The adsorption performance of the Cordyceps militaris sponge on different metal ions is shown in figure 2, the adsorption capacity is gradually increased along with the increase of time, and the rapid adsorption balance can be achieved at 90 min, which benefits from the three-dimensional porous structure of the Cordyceps militaris sponge and the high specific surface area of the fungal hyphae. And at 0-20 min, the adsorption capacity increases significantly with the adsorption time.
Example 3
The Cordyceps militaris sponge in example 1 was placed in 25 mg/mL of heavy metal ion (Cd, pb) solution for 20min. Taking out Cordyceps militaris sponge in heavy metal ion solution, and lyophilizing with vacuum lyophilizing machine 24 h. The cordyceps militaris sponge after being adsorbed with the heavy metal solution and freeze-dried is placed under an electron microscope, as shown in fig. 3, the heavy metal ions are observed to be uniformly adsorbed on the surface of hyphae, and the adsorption effect of the cordyceps militaris sponge on Cd ions can be seen to be better in an EDS scanning chart.
Example 4
The Cordyceps militaris sponge in example 1 was placed in 25 mg/mL of heavy metal ion (Cd, pb) solution for 20min. Taking out Cordyceps militaris sponge in heavy metal ion solution, and lyophilizing with vacuum lyophilizing machine 24 h. The cordyceps militaris sponge after being adsorbed with the heavy metal solution and freeze-dried is judged to cause the change of the chemical structure of the cordyceps militaris sponge after the heavy metal ions are adsorbed by the cordyceps militaris sponge by utilizing a Fourier infrared spectrum technology, as shown in fig. 4, which shows that the adsorption of the heavy metal ions by the cordyceps militaris sponge is not only simple physical adsorption but also chemical adsorption.
Comparative example 1
The cordyceps militaris sponge in the embodiment 1 is cut into a cubic sponge with the length of 0.5cm multiplied by 1cm multiplied by 2 cm, one end of the cordyceps militaris sponge is contacted with water, the time is recorded at the moment of contact, and the water is absorbed when the water is absorbed from the bottom to the whole top of the cordyceps militaris sponge. Correspondingly, a common commercial sponge (polyurethane, melamine) is cut into a cubic sponge of 0.5cm multiplied by 1cm multiplied by 2 cm, one end is contacted with water, the time is recorded at the moment of contact, and the commercial sponge absorbs water to the top, namely the water absorption is finished. As shown in FIG. 5, the Cordyceps militaris sponge has stronger water absorbing capacity than commercial sponge (polyurethane, melamine), and as can be seen from FIG. 5, at the beginning of the contact of three sponges with water, the melamine sponge does not absorb water at all, the Cordyceps militaris sponge and the polyurethane sponge show rapid water absorbing property, meanwhile, the water absorbing rate of the Cordyceps militaris sponge is faster, the water absorbing height of the polyurethane sponge is not increased after 10s absorption, but the Cordyceps militaris sponge can absorb water all the time until 150s, and the whole water absorption of the Cordyceps militaris sponge is completed.
Comparative example 2
The cordyceps militaris sponge in the embodiment 1 is cut into a cubic sponge with the length of 0.5cm multiplied by 1cm multiplied by 2 cm, one end of the cordyceps militaris sponge is contacted with the aqueous solution of the rose bengal (the rose bengal is dissolved in deionized water, the concentration is 10 g/L), the time is recorded at the moment of contact, and the adsorption is completed when the cordyceps militaris sponge is completely dyed by the rose bengal dye. Correspondingly, a common commercial sponge (polyurethane, melamine) is cut into a cubic sponge with the length of 0.5cm multiplied by 1cm multiplied by 2 cm, one end is contacted with an aqueous solution of the rose bengal (the rose bengal is dissolved in deionized water and has the concentration of 10 g/L), the time is recorded at the moment of contact, and the adsorption is completed when the commercial sponge is completely dyed by the rose bengal dye. As shown in fig. 6, it was shown that the cordyceps militaris sponge had better dye adsorption capacity than the commercial sponge (polyurethane, melamine). As can be seen from fig. 6, when the Cordyceps militaris sponge dye is adsorbed for 30s, the height of the Cordyceps militaris sponge dye is obviously higher than that of the other two sponges, when the Cordyceps militaris sponge is adsorbed for 150s, the dye is adsorbed to the top, and the other two sponge dyes are adsorbed Gao Duyuan far less than the Cordyceps militaris sponge, so that the Cordyceps militaris sponge is far more than the commercial sponge in the capability of autonomously adsorbing the rose water solution of the Bengale under the same time and the same volume.
Comparative example 3
The cordyceps militaris sponge in the embodiment 1 is cut into a cubic sponge with the length of 0.5cm multiplied by 1cm multiplied by 2 cm, one end of the cordyceps militaris sponge is contacted with methylene blue ethanol solution (the concentration of methylene blue is 10g/L in the ethanol solution), the time is recorded at the moment of contact, and the adsorption is completed when the cordyceps militaris sponge is completely dyed by methylene blue dye. In response, a common commercial sponge (polyurethane, melamine) is cut into a cubic sponge of 0.5cm multiplied by 1cm multiplied by 2 cm, one end of the cubic sponge is contacted with methylene blue ethanol solution (the methylene blue is dissolved in the ethanol solution, the concentration is 10 g/L), the time is recorded at the moment of contact, and the adsorption is completed when the commercial sponge is completely dyed by the methylene blue ethanol solution. As shown in fig. 7, it was shown that the cordyceps militaris sponge had better dye adsorption capacity than the commercial sponge (polyurethane, melamine). As can be seen from fig. 7, when the Cordyceps militaris sponge and the polyurethane sponge have good adsorption capacity on methyl blue ethanol solution, when the Cordyceps militaris sponge is adsorbed for 5s, the dye of the Cordyceps militaris sponge is obviously higher than that of other two sponges, when the Cordyceps militaris sponge is adsorbed for 52s, only the Cordyceps militaris sponge adsorbs the dye on the top, and the other two sponge dyes are adsorbed Gao Duyuan far less than that of the Cordyceps militaris sponge, so that the Cordyceps militaris sponge has the capability of autonomously adsorbing methylene blue ethanol solution in the same time and the same volume, and is far beyond that of commercial sponge.
Although embodiments of the present invention have been disclosed for illustrative purposes, those skilled in the art will appreciate that: various substitutions, changes and modifications are possible without departing from the spirit and scope of the invention and the appended claims, and therefore the scope of the invention is not limited to the disclosure of the embodiments.
Claims (3)
1. A method for preparing biological sponge for adsorbing sewage by using cordyceps militaris is characterized by comprising the following steps: the method comprises the following steps:
(1) Inoculating Cordyceps militaris spore suspension into potato liquid culture medium, culturing at 25deg.C and 120rpm in dark for 72 h, and winding mycelia into uniform size after Cordyceps militaris spores germinate in shake flask; taking out the fungus balls from the shake flask, and cleaning the residual culture medium and broken hyphae by using sterile water;
(2) Putting the fungus balls obtained in the step (1) into a glassware, freezing for 2-4 hours at the temperature of minus 80 ℃, and transferring into freeze drying to obtain Cordyceps militaris sponge;
The Cordyceps militaris is Cordyceps militaris ACCC50632.
2. The biological sponge according to claim 1, wherein the biological sponge is used for adsorbing sewage.
3. The biological sponge according to claim 1, wherein the biological sponge is used for adsorbing heavy metal ions Cd and/or Pb in sewage.
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