Single cell surface stereocomplex polylactic acid silicon dioxide radiation refrigeration shell material and preparation method thereof

Abstract

The invention discloses a single cell surface stereocomplex polylactic acid silicon dioxide radiation refrigeration shell layer material and a preparation method thereof, L-polylactic acid, D-polylactic acid and silicon dioxide are adopted as raw materials, a radiation refrigeration shell layer is constructed on the surface of a yeast single cell, and yeast cells are endowed with good environmental tolerance; the cell activity of yeast cells in a high-temperature environment is improved, so that the ethanol yield is improved. The stereocomplex polylactic acid silicon dioxide radiation refrigeration shell material can reduce the surface temperature of an object with zero energy consumption by reflecting visible light and near-infrared light and radiating mid-infrared light, improves the tolerance of cells to light and heat, provides protection for the cells, and further provides light, heat and high temperature resistant immobilized cells for simultaneous saccharification and fermentation. The radiation refrigeration material can reflect visible light and near infrared rays and emit intermediate infrared rays, so that cooling and refrigeration are achieved. A radiation refrigeration technology and a cell immobilization technology are combined, and cells are immobilized by a radiation refrigeration material, so that the tolerance of yeast cells to light and heat is improved.

Description

Technical Field

[0001] The present invention belongs to the technical field of radiation refrigeration material preparation, and relates to a single-cell surface stereocomposite polylactic acid silicon dioxide radiation refrigeration shell material and a preparation method thereof. Background Art

[0002] With the acceleration of population growth and industrial modernization, the contradiction between supply and demand of traditional fossil energy is becoming increasingly severe, and the energy crisis has become a major challenge hindering sustainable economic development. Vigorously developing bioethanol using cellulose as raw material is in line with the requirements of optimizing the energy structure and achieving a green and low-carbon transition. In the bioconversion process of bioethanol, the simultaneous saccharification and fermentation (SSF) process has attracted much attention because it places yeast cells and cellulase in the same fermentation system, effectively improving substrate utilization and shortening the production cycle. However, its core bottleneck lies in the significant difference in the optimal temperature of cellulase and yeast cells. Therefore, improving the tolerance limit of yeast cells in high temperature environments is the key to improving the efficiency of the SSF process. Light causes the solution temperature to rise, and high temperature destroys the cell membrane structure, affecting the normal metabolic activities and production work of the cells. Summary of the Invention

[0003] The purpose of the present invention is to solve the problems of the prior art and provide a single-cell surface stereocomposite polylactic acid silica radiation refrigeration shell material and a preparation method.

[0004] In order to achieve the above object, the present invention adopts the following technical solutions:

[0005] The present invention provides a method for preparing a single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material, comprising the following steps:

[0006] Step 1: After adding PDDA solution to the yeast cell resuspension and shaking, the mixture was centrifuged and washed with a buffer solution to prepare yeast cells with PDDA nanoshells assembled on their surfaces, which were designated as Cell@PDDA.

[0007] Step 2: Cell@PDDA was placed in a buffer solution, nano-SiO2 dispersion was added and shaken, and then centrifuged and washed with the buffer solution to obtain yeast cells with PDDA / SiO2 nanoshells assembled on their surfaces, which were recorded as Cell@PDDA@SiO2.

[0008] Step 3: Cell@PDDA@SiO2 was placed in a buffer solution, PLLA solution was added and shaken, and then centrifuged and washed with the buffer solution to obtain yeast cells with PDDA / SiO2 / PLLA nanoshells assembled on their surfaces, which were recorded as Cell@PDDA@SiO2@PLLA.

[0009] Step 4: Place Cell@PDDA@SiO2@PLLA in a buffer solution, add PDLA solution and shake, then centrifuge and wash with buffer solution to obtain yeast cells with PDDA / SiO2 / SC-PLA nanoshells assembled on the surface, which are recorded as Cell@PDDA@SiO2@SC-PLA.

[0010] Preferably, the preparation method of the PDDA solution is as follows:

[0011] Dissolve PDDA in PBS buffer solution and stir until completely dissolved to prepare PDDA solution;

[0012] The ratio of PDDA to PBS buffer solution is (0.01 g to 0.05 g): (5 mL to 20 mL), and the pH of the PBS buffer solution is 5 to 7.

[0013] Preferably, the preparation method of the nano-SiO2 dispersion is as follows:

[0014] Nano-SiO2 was dispersed in PBS buffer solution and ultrasonicated until uniformly dispersed to prepare nano-SiO2 dispersion.

[0015] The ratio of nano-SiO2 to PBS buffer solution is (0.01g-0.05g):(5mL-20mL), the particle size of nano-SiO2 is 20nm-100nm, the ultrasonic time is 20min-60min, and the pH value of the PBS buffer solution is 5-7.

[0016] Preferably, the preparation method of the PLLA solution and the PDLA solution is as follows:

[0017] HFIP solution was prepared using PBS buffer solution, PLLA and PDLA were dissolved in HFIP solution respectively, and heated and stirred until completely dissolved to prepare PLLA solution and PDLA solution;

[0018] The mass percentage concentration of the HFIP solution is 5% to 10%, and the ratio of the HFIP solution, PLLA and PDLA is (5 mL to 20 mL):(0.01 g to 0.05 g):(0.01 g to 0.05 g); the weight average molecular weight ratio of PLLA to PDLA is 1:1.

[0019] Preferably, in step 1, the ratio of the yeast cell suspension, the buffer solution and the PDDA solution is (1 mL to 5 mL): (1 mL to 3 mL): (1 mL to 3 mL); and the pH of the buffer solution is 5 to 7.

[0020] Preferably, in step 2, the ratio of the buffer solution to the nano-SiO2 dispersion is (1 mL to 3 mL): (1 mL to 3 mL), and the pH of the buffer solution is 5 to 7.

[0021] Preferably, in step 3, the ratio of the buffer solution to the PLLA solution is (1 mL-3 mL):(1 mL-3 mL), and the pH of the buffer solution is 5-7.

[0022] Preferably, in step 4, the ratio of the buffer solution to the PDLA solution is (1 mL-3 mL):(1 mL-3 mL), and the pH of the buffer solution is 5-7.

[0023] Preferably, the oscillation is carried out in an air bath constant temperature oscillator, the temperature of the air bath constant temperature oscillator is 20-40 DEG C, the rotation speed of the air bath constant temperature oscillator is 150-250 rpm, and the oscillation time of the air bath constant temperature oscillator is 20-60 min.

[0024] The application provides a single-cell surface stereocomplex polylactic acid silica radiation refrigeration shell material.

[0025] Compared with the prior art, the application has the following beneficial effects:

[0026] The application provides a preparation method of a single-cell surface stereocomplex polylactic acid silica radiation refrigeration shell material. Compared with the prior art, the application has the following beneficial effects: BRIEF DESCRIPTION OF DRAWINGS

[0027] In order to more clearly illustrate the technical solutions of the embodiments of the application, the following will briefly introduce the drawings needed to be used in the embodiments. It should be understood that the following drawings only show some of the embodiments of the application, and therefore should not be regarded as a limitation to the scope. For those skilled in the art, other related drawings can also be obtained without creative labor on the basis of these drawings.

[0028] Figure 1Schematic diagram of the preparation of stereocomposite polylactic acid / silica radiative cooling nanoshells on the surface of yeast single cells according to the present invention ((a) yeast cell surface with negative charge; (b) yeast cell surface after assembly with PDDA nanoshell; (c) yeast cell surface after assembly with SiO2 nanoshell; (d) yeast cell surface after assembly with PLLA nanoshell; (e) yeast cell surface after assembly with PDLA nanoshell).

[0029] Figure 2 These are the C1s high-resolution XPS spectra of yeast cells and PDDA, SiO2, PLLA, and PDLA-encapsulated cells layer by layer during the preparation of the radiorefrigeration nanoshells of the present invention ((a) is yeast cells; (b) is Cell@PDDA; (c) is Cell@PDDA@SiO2; (d) is Cell@PDDA@SiO2@PLLA; (e) is Cell@PDDA@SiO2@SC-PLA);

[0030] Figure 3 This is the ethanol production by the Cell@PDDA@SiO2@SC-PLA prepared by the present invention and free yeast cells under photothermal fermentation. DETAILED DESCRIPTION

[0031] To make the objectives, technical solutions, and advantages of the embodiments of the present invention more clear, the technical solutions of the embodiments of the present invention will be clearly and completely described below in conjunction with the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Generally, the components of the embodiments of the present invention described and shown in the drawings herein can be arranged and designed in various different configurations.

[0032] Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the invention as claimed, but rather merely represents selected embodiments of the present invention. All other embodiments derived by persons of ordinary skill in the art based on the embodiments of the present invention without creative effort shall fall within the scope of protection of the present invention.

[0033] It should be noted that similar reference numerals and letters denote similar items in the following drawings, and therefore, once an item is defined in one drawing, it does not need to be further defined or explained in subsequent drawings.

[0034] The present invention is described in further detail below with reference to the accompanying drawings:

[0035] The present invention is a method for preparing a single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material, comprising the following steps:

[0036] Step 1, dissolve PDDA in PBS buffer solution, stir until completely dissolved to prepare PDDA solution.

[0037] The ratio of PDDA to PBS buffer solution is (0.01g-0.05g):(5mL-20mL), and the pH of the PBS buffer solution is 5-7.

[0038] Weigh nano-SiO2 and disperse it in PBS buffer solution, and ultrasonic until uniformly dispersed to prepare nano-SiO2 dispersion.

[0039] The ratio of nano-SiO2 to PBS buffer solution is (0.01g-0.05g):(5mL-20mL), the particle size of nano-SiO2 is 20nm-100nm, the ultrasonic time is 20min-60min, and the pH of the PBS buffer solution is 5-7.

[0040] Prepare HFIP solution using PBS buffer solution, dissolve PLLA and PDLA in HFIP solution respectively, and heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0041] The mass percentage concentration of HFIP is 5%-10%, and the ratio of HFIP solution, PLLA and PDLA is (5mL-20mL):(0.01g-0.05g):(0.01g-0.05g).

[0042] The weight average molecular weight ratio of PLLA and PDLA is 1:1.

[0043] Step 2, take the yeast cell suspension and place it in a centrifuge tube, centrifuge and wash it with PBS buffer solution for 3 times, then place the yeast cells in PBS buffer solution to obtain yeast cell resuspension. Then add the PDDA solution prepared in step (1), and put it into an air bath constant temperature oscillator. Then, centrifuge and wash it with PBS buffer solution for 3 times to obtain yeast cells with PDDA nanoshell assembled on the surface, marked as Cell@PDDA.

[0044] The ratio of yeast cell suspension, buffer solution and PDDA solution is (1mL-5mL):(1mL-3mL):(1mL-3mL), and the pH of the PBS buffer solution is 5-7.

[0045] The temperature of the air bath constant temperature oscillator is 20℃-40℃, the rotation speed of the air bath constant temperature oscillator is 150rpm-250rpm, and the oscillation time of the air bath constant temperature oscillator is 20min-60min.

[0046] Step 3: Place the Cell@PDDA prepared in step (2) in a PBS buffer solution, add the nano-SiO2 dispersion prepared in step (1), and oscillate in an air bath thermostat. Subsequently, wash three times by centrifugation with PBS buffer solution to obtain yeast cells with PDDA / SiO2 nanoshells assembled on their surfaces, designated as Cell@PDDA@SiO2.

[0047] The ratio of PBS buffer solution to nano-SiO2 dispersion is (1 mL-3 mL): (1 mL-3 mL), and the pH value of PBS buffer solution is 5-7.

[0048] The temperature of the air bath constant temperature oscillator is 20° C. to 40° C., the speed of the air bath constant temperature oscillator is 150 rpm to 250 rpm, and the oscillation time of the air bath constant temperature oscillator is 20 min to 60 min.

[0049] Step 4: Place the Cell@PDDA@SiO2 prepared in step (3) in a PBS buffer solution, add the PLLA solution prepared in step (1), and oscillate in an air bath constant temperature oscillator. Subsequently, centrifuge and wash three times with PBS buffer solution to obtain yeast cells with PDDA / SiO2 / PLLA nanoshells assembled on their surfaces, designated as Cell@PDDA@SiO2@PLLA.

[0050] The ratio of PBS buffer solution to PLLA solution is (1 mL to 3 mL): (1 mL to 3 mL), and the pH of the PBS buffer solution is 5 to 7.

[0051] The temperature of the air bath constant temperature oscillator is 20° C. to 40° C., the speed of the air bath constant temperature oscillator is 150 rpm to 250 rpm, and the oscillation time of the air bath constant temperature oscillator is 20 min to 60 min.

[0052] Step 5: Place the Cell@PDDA@SiO2@PLLA prepared in step (4) in a PBS buffer solution, add the PDLA solution prepared in step (1), and oscillate in an air bath constant temperature oscillator. PLLA and PDLA form SC-PLA. Subsequently, the cells are washed three times by centrifugation in a PBS buffer solution to obtain yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated as Cell@PDDA@SiO2@SC-PLA.

[0053] The ratio of PBS buffer solution to PDLA solution is (1 mL to 3 mL): (1 mL to 3 mL), and the pH of the PBS buffer solution is 5 to 7.

[0054] The temperature of the air bath constant temperature oscillator is 20° C. to 40° C., the speed of the air bath constant temperature oscillator is 150 rpm to 250 rpm, and the oscillation time of the air bath constant temperature oscillator is 20 min to 60 min.

[0055] The following is a detailed description of this method:

[0056] Example 1

[0057] Weigh 0.03g of PDDA and dissolve it in 10mL of PBS buffer (0.01M, pH=6.5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.01g of 20nm nano-SiO2 and disperse it in 5mL of PBS buffer. Ultrasonicate for 20min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 10% HFIP solution. Weigh 0.05g of PLLA and 0.05g of PDLA, add 20mL of 10% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0058] 2 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 6.5). The yeast cells were then resuspended in 2 mL of PBS buffer to obtain a yeast cell suspension. 2 mL of the prepared PDDA solution was then added, and the tube was shaken in an air bath at 20°C and 150 rpm for 30 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0059] The prepared Cell@PDDA was placed in 2 mL of PBS buffer (0.01 M, pH 6.5), and 2 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator and shaken at 150 rpm at 20°C for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0060] The prepared Cell@PDDA@SiO2 was placed in 2 mL of PBS buffer (0.01 M, pH 6.5), and 2 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 150 rpm at 20°C for 30 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0061] The prepared Cell@PDDA@SiO2@PLLA was placed in 2 mL of PBS buffer (0.01 M, pH 6.5), and 2 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 150 rpm at 20°C for 30 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0062] Example 2

[0063] Weigh 0.02g of PDDA and dissolve it in 20mL of PBS buffer (0.01M, pH=7). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.04g of 50nm nano-SiO2 and disperse it in 20mL of PBS buffer. Ultrasonicate for 60min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 5% HFIP solution. Weigh 0.01g of PLLA and 0.01g of PDLA, add 5mL of 5% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0064] 3 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 7) by centrifugation. The yeast cells were then placed in 3 mL of PBS buffer to obtain a resuspension. 3 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath with a constant temperature shaker at 30°C and 250 rpm for 50 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to produce yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0065] The prepared Cell@PDDA was placed in 3 mL of PBS buffer (0.01 M, pH 7), and 3 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator at 30°C and 250 rpm for 50 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0066] The prepared Cell@PDDA@SiO2 was placed in 3 mL of PBS buffer (0.01 M, pH 7), and 3 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 250 rpm at 30°C for 50 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0067] The prepared Cell@PDDA@SiO2@PLLA was placed in 3 mL of PBS buffer (0.01 M, pH 7), and 3 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 250 rpm at 30°C for 50 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0068] Example 3

[0069] Weigh 0.01g of PDDA and dissolve it in 5mL of PBS buffer (0.01M, pH=5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.02g of 30nm nano-SiO2 and disperse it in 10mL of PBS buffer. Ultrasonicate for 30min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 10% HFIP solution. Weigh 0.02g of PLLA and 0.02g of PDLA, add 8mL of 10% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0070] 2 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 5) by centrifugation. The yeast cells were then resuspended in 1 mL of PBS buffer to obtain a yeast cell suspension. 1 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath oscillator at 35°C and 200 rpm for 35 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0071] The prepared Cell@PDDA was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared nano-SiO2 dispersion was added. The mixture was then placed in an air bath oscillator and shaken at 200 rpm at 35°C for 35 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0072] The prepared Cell@PDDA@SiO2 was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared PLLA solution was added. The cells were then placed in an air bath and shaken at 200 rpm at 35°C for 35 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0073] The prepared Cell@PDDA@SiO2@PLLA was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 35°C for 35 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0074] Example 4

[0075] Weigh 0.04g of PDDA and dissolve it in 15mL of PBS buffer (0.01M, pH=5.5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.02g of 100nm nano-SiO2 and disperse it in 20mL of PBS buffer. Ultrasonicate for 40min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare an 8% HFIP solution. Weigh 0.03g of PLLA and 0.03g of PDLA, add 10mL of 8% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0076] 1 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 5.5). The yeast cells were then placed in 1 mL of PBS buffer to obtain a resuspension. 1 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath with a constant temperature shaker at 40°C and 180 rpm for 30 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0077] The prepared Cell@PDDA was placed in 1 mL of PBS buffer (0.01 M, pH 5.5), and 1 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator at 40°C and 180 rpm for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0078] The prepared Cell@PDDA@SiO2 was placed in 1 mL of PBS buffer (0.01 M, pH 5.5), and 1 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath oscillator at 40°C and 180 rpm for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0079] The prepared Cell@PDDA@SiO2@PLLA was placed in 1 mL of PBS buffer (0.01 M, pH 5.5), and 1 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 180 rpm at 40°C for 30 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0080] Example 5

[0081] Weigh 0.05g of PDDA and dissolve it in 10mL of PBS buffer (0.01M, pH 6). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.05g of 80nm nano-SiO2 and disperse it in 20mL of PBS buffer. Ultrasonicate for 50 minutes until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 7% HFIP solution. Weigh 0.04g of PLLA and 0.04g of PDLA, add 10mL of 7% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0082] 2 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 6) by centrifugation. The yeast cells were then placed in 3 mL of PBS buffer to obtain a resuspension. 3 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath oscillator at 25°C and 200 rpm for 60 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to produce yeast cells with PDDA nanoshells on their surfaces, designated Cell@PDDA.

[0083] The prepared Cell@PDDA was placed in 3 mL of PBS buffer (0.01 M, pH 6), and 3 mL of the prepared nano-SiO2 dispersion was added. The mixture was then placed in an air bath oscillator and shaken at 200 rpm at 25°C for 60 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0084] The prepared Cell@PDDA@SiO2 was placed in 3 mL of PBS buffer (0.01 M, pH 6), and 3 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 25°C for 60 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0085] The prepared Cell@PDDA@SiO2@PLLA was placed in 3 mL of PBS buffer (0.01 M, pH 6), and 3 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 25°C, 200 rpm, for 60 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0086] Example 6

[0087] Weigh 0.02g of PDDA and dissolve it in 10mL of PBS buffer (0.01M, pH 6.5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.02g of 30nm nano-SiO2 and disperse it in 10mL of PBS buffer. Ultrasonicate for 30min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 7% HFIP solution. Weigh 0.02g of PLLA and 0.02g of PDLA, add 10mL of 7% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0088] 2 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 6.5). The yeast cells were then resuspended in 1 mL of PBS buffer to obtain a yeast cell suspension. 1 mL of the prepared PDDA solution was then added, and the suspension was shaken in an air bath at 30°C and 200 rpm for 30 minutes. Subsequently, the suspension was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells on their surfaces, designated Cell@PDDA.

[0089] The prepared Cell@PDDA was placed in 1 mL of PBS buffer (0.01 M, pH 6.5), and 1 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator and shaken at 200 rpm at 30°C for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0090] The prepared Cell@PDDA@SiO2 was placed in 1 mL of PBS buffer (0.01 M, pH 6.5), and 1 mL of the prepared PLLA solution was added. The cells were then placed in an air bath and shaken at 200 rpm at 30°C for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0091] The prepared Cell@PDDA@SiO2@PLLA was placed in 1 mL of PBS buffer (0.01 M, pH 6.5), and 1 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 30°C for 30 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0092] Example 7

[0093] Weigh 0.02g of PDDA and dissolve it in 6mL of PBS buffer (0.01M, pH 6). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.05g of 30nm nano-SiO2 and disperse it in 15mL of PBS buffer. Ultrasonicate for 50min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare an 8% HFIP solution. Weigh 0.03g of PLLA and 0.03g of PDLA, add 15mL of 8% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0094] 1 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with 0.01 M PBS buffer (pH 6). The yeast cells were then resuspended in 2 mL of PBS buffer to obtain a yeast cell suspension. 2 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath oscillator at 35°C and 200 rpm for 20 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells on their surfaces, designated Cell@PDDA.

[0095] The prepared Cell@PDDA was placed in 2 mL of PBS buffer (0.01 M, pH 6), and 2 mL of the prepared nano-SiO2 dispersion was added. The mixture was then placed in an air bath oscillator and shaken at 200 rpm at 35°C for 20 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0096] The prepared Cell@PDDA@SiO2 was placed in 2 mL of PBS buffer (0.01 M, pH 6), and 2 mL of the prepared PLLA solution was added. The cells were then placed in an air bath and shaken at 200 rpm at 35°C for 20 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0097] The prepared Cell@PDDA@SiO2@PLLA was placed in 2 mL of PBS buffer (0.01 M, pH 6.5), and 2 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 35°C for 20 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0098] Example 8

[0099] Weigh 0.04g of PDDA and dissolve it in 10mL of PBS buffer (0.01M, pH=5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.02g of 60nm nano-SiO2 and disperse it in 15mL of PBS buffer. Ultrasonicate for 40min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 7% HFIP solution. Weigh 0.02g of PLLA and 0.02g of PDLA, add 10mL of 7% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0100] 2 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 5) by centrifugation. The yeast cells were then placed in 1 mL of PBS buffer to obtain a resuspension. 1 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath oscillator at 30°C and 250 rpm for 50 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to produce yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0101] The prepared Cell@PDDA was placed in 1 mL of PBS buffer (0.01 M, pH 5), and 1 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator at 30°C and 250 rpm for 50 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0102] The prepared Cell@PDDA@SiO2 was placed in 1 mL of PBS buffer (0.01 M, pH 5), and 1 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 250 rpm at 30°C for 50 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0103] The prepared Cell@PDDA@SiO2@PLLA was placed in 1 mL of PBS buffer (0.01 M, pH 5), and 1 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 250 rpm at 30°C for 50 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0104] Example 9

[0105] Weigh 0.05g of PDDA and dissolve it in 20mL of PBS buffer (0.01M, pH 6.5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.03g of 40nm nano-SiO2 and disperse it in 10mL of PBS buffer. Ultrasonicate for 40min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare a 6% HFIP solution. Weigh 0.05g of PLLA and 0.05g of PDLA, add 15mL of 6% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0106] 5 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 6.5). The yeast cells were then resuspended in 3 mL of PBS buffer to obtain a yeast cell suspension. 3 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath oscillator at 40°C and 200 rpm for 40 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells assembled on their surfaces, designated Cell@PDDA.

[0107] The prepared Cell@PDDA was placed in 3 mL of PBS buffer (0.01 M, pH 6.5), and 3 mL of the prepared nano-SiO2 dispersion was added. The mixture was then placed in an air bath oscillator and shaken at 200 rpm at 40°C for 40 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0108] The prepared Cell@PDDA@SiO2 was placed in 3 mL of PBS buffer (0.01 M, pH 6.5), and 3 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 40°C for 40 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0109] The prepared Cell@PDDA@SiO2@PLLA was placed in 3 mL of PBS buffer (0.01 M, pH 6.5), and 3 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 200 rpm at 40°C for 40 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the cells were washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0110] Example 10

[0111] Weigh 0.02g of PDDA and dissolve it in 20mL of PBS buffer (0.01M, pH=5). Stir until completely dissolved to prepare a PDDA solution. Weigh 0.02g of 30nm nano-SiO2 and disperse it in 10mL of PBS buffer. Ultrasonicate for 30min until uniformly dispersed to prepare a nano-SiO2 dispersion. Use PBS buffer to prepare an 8% HFIP solution. Weigh 0.01g of PLLA and 0.01g of PDLA, add 8mL of 8% HFIP solution respectively, heat and stir until completely dissolved to prepare PLLA solution and PDLA solution.

[0112] 3 mL of yeast cell suspension was placed in a centrifuge tube and washed three times with PBS buffer (0.01 M, pH 5) by centrifugation. The yeast cells were then resuspended in 2 mL of PBS buffer to obtain a yeast cell suspension. 2 mL of the prepared PDDA solution was then added, and the tube was placed in an air bath with a constant temperature shaker at 20°C and 150 rpm for 30 minutes. Subsequently, the tube was washed three times with PBS buffer by centrifugation to obtain yeast cells with PDDA nanoshells on their surfaces, designated Cell@PDDA.

[0113] The prepared Cell@PDDA was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared nano-SiO2 dispersion was added. The cells were then placed in an air bath oscillator and shaken at 150 rpm at 20°C for 30 minutes. Subsequently, the cells were washed three times by centrifugation with PBS buffer to obtain yeast cells with surface-assembled PDDA / SiO2 nanoshells, designated Cell@PDDA@SiO2.

[0114] The prepared Cell@PDDA@SiO2 was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared PLLA solution was added. The mixture was then placed in an air bath and shaken at 150 rpm at 20°C for 30 minutes. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / PLLA nanoshells, designated Cell@PDDA@SiO2@PLLA.

[0115] The prepared Cell@PDDA@SiO2@PLLA was placed in 2 mL of PBS buffer (0.01 M, pH 5), and 2 mL of the prepared PDLA solution was added. The mixture was then placed in an air bath and shaken at 150 rpm at 20°C for 30 minutes. PLLA and PDLA formed SC-PLA. Subsequently, the mixture was washed three times by centrifugation with PBS buffer to produce yeast cells with surface-assembled PDDA / SiO2 / SC-PLA nanoshells, designated Cell@PDDA@SiO2@SC-PLA.

[0116] Example 1 of the present invention is taken as experimental group 1, Example 2 is taken as experimental group 2, Example 3 is taken as experimental group 3, Example 4 is taken as experimental group 4, Example 5 is taken as experimental group 5, Example 6 is taken as experimental group 6, Example 7 is taken as experimental group 7, Example 8 is taken as experimental group 8, Example 9 is taken as experimental group 9, and Example 10 is taken as experimental group 10.

[0117] The immobilized cells Cell@PDDA@SiO2@SC-PLA prepared in the above 10 experiments were subjected to ethanol fermentation experiments under photothermal conditions and analyzed to obtain the following results:

[0118] A gas chromatograph was used to measure ethanol production under photothermal conditions in 10 experimentally prepared immobilized cells (Cell@PDDA@SiO2@SC-PLA). XPS characterized the layer-by-layer shells on the surface of the single cells and analyzed their chemical structure. Example 6 achieved the best results, achieving the highest ethanol yield.

[0119] Figure 1 (a) The yeast cell surface is negatively charged; (b) is the yeast cell surface after assembling the PDDA nanoshell; (c) is the yeast cell surface after assembling the SiO2 nanoshell; (d) is the yeast cell surface after assembling the PLLA nanoshell; (e) is the yeast cell surface after assembling the PDLA nanoshell.

[0120] Figure 2 (a) is the C1s XPS spectrum of yeast cells, 284.80 eV, 286.44 eV and 288.20 eV are attributed to the CC / CH, CO and C=O bonds of yeast cells, respectively. Figure 2 (b) is the C1s XPS spectrum of Cell@PDDA. 284.80eV, 286.66eV and 287.96eV are attributed to the CC / CH of Cell@PDDA and the CO and C=O bonds of yeast cells, respectively. 286.16eV is attributed to the CN of PDDA. + , proving the successful preparation of Cell@PDDA. Figure 2 (c) is the C1s XPS spectrum of Cell@PDDA@SiO2. The 284.80eV, 286.43eV and 287.84eV are attributed to the CC / CH of Cell@PDDA and the CO and C=O bonds of yeast cells, respectively. + The disappearance of the peak is due to the assembly of SiO2 nanoshells blocking the CN + The signal was detected, proving the successful assembly of the SiO2 nanoshell. Figure 2(d) is the C1s XPS spectrum of Cell@PDDA@SiO2@PLLA. 284.80 eV, 286.48 eV and 287.52 eV are attributed to the CC / CH, CO and C=O bonds of Cell@PDDA@SiO2@PLLA, respectively, and 289.06 eV is attributed to the OC=O of PLLA, indicating that PLLA is successfully assembled on the surface of immobilized cells. Figure 2 (e) is the C1s XPS spectrum of Cell@PDDA@SiO2@SC-PLA. 284.80eV, 286.46eV and 287.57eV are attributed to the CC / CH, CO and C=O bonds of Cell@PDDA@SiO2@SC-PLA, respectively. 288.91eV is attributed to the OC=O of SC-PLA. Meanwhile, the CC / CH peak is higher than Figure 2 (d), demonstrating the successful preparation of Cell@PDDA@SiO2@SC-PLA.

[0121] Figure 3 Figure 2 is the ethanol yield of Cell@PDDA@SiO2@SC-PLA and free yeast cells under photothermal fermentation. The ethanol yield of Cell@PDDA@SiO2@SC-PLA is 1.3 mg / mL, which is higher than that of free yeast cells.

[0122] The above are merely preferred embodiments of the present invention and are not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the present invention.

Claims

1. A method for preparing a single cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material, characterized in that: The steps include: Step 1: After adding PDDA solution to the yeast cell resuspension and shaking, the mixture was centrifuged and washed with a buffer solution to prepare yeast cells with PDDA nanoshells assembled on their surfaces, which were designated as Cell@PDDA. Step 2: Cell@PDDA was placed in a buffer solution, nano-SiO2 dispersion was added and shaken, and then centrifuged and washed with the buffer solution to obtain yeast cells with PDDA / SiO2 nanoshells assembled on their surfaces, which were recorded as Cell@PDDA@SiO2. Step 3: Cell@PDDA@SiO2 was placed in a buffer solution, PLLA solution was added and shaken, and then centrifuged and washed with the buffer solution to obtain yeast cells with PDDA / SiO2 / PLLA nanoshells assembled on their surfaces, which were recorded as Cell@PDDA@SiO2@PLLA. Step 4: Place Cell@PDDA@SiO2@PLLA in a buffer solution, add PDLA solution and shake, then centrifuge and wash with buffer solution to obtain yeast cells with PDDA / SiO2 / SC-PLA nanoshells assembled on the surface, which are recorded as Cell@PDDA@SiO2@SC-PLA.

2. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: The preparation method of the PDDA solution is as follows: Dissolve PDDA in PBS buffer solution and stir until completely dissolved to prepare PDDA solution; The ratio of PDDA to PBS buffer solution is (0.01 g to 0.05 g): (5 mL to 20 mL), and the pH of the PBS buffer solution is 5 to 7.

3. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: The preparation method of the nano-SiO2 dispersion is as follows: Nano-SiO2 was dispersed in PBS buffer solution and ultrasonicated until uniformly dispersed to prepare nano-SiO2 dispersion. The ratio of nano-SiO2 to PBS buffer solution is (0.01g-0.05g):(5mL-20mL), the particle size of nano-SiO2 is 20nm-100nm, the ultrasonic time is 20min-60min, and the pH value of the PBS buffer solution is 5-7.

4. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: The preparation methods of the PLLA solution and the PDLA solution are as follows: HFIP solution was prepared using PBS buffer solution, PLLA and PDLA were dissolved in HFIP solution respectively, and heated and stirred until completely dissolved to prepare PLLA solution and PDLA solution; The mass percentage concentration of the HFIP solution is 5% to 10%, and the ratio of the HFIP solution, PLLA and PDLA is (5 mL to 20 mL):(0.01 g to 0.05 g):(0.01 g to 0.05 g); the weight average molecular weight ratio of PLLA to PDLA is 1:

1.

5. The method for preparing the single cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: In step 1, the ratio of yeast cell suspension, buffer solution and PDDA solution is (1 mL to 5 mL): (1 mL to 3 mL): (1 mL to 3 mL); the pH of the buffer solution is 5 to 7.

6. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: In step 2, the ratio of the buffer solution to the nano-SiO2 dispersion is (1 mL to 3 mL): (1 mL to 3 mL), and the pH value of the buffer solution is 5 to 7.

7. The method for preparing the single cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: In step 3, the ratio of the buffer solution to the PLLA solution is (1 mL to 3 mL): (1 mL to 3 mL), and the pH of the buffer solution is 5 to 7.

8. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: In step 4, the ratio of the buffer solution to the PDLA solution is (1 mL to 3 mL): (1 mL to 3 mL), and the pH of the buffer solution is 5 to 7.

9. The method for preparing the single-cell surface stereocomposite polylactic acid-silica radiation refrigeration shell material according to claim 1, characterized in that: Oscillate in an air bath constant temperature oscillator, the temperature of the air bath constant temperature oscillator is 20°C to 40°C, the speed of the air bath constant temperature oscillator is 150 rpm to 250 rpm, and the oscillation time of the air bath constant temperature oscillator is 20 min to 60 min.

10. A single cell surface stereocomposite polylactic acid silica radiation refrigeration shell material, characterized in that: The method is prepared by any one of claims 1 to 9.

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