CN113800622B

CN113800622B - Method and device for relieving photoinhibition of photosynthetic microorganisms and application of method and device

Info

Publication number: CN113800622B
Application number: CN202111182203.3A
Authority: CN
Inventors: 孙健; 姚欣媛; 白晓燕
Original assignee: Guangdong University of Technology
Current assignee: Guangdong University of Technology
Priority date: 2021-10-11
Filing date: 2021-10-11
Publication date: 2023-08-11
Anticipated expiration: 2041-10-11
Also published as: CN113800622A

Abstract

The application belongs to the technical field of bioelectrochemistry, and particularly relates to a method and a device for relieving photoinhibition of photosynthetic microorganisms and application thereof. The application provides a method for relieving photoinhibition of photosynthetic microorganisms; wherein the method for relieving photoinhibition of photosynthetic microorganisms comprises the steps of: the photosynthetic microorganisms are irradiated with light to provide hydrogen and carbon sources for the photosynthetic microorganisms and adhere the photosynthetic microorganisms to the working electrode, and partial generated photosynthetic electrons are transferred from the photosynthetic microorganisms to the working electrode, so that the photosynthesis of the photosynthetic microorganisms, synthesis of organic matters such as proteins and the like and metabolism of the synthesized organic matters in vivo are promoted, and the photoinhibition of the photosynthetic microorganisms is relieved.

Description

Method and device for relieving photoinhibition of photosynthetic microorganisms and application of method and device

Technical Field

The application belongs to the technical field of bioelectrochemistry, and particularly relates to a method and a device for relieving photoinhibition of photosynthetic microorganisms and application thereof.

Background

Photosynthetic microorganisms are microorganisms living with light as the only or main energy source, and can synthesize organic matters in vivo by utilizing various inorganic matters and organic matters in nature under illumination conditions, so that the photosynthetic microorganism treatment is a common method for removing inorganic and organic pollutants in wastewater.

However, since the self-protection reaction of the photosynthetic system of the photosynthetic microorganism itself, namely the photoinhibition of the photosynthetic microorganism, is specifically represented by the phenomenon that the photosynthetic function of the photosynthetic microorganism is reduced when the illumination intensity exceeds the intensity that can be utilized by the photosynthetic system of the photosynthetic microorganism itself, the existing photosynthetic microorganism has low efficiency in removing inorganic and organic pollutants in wastewater and synthesizing organic matters such as proteins in vivo.

Disclosure of Invention

In view of the above, the present application provides a method and an apparatus for removing photoinhibition of photosynthetic microorganisms, and an application thereof, which promote the growth and metabolism of the photosynthetic microorganisms by applying electrical stimulation to the photosynthetic microorganisms, and extract more photosynthetic electrons, thereby removing photoinhibition of the photosynthetic microorganisms, and solving the problems of the prior art that the photosynthetic function of the photosynthetic microorganisms is reduced and photoinhibition occurs when the illumination intensity exceeds the intensity available to the photosynthetic system of the photosynthetic microorganisms themselves.

The first aspect of the present application provides a method for relieving photoinhibition of photosynthetic microorganisms, comprising the steps of:

step one, applying light to photosynthetic microorganisms;

providing a hydrogen body and carbon source for the photosynthetic microorganism;

attaching the photosynthetic microorganisms to an electrode;

step four, applying an electrode potential to the electrode;

the electrode is a working electrode in a three-electrode electrochemical system.

It is noted that, the photosynthetic microorganism is attached to the working electrode in the three-electrode electrochemical system, and the three-electrode electrochemical system forms a closed loop, and electrode potential is applied at the same time, so that the growth and metabolism of the photosynthetic microorganism are stimulated, more photosynthetic electrons can be extracted and transferred to the working electrode, extracellular diversion is realized, the photosynthetic metabolism of the photosynthetic microorganism is enhanced, and the purpose of relieving the photoinhibition of the photosynthetic microorganism is achieved.

Preferably, the applying electrode potential comprises: an electrode potential of-0.3V, 0V, +0.3V was applied.

Although the electrical stimulation accelerates the growth and metabolism of the photosynthetic microorganisms and releases the photoinhibition, if the applied electrode potential is too large, a large amount of photosynthetic microorganisms die, and therefore, the control of the applied electrode potential is beneficial to avoiding the death of a large amount of photosynthetic microorganisms.

Preferably, the hydrogen-body-carbon source includes: one, two or more of ammonia nitrogen, phosphate and sulfadiazine.

The ammonia nitrogen, the phosphate and the sulfadiazine are common inorganic and organic pollutants in the domestic and industrial wastewater, and photosynthetic microorganisms growing in the domestic and industrial wastewater can adapt to the domestic and industrial wastewater containing the ammonia nitrogen, the phosphate and the sulfadiazine due to natural domestication, and utilize the ammonia nitrogen, the phosphate and the sulfadiazine to carry out photosynthesis, growth and metabolism and synthesize organic matters such as protein.

In a second aspect, the present application provides an apparatus for performing a method for relieving photoinhibition of photosynthetic microorganisms, comprising:

a photosynthetic microorganism photosynthetic reactor, an electrochemical workstation;

the working electrode of the electrochemical workstation is placed in the photosynthetic microorganism photosynthetic reactor.

The photosynthetic microorganisms in the photosynthetic microorganism photosynthetic reactor can utilize hydrogen and carbon sources to carry out photosynthesis, organic matters are synthesized in the body and metabolized, so that photosynthetic electrons are generated, and the photosynthetic microorganisms are loaded on the working electrode of the electrochemical workstation, so that the working electrode can transfer the photosynthetic electrons generated in the photosynthesis and organic matter metabolism processes of the photosynthetic microorganisms, the photosynthesis of the photosynthetic microorganisms is promoted, and the photoinhibition of the photosynthetic microorganisms is relieved.

Preferably, the photosynthetic microorganism photosynthetic reactor comprises a light source, a vessel;

the container may contain a gas, liquid or solid containing a hydrogen and carbon source and a photosynthetic microorganism culture solution, and the working electrode carrying the photosynthetic microorganism is placed in the gas, liquid or solid containing the hydrogen and carbon source, and the photosynthetic microorganism performs photosynthesis to synthesize an organic substance and the organic substance decomposed and synthesized in the body performs growth metabolism under irradiation of light source simulated sunlight.

Preferably, the working electrode is a graphite plate.

Graphite has excellent stability and conductivity, and can avoid interference when being used as a working electrode of an electrochemical workstation, and transfer photosynthetic electrons, thus well reflecting photosynthesis and growth metabolism of photosynthetic microorganisms on the working electrode.

Preferably, the light source is one, two or more of a flame, a candle, an incandescent lamp, a sodium lamp, a mercury lamp, a fluorescent lamp, an LED, and an OLED.

Preferably, the light source is an OLED.

The relative color temperature (CCT) of the incandescent lamp is about 2700K, the relative color temperature (CCT) of the cold fluorescent lamp is about 4000-5000K, which is far lower than 2500-8000K of sunlight, and the relative color temperature (CCT) of the OLED is close to sunlight, so that the OLED has the performance of simulating sunlight, and further, photosynthesis of photosynthetic microorganisms by utilizing hydrogen and a carbon source can be better promoted.

Preferably, the graphite plate has a size of 30×20×5mm.

Preferably, the graphite sheet is a treated graphite sheet.

Preferably, the treatment method comprises the following steps: sequentially placing the graphite plates into acetone and deionized water, respectively carrying out ultrasonic treatment for 10min and 20min, and then placing the graphite plates into an oven for drying for later use. And putting the electrode into an oven for drying after the electrode is ultrasonically treated with deionized water for 20 min.

Preferably, the distance between the light source and the working electrode is 5cm.

Preferably, the container is a glass container.

The glass container has good light transmittance, so that the light of the light source can be irradiated on the surface of the photosynthetic microorganism as much as possible, and the growth and metabolism of the photosynthetic microorganism are accelerated.

In a third aspect, the application provides a method and apparatus for relieving photoinhibition of photosynthetic microorganisms, and application thereof in the field of wastewater treatment.

Nitrogen and phosphorus contained in the wastewater promote the growth of algae and aquatic plants, thereby deteriorating water quality and reducing dissolved oxygen. The organic pollutant antibiotics in the wastewater generally coexist with pollutants containing nitrogen and phosphorus, the antibiotics can have extremely low concentration to influence the environment, and have the characteristics of durability, nondegradability and accumulation in the environment, the existing wastewater treatment efficiency by utilizing the photosynthetic microorganisms is not high due to the photoinhibition phenomenon of the photosynthetic microorganisms, and the growth metabolism and photosynthesis of the photosynthetic microorganisms can be promoted by relieving the photoinhibition of the photosynthetic microorganisms, so that the aim of improving the wastewater treatment efficiency of the photosynthetic microorganisms is fulfilled.

In summary, the application provides a method and a device for removing photoinhibition of photosynthetic microorganisms and application thereof; wherein the method for relieving photoinhibition of photosynthetic microorganisms comprises the steps of: applying light to the photosynthetic microorganism to provide a hydrogen body and a carbon source for the photosynthetic microorganism and attach the photosynthetic microorganism to the working electrode, when the photosynthetic microorganism utilizes the hydrogen body and the carbon source to carry out photosynthesis under the light condition, organic matters are synthesized in the body and the organic matters are accompanied with the generation of photosynthetic electrons in the metabolic process of the photosynthetic microorganism in the body, and when part of generated photosynthetic electrons are transferred from the photosynthetic microorganism to the working electrode, the photosynthesis of the photosynthetic microorganism and the metabolism of the synthesized organic matters in the body are promoted, so that the photoinhibition of the photosynthetic microorganism is relieved; therefore, the application provides a method and a device for relieving the photoinhibition of photosynthetic microorganisms and application thereof, which can solve the technical problem that the photosynthetic function of the photosynthetic microorganisms is reduced when the illumination intensity exceeds the intensity available by the photosynthetic system of the photosynthetic microorganisms in the prior art.

Description of the drawings:

in order to more clearly illustrate the embodiments of the application or the technical solutions of the prior art, the drawings which are used in the description of the embodiments or the prior art will be briefly described, it being obvious that the drawings in the description below are only some embodiments of the application, and that other drawings can be obtained from these drawings without inventive faculty for a person skilled in the art.

FIG. 1 shows ammonia nitrogen removal efficiency at 3000Lux light intensity in accordance with the present application;

FIG. 2 is a graph showing ammonia nitrogen removal efficiency of the present application at 8000Lux light intensity;

FIG. 3 is a graph showing ammonia nitrogen removal efficiency of the present application at 23000Lux illumination intensity;

FIG. 4 is a graph showing the effect of phosphate removal at 3000Lux light intensity according to the present application;

FIG. 5 is a graph showing the effect of phosphate removal at 8000Lux light intensity according to the present application;

FIG. 6 is a graph showing the effect of phosphate removal at 23000Lux light intensity according to the present application;

FIG. 7 is a graph showing the effect of antibiotic degradation under 3000Lux illumination according to the present application;

FIG. 8 is a graph showing the effect of antibiotic degradation of the present application under 8000Lux illumination;

FIG. 9 is a graph showing the effect of antibiotic degradation under 23000Lux illumination according to the present application;

FIG. 10 is a stacked bar graph of photosynthetic microorganism protein content under illumination intensity of 3000Lux, 8000Lux, 23000Lux according to the present application.

The specific embodiment is as follows:

the application provides a method and a device for relieving photoinhibition of photosynthetic microorganisms and application thereof, which can solve the technical problem that the photosynthetic function of the photosynthetic microorganisms is reduced when the illumination intensity exceeds the intensity available by the photosynthetic system of the photosynthetic microorganisms in the prior art.

The following description of the technical solutions in the embodiments of the present application will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present application, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the application without making any inventive effort, are intended to be within the scope of the application.

Among them, the reagents and raw materials used in the following examples are all commercially available or homemade.

Example 1

The embodiment provides a first method for relieving photoinhibition of photosynthetic microorganisms, which comprises the following steps:

firstly, using a graphite plate as a working electrode of an electrochemical workstation, using a titanium wire as a counter electrode, and using a saturated calomel electrode as a reference electrode to construct a three-electrode electrochemical system;

placing a graphite plate working electrode loaded with photosynthetic microorganisms, a titanium wire counter electrode, a saturated calomel electrode and photosynthetic microorganism culture solution in a glass container to form a closed loop;

controlling the OLED lamp to irradiate the glass container with light intensities of 3000lux, 8000lux and 23000 lux;

and step four, applying an electrode potential of 0V to a working electrode of the three-electrode electrochemical system.

Wherein the photosynthetic microorganisms are photosynthetic microorganisms which are collected in factory wastewater and are in logarithmic growth phase, and the concentration of ammonia nitrogen in the photosynthetic microorganism culture solution is 125mg/L.

The working electrode, the counter electrode and the reference electrode in the three-electrode electrochemical system form a closed loop, and under the condition that 0V electrode potential is applied to the counter electrode, the photosynthetic microorganisms are electrically stimulated by being attached to the working electrode, so that the growth metabolism is quickened, more photosynthetic electrons generated in the growth metabolism process are guided outwards through the working electrode, the photosynthetic metabolism of the photosynthetic microorganisms is promoted, and the purpose of relieving the photoinhibition of the photosynthetic microorganisms is achieved.

Example 2

This example provides a second method of relieving photoinhibition of photosynthetic microorganisms, which differs from example 1 in that it further comprises a step five of applying an electric potential of-0.3V through an electrochemical workstation.

Example 3

This example provides a third method of relieving photoinhibition of photosynthetic microorganisms, which differs from example 1 in that it further comprises a step five of applying a potential of +0.3v through an electrochemical workstation.

Example 4

The present embodiment provides a fourth method for relieving photoinhibition of photosynthetic microorganisms, comprising the steps of:

Wherein the photosynthetic microorganisms are photosynthetic microorganisms which are collected in factory wastewater and in logarithmic growth phase, and the concentration of phosphate in the photosynthetic microorganism culture solution is 15mg/L.

Example 5

This example provides a fifth method for relieving photoinhibition of photosynthetic microorganisms, which differs from example 4 in that it further comprises a step five of applying an electric potential of-0.3V through an electrochemical workstation.

Example 6

This example provides a sixth method for relieving photoinhibition of photosynthetic microorganisms, which differs from example 4 in that it further comprises a step five of applying an electric potential of +0.3V through an electrochemical workstation.

Example 7

The present embodiment provides a seventh method for relieving photoinhibition of photosynthetic microorganisms, comprising the steps of:

Wherein the photosynthetic microorganisms are photosynthetic microorganisms which are collected in factory wastewater and in logarithmic growth phase, and the concentration of sulfadiazine in the photosynthetic microorganism culture solution is 1mg/L.

Example 8

This example provides an eighth method for relieving photoinhibition of photosynthetic microorganisms, which differs from example 7 in that it further comprises a step five of applying an electric potential of-0.3V through an electrochemical workstation.

Example 9

This example provides a ninth method of relieving photoinhibition of photosynthetic microorganisms, which differs from example 7 in that it further comprises a step five of applying an electric potential of +0.3v through an electrochemical workstation.

Example 10

The present embodiment provides a tenth method for relieving photoinhibition of photosynthetic microorganisms, comprising the steps of:

step two, placing the working electrode of the graphite plate loaded with photosynthetic microorganisms, the titanium wire counter electrode, the saturated calomel electrode and the factory wastewater of the photosynthetic microorganism culture solution into a glass container to form a closed loop;

Wherein the photosynthetic microorganisms are photosynthetic microorganisms which are collected in factory wastewater and in logarithmic growth phase, the concentration of ammonia nitrogen in a photosynthetic microorganism culture solution is 125mg/L, the concentration of phosphate is 15mg/L, the concentration of sulfadiazine is 1mg/L, and the photosynthetic microorganism culture solution is prepared by a formula method which is conventional in the field.

Example 11

This example provides an eleventh method of relieving photoinhibition of photosynthetic microorganisms, which differs from example 10 in that it further comprises a step five of applying an electric potential of-0.3V through an electrochemical workstation.

Example 12

This example provides a twelfth method of relieving photoinhibition of photosynthetic microorganisms, which differs from example 10 in that it further comprises a step five of applying an electric potential of +0.3v through an electrochemical workstation.

Comparative example 1:

placing the photosynthetic microorganism loaded in a glass container containing photosynthetic microorganism culture solution;

and secondly, controlling the OLED lamp to irradiate the glass container with light intensities of 3000lux, 8000lux and 23000 lux.

The photosynthetic microorganisms are photosynthetic microorganisms which are collected in factory wastewater and in logarithmic growth phase, the concentration of ammonia nitrogen in a photosynthetic microorganism culture solution is 125mg/L, the concentration of phosphate is 15mg/L, the concentration of sulfadiazine is 1mg/L, and the preparation method of the photosynthetic microorganism culture solution is the prior art and is not described here.

Example 13

This example is an example of measuring the efficiency of removing ammonia nitrogen from a photosynthetic microorganism culture broth in the method of removing photoinhibition of photosynthetic microorganisms described in examples 1-3.

The detection step comprises the following steps: when three repeatable currents are detected by an electrochemical workstation, starting timing, and taking out partial photosynthetic microorganism culture solution containing ammonia nitrogen at the time of 0, 6, 12, 19, 24, 36, 48, 72, 96 and 120 hours, wherein the partial photosynthetic microorganism culture solution is respectively recorded as samples N1-N10;

step two, filtering samples N1 to N10 with a water-based filter membrane with the aperture of 0.22 mu m, and then placing the filtered samples in a refrigerator at the temperature of 4 ℃ for preservation and test;

and step three, measuring the ammonia nitrogen concentration in the filtered samples N1-N10 by adopting a Nahner reagent spectrophotometry, wherein the result is shown in the attached figures 1-3 of the specification.

Example 14

This example is an example of measuring the efficiency of phosphate removal from photosynthetic microorganism culture broth in the method of relieving photoinhibition of photosynthetic microorganisms described in examples 4-6.

The detection step comprises the following steps: step one, when the electrochemical workstation detects three repeatable currents, starting timing, and taking out part of photosynthetic microorganism culture solution containing phosphate at the time of 0, 6, 12, 19, 24, 36, 48, 72, 96 and 120 hours, and respectively marking the photosynthetic microorganism culture solution as samples P1-P10;

step two, filtering the samples P1 to P10 with a water-based filter membrane with the aperture of 0.22 mu m, and then placing the filtered samples in a refrigerator at the temperature of 4 ℃ for preservation and test;

and step three, measuring the phosphate concentration in the filtered samples P1-P10 by adopting an ammonium molybdate spectrophotometry, wherein the result is shown in the attached figures 4-6 of the specification.

Example 15

This example is an example of measuring the efficiency of sulfadiazine removal from photosynthetic microorganism culture broth in the method of relieving photoinhibition of photosynthetic microorganisms described in examples 7 to 9.

The detection step comprises the following steps: step one, when three repeatable currents are detected by an electrochemical workstation, starting timing, and taking out part of photosynthetic microorganism culture solution containing sulfadiazine at the time of 0, 3, 6, 12, 24, 36, 48, 72, 96 and 120 hours, wherein the photosynthetic microorganism culture solution is respectively marked as samples S1-S10;

step two, filtering the samples S1 to S10 by using a water-based filter membrane with the aperture of 0.22 mu m, and then placing the filtered samples in a refrigerator at the temperature of 4 ℃ for preservation and test;

and step three, measuring the concentration of sulfadiazine in the filtered samples S1-S10 by adopting high performance liquid chromatography, wherein the result is shown in the attached figures 7-9 of the specification.

Example 16

This example is an example of measuring the protein content in a photosynthetic microorganism in the method for relieving photoinhibition of a photosynthetic microorganism described in examples 10 to 12.

The detection step comprises the following steps: step one, applying an electrode potential of 0V to the working electrode under 3000lux illumination intensity, scraping the photosynthetic biological film attached to the working electrode from the working electrode by using a sterile brush when three repeatable currents are detected by the electrochemical workstation, recording the illumination time h of the photosynthetic microorganisms, and centrifuging at 4000rpm for 5min so as to collect the photosynthetic microorganisms as much as possible. The total protein extraction kit was used to extract proteins. Mu.l TPEBuffer-I was added per 100. Mu.l cell pellet. The tube was vortexed for 1-2 minutes and then placed on ice. For each 100. Mu.l TPEBuffer-I used, 60. Mu.l TPE Buffer-II was added. The tube may be tapped with a finger to mix gently. The tube was vortexed for 30 seconds to achieve complete mixing, if necessary. The tube was placed in a boiling hot water bath for 30 seconds.

The tube was removed and vortexed for 30 seconds. This heating and vortexing process was repeated until a clear solution was seen. The tube was incubated in a boiling water bath for an additional 10 minutes, after which time the tube was centrifuged at 15000Xg for 5 minutes at 4℃to remove all cell debris. The supernatant, the solution containing the protein, was transferred to a clean tube. The supernatant may be stored at-20℃to 70℃and designated as sample CH1;

step two to step nine, the samples collected at an electrode potential of-0.3V, an electrode potential of +0.3V and an electrode potential of 8000lux, an electrode potential of +0.3V, an electrode potential of 8000lux, an electrode potential of +0.3V and an electrode potential of 23000lux, an electrode potential of 0V, an electrode potential of 23000lux, an electrode potential of +0.3V and an electrode potential of +0.3V were recorded as CH 2-CH 9;

and step ten, protein content in the filtered samples CH1 to CH9 is determined by using a BCA protein concentration determination kit, and the result is shown in figure 10 of the specification.

Example 17

This example is an example of detecting the protein content in the photosynthetic microorganism of comparative example 1.

Step one, the photosynthetic microorganisms in the glass vessel were placed in a centrifuge with a sterile brush and centrifuged at 4000rpm for 5min under an illumination intensity of 3000lux for h, thereby collecting as much photosynthetic microorganisms as possible. The total protein extraction kit was used to extract proteins. Mu.l TPE Buffer-I was added per 100. Mu.l cell pellet. The tube was vortexed for 1-2 minutes and then placed on ice. For each 100. Mu.l TPE Buffer-I used, 60. Mu.l TPE Buffer-II was added. The tube may be tapped with a finger to mix gently. The tube was vortexed for 30 seconds to achieve complete mixing, if necessary. The tube was placed in a boiling hot water bath for 30 seconds. The tube was removed and vortexed for 30 seconds. This heating and vortexing process was repeated until a clear solution was seen. The tube was incubated in a boiling water bath for an additional 10 minutes, after which time the tube was centrifuged at 15000Xg for 5 minutes at 4℃to remove all cell debris. The supernatant, the solution containing the protein, was transferred to a clean tube. The supernatant may be stored at-20℃to 70℃and designated as sample CH10;

step two to step three, respectively illuminating for h hours under 8000lux illumination intensity and 23000lux illumination intensity, and recording collected samples as CH 11-CH 12;

and step four, protein content in the filtered samples CH 10-CH 12 is determined by using a BCA protein concentration determination kit, and the result is shown in figure 10 of the specification.

It can be understood from FIG. 10 that the application of the electrode potential to stimulate the photosynthetic microorganisms does not cause the protein content in the photosynthetic microorganisms to be substantially the same in the strong light, the medium light and the weak light, which means that the photosynthetic microorganisms are photo-inhibited when the electrode potential is not applied, and the protein content in the photosynthetic microorganisms is significantly improved when the electrode potential of-0.3V, 0V and +0.3V is applied, which means that the application of the electrode potential can promote the growth and metabolism of the photosynthetic microorganisms, extract more photosynthetic electrons and relieve the photo-inhibition of the photosynthetic microorganisms.

The foregoing is merely a preferred embodiment of the present application and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present application, which are intended to be comprehended within the scope of the present application.

Claims

1. A method of relieving photoinhibition of a photosynthetic microorganism, comprising the steps of:

step one, applying illumination with 23000Lux illumination intensity to photosynthetic microorganisms;

providing a hydrogen body and carbon source for the photosynthetic microorganism, wherein the hydrogen body and carbon source comprises: ammonia nitrogen, phosphate and sulfadiazine;

attaching the photosynthetic microorganisms to an electrode;

step four, applying an electrode potential of-0.3V or +0.3V to the electrode;

the electrode is a working electrode in a three-electrode electrochemical system;

the photosynthetic microorganisms grow photosynthetic microorganisms in domestic and industrial wastewater.