KR101270906B1 - Fluidizable carrier with biofilm formation-inhibiting microorganisms immobilized therein and membrane water treatment apparatus using the same - Google Patents

Abstract

The present invention relates to a membrane water treatment technology that can at the same time obtain a membrane fouling mitigation effect by physical washing while reducing the membrane fouling of the membrane from the molecular biology perspective by maintaining the activity of the microorganisms inhibiting the formation of biofilms, A membrane comprising a water treatment reactor and a membrane module for water treatment, in which biofilm formation inhibiting microorganisms are immobilized and biofilm formation suppression microorganism immobilized fluidized bed carriers having fluidity by aeration in water, and biofilm formation suppression microorganism immobilized fluidized bed carriers are disposed therein. Provided is a water treatment device.

Description

Fluidized carrier with biofilm formation-inhibiting microorganisms immobilized therein and membrane water treatment apparatus using the same}

The present invention relates to a technology for suppressing biomembrane fouling by biofilm formed by growing on the membrane surface during operation of a membrane water treatment device, and more specifically, immobilizing a microorganism capable of inhibiting biofilm formation. The present invention relates to a fluidized bed carrier and a separator water treatment device capable of stably maintaining the permeation performance of the membrane for a long time without loss or death of biofilm formation inhibiting microorganisms.

As the recent shortage of water resources and deterioration of water quality, demand for water treatment technology is increasing rapidly worldwide. In order to cope with such a situation, global attention has been focused on water treatment technology, and membranes including reverse osmosis / nanofiltration processes used for seawater desalination and water treatment and membrane bioreactor (MBR) processes used for wastewater treatment. Water treatment processes are emerging due to advantages such as continuity of treatment, simplicity of process and high treatment efficiency. Indeed, data from the BBC research in 2008 predicted that the international MBR market size would increase from $ 296 million in 2008 to $ 488 million in 2013.

However, in the membrane water treatment process, which is attracting attention, microorganisms adhere to and grow on the surface of the membrane to form a biofilm, which causes membrane fouling, thereby reducing permeability, reducing the cleaning cycle and lifespan of the membrane, There is a problem in that the economic performance of the membrane water treatment process is deteriorated by lowering the filtration performance of the membrane such as an increase in energy consumption required for filtration.

To solve this problem, various researches have been conducted for the last 20 years, and various physical and chemical methods have been studied. The physical methods of the biofilm reduction technology on the surface of the separator are biodegradation induction by increasing shear force through aeration and biofilm desorption through backwashing, and chemical methods include polymer coagulant for increasing particle size. There are methods such as injection or modification to increase the hydrophilicity of the membrane surface. These studies have shown that the characteristics of the water treatment process where water and nutrients necessary for the growth of microorganisms are present everywhere, the characteristics of the membrane process where driving force exists in the direction of the filtration of the membrane, and once formed are high in external physical and chemical impacts. Due to the nature of the resistant biofilm itself, it has not been a satisfactory solution so far. This is because the complex biological structure derived from microorganisms is regarded as a simple pollutant such as scaling and organic adsorption, and has been approached to be solved by physical and chemical methodological approaches. There is a desperate need for an understanding of and a biological approach based on it.

On the other hand, microorganisms recognize surrounding cell density by synthesizing specific signal molecules in response to changes in various environmental conditions such as temperature, pH, nutrients, and releasing / absorbing them to the outside of cells. When the cell density increases and the concentration of these signal molecules reaches a certain level, expression of specific genes begins, resulting in group behavior regulation of the microbial population, which is called quorum sensing phenomena. In general, it occurs under high density of cells. Representative examples of such quorum sensing include symbiosis, virulence, competetion, conjugation, antibiotic production, motility, sporulation, biofilm formation (eg, biofilm formation) have been reported (Fuqua et al ., Ann . Rev. Microbiol ., 2001, Vol. 50, pp. 725-751).

Especially, in the biofilm condition where the cell density is much higher than that of the floating phase, the mechanism of detecting the quorum of the microorganism can be easily and actively generated. In 1998, Davies et al. (Science, Vol. 280, pp. 295 ~ 298) reported that the mechanism of quantitative detection of microbial quorum is characterized by the physical structure characteristics, such as the extent, thickness and morphology of the biofilm formation of the pathogenic microorganism Pseudomonas aeruginosa . preventing contamination of medical devices in some areas, such as studies to inhibit biofilm formation through artificial control of the quorum sensing mechanism since the report represent a closely related with a variety of characteristics such as biofilm resistance recent medical and agriculture (Baveja et al ., Biomaterials , 2004, Vol. 50, pp. 5003-5012), plant diseases (Dong et al ., Nature , 2001, Vol. 411, pp. 813-817).

Conventional methods for controlling biofilm formation by controlling the microbial quorum sensing mechanism are classified into the following two types. First, it is possible to inhibit biofilm formation by injecting an antagonist, which has a similar structure to the signal molecule used in the quan- tity sensing mechanism and is competitive with the signal molecule and the gene expression site, and the most typical antagonist Delisea , a kind of algae Furanone and its halogenated derivatives secreted by pulchra have been reported (Henzer et < RTI ID = 0.0 > al ., EMBO Journal, Vol. 22, 3803-3815). Second, it inhibits biofilm formation by using an enzyme that decomposes signal molecules used in the quorum sensing mechanism (biofilm formation inhibiting enzymes such as a microbial quorum sensing inhibitory enzyme: for example, lactonease, acylase) or a microorganism that produces the enzyme can do. For example, Xu et al. (2004) developed a method to inhibit biofilm formation on various surfaces by injecting acylase solution, an enzyme that degrades acyl-homoserine lactone (AHL), a signal molecule of gram-negative bacteria, (U.S. Patent No. 6,777,223).

However, if the acylase enzyme solution is directly injected to inhibit the formation of biofilms, not only the loss of the enzyme is severe but also the inactivation is rapidly progressed due to the denaturation of the enzyme. have.

Recently, the acylase enzyme is immobilized on a magnetic carrier by a layer-by-layer to prevent deactivation due to degeneration of the enzyme, and to facilitate the separation and recovery of the enzyme-immobilized magnetic carrier using a magnetic field. The results of inhibiting biofilm contamination on the surface of a separator by applying to a submerged membrane bioreactor (sMBR) have been reported by Lee et al. (2009) (registered patent No. 981519). However, there is a high concentration of microbial flocs, and due to the nature of the MBR process, in which these flocs are continuously discharged to maintain sludge retention time, even if the carrier is a magnetic carrier, the carrier mixed in the floc is a magnetic field. There is a limit to recovering the whole quantity by bay. In addition, in order to maximize the recovery rate of the magnetic carrier, the carrier should be an immersion type reaction tank existing only in the reaction tank without circulating the inside of the system (eg, tubing, valve, fitting, etc.), and high pressure should be used, such as a nanofiltration process or a reverse osmosis membrane process. In the case of the separation membrane process is pointed out that the problem is difficult to apply because most of the external pressure type. Furthermore, in the case of the above-described Hyogo-immobilized magnetic carrier, the enzyme can be immobilized by culturing the microorganism and extracting and purifying the enzyme to produce the enzyme using a conventional microbial recombination technology, and thus the cost is high. It is also pointed out that immobilization of the purified enzyme through the lamination method also requires a lot of cost and time.

Accordingly, the present inventors solve the various problems arising from the above-described enzyme utilization technology by immobilizing a biofilm-forming inhibitory microorganism that produces the enzyme as described above instead of a biofilm-forming inhibitory enzyme to a carrier through a predetermined means and introducing the same into a water treatment reactor. In order to implement biofilm formation suppression technology in a membrane water treatment process economically and stably, and to anticipate physical membrane fouling mitigation effects at the molecular biological perspective, the research has been intensively studied.

U.S. Patent 6,777,223 Registered Patent No. 981519

Henzer et al., EMBO Journal, Vol. 22, 3803-3815

The present invention aims to solve and alleviate the membrane fouling phenomenon of biofilms by physical washing, in addition to the stable implementation of the mechanism of inhibiting biofilm formation from a molecular biological point of view based on the understanding of the mechanism of biofilm formation in a water treatment membrane process.

As a result of the research for achieving the above object, the present inventors use the biofilm formation inhibiting microorganism immobilized fluidized bed carrier having the fluidity of the biofilm formation inhibiting microorganisms immobilized therein and fluidity by aeration in the water, thereby promoting the activity of the biofilm formation inhibiting microorganisms. The present inventors have found that the membrane fouling mitigation effect can be simultaneously obtained by physical washing while maintaining the stability and reducing the membrane fouling of the membrane from the molecular biological point of view.

More specifically, the present invention provides a biofilm-forming inhibitory microorganism-immobilized fluidized bed carrier in which biofilm formation-inhibiting microorganisms are immobilized inside the carrier and have fluidity by aeration in water. As a result, biofilm formation can be suppressed molecularly by the biofilm formation inhibiting microorganism immobilized inside the carrier, and due to the fluidity of the carrier under the aeration condition in water, the biofilm formed on the surface of the membrane is detached by directly hitting the membrane surface. By inducing it is possible to induce detachment of the physical biofilm.

In the present invention, the fluidized bed carrier may include a hydrogel composed of a hydrophilic polymer as a main component, and more specifically, the hydrogel may be at least one selected from the group consisting of alginate, PVA, polyethylene glycol, and polyurethane (these And a complex of components), and the use of a fluidized bed carrier of such a material facilitates mass transfer through and out of the carrier and prevents damage to the surface of the immersion type membrane under aeration conditions.

Hydrogel in the present invention is preferably having a three-dimensional network structure through the internal chemical cross-linking in that the biofilm formation inhibitory microorganisms can be trapped between the chemical cross-linking to be able to continuously grow in the carrier.

For example, alginate is a carrier material containing a hydrophilic natural polymer as a main component, and forms solids in a network structure that minimizes resistance to mass transfer through crosslinking, which is a chemical bond in a calcium chloride solution. As a result, it is possible to immobilize not only the biofilm formation inhibiting microorganism but also the enzyme produced by the microorganism. The biocompatibility is excellent, so it is suitable for use in a reaction tank in which microorganisms for water treatment are present. It is also preferable in that it is harmless to a human body.

Furthermore, the fluidized bed carrier of the present invention may be substantially spherical or have a shape close to a spherical shape, thereby preventing damage to the surface of the immersion type membrane under water aeration conditions.

The biofilm formation inhibiting microorganism applicable to the present invention may be any kind of recombinant microorganism or natural microorganism as long as it is a microorganism of a kind capable of producing a biofilm formation inhibitory enzyme. In addition, microorganisms capable of producing quorum detection inhibitory enzymes that degrade signal molecules used in quorum detection mechanisms can be used, and preferably microorganisms that produce quorum detection inhibitory enzymes such as lactonase or acylase can be used. For example, Bacillus in E. coli XL1-blue, which is widely used for gene recombination. thuringiensis subsp. Escherichia coli recombinant aiiA gene extracted from kurstaki (gene related to lactonase production), or natural microorganisms in nature (eg, Rhodococcus qingshengii species strains) can be used. We believe that Rhodococcus is suitable for use in water treatment processes. In order to obtain strains of qingshengii species, sludges were collected from the bioreactors of urban sewage treatment plants, and microorganisms were isolated and identified, and then, using the enrichment culture method from the various microorganisms, Rhodoccocus microorganisms (Rhodococcus genus qingshengii, etc.).

In the present invention, the method of immobilizing the microorganisms to inhibit the biofilm formation in the carrier is not particularly limited as long as it is a method of immobilizing the microorganisms in the fluidized bed carrier, and it is not limited to adhesion, entrapment, encapsulation, etc. Microbial immobilization method can be used. For example, a highly concentrated microbial suspension in which biofilm formation inhibiting microorganisms are suspended in water can be mixed with a hydrogel and added dropwise to a calcium chloride solution at a constant flow rate to prepare a fluidized bed carrier of a constant size.

The present invention also provides a membrane water treatment apparatus including a water treatment reactor and a membrane module for water treatment in which a biofilm formation inhibiting microorganism immobilized fluidized bed carrier is disposed therein. The membrane module applicable to the membrane water treatment device of the present invention is not particularly limited as long as the membrane is formed on the surface of the membrane and thus the permeation performance of the membrane is deteriorated. Nanofiltration in addition to membrane bioreactor (MBR) devices in which biofilms are formed on the membrane surface by microorganisms for water treatment, as well as conventional membrane water treatment devices in which biofilms are formed on the membrane surface due to microorganisms often present in the water to be treated. Advanced water treatment apparatuses, such as an apparatus and a reverse osmosis filtration apparatus, are mentioned.

When the biofilm formation inhibiting microorganism-immobilized fluidized bed carrier of the present invention is applied to a membrane water treatment device, the biofilm formation suppression technique in terms of molecular biology, such as a quorum suppression mechanism, reduces the biofilm formation on the surface of the membrane while showing fluidity inside the reactor. The carrier physically strikes the surface of the separator and has the effect of detaching the biofilm attached to the surface. As a result, the membrane washing cycle is longer than the conventional membrane water treatment process and the consumption of cleaning chemicals is reduced, and the filtration process can be performed for a long time.

In addition, compared to the existing biofilm suppressant enzyme use and technology, there is no need for complicated and expensive processes such as enzyme extraction and immobilization, and only the biofilm forming inhibitory microorganisms need to be mixed when preparing a fluidized bed carrier using a pump. There is an advantage in that it can be easily recovered and reused by using the characteristic of sinking in a short time at the time of aeration even without a separate recovery device as in the conventional enzyme-immobilized magnetic carrier technology.

Figures 1a and 1b is a schematic diagram of the biofilm formation inhibiting microorganism immobilized fluid bed carrier of the present invention and is a photograph of the carrier actually produced.
2A and 2B are actual photographs of a bioreactor (FIG. 2A: no aeration; FIG. 2B: aeration) in which the biofilm formation inhibiting microorganism immobilized fluidized bed carrier of the present invention is added.
3 is a view showing an embodiment of a method for producing a biofilm formation inhibiting microorganism immobilized fluid bed carrier.
4 is a diagram showing the signal molecule decomposition activity of biofilm formation inhibiting microorganism immobilized fluid bed carrier.
FIG. 5 is a schematic diagram of a membrane bioreactor apparatus in which a biofilm formation inhibiting microorganism-immobilized fluidized bed carrier of the present invention is introduced into a bioreactor and operated.
6 is a view showing the degree of increase (membrane contamination) of the interlayer differential pressure according to the operating time in the membrane bioreactor apparatus of the Examples and Comparative Examples of the present invention.
FIG. 7 is a diagram showing signal molecule decomposition activity (relative activity) of biofilm formation inhibiting microorganism-immobilized fluidized bed carrier according to operating time of the membrane bioreactor according to the present invention.
8 is a view showing the wet weight of the biofilm formation inhibiting microorganism immobilized fluid bed carrier according to the operating time of the membrane bioreactor device of the present invention.

Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not limited thereto.

Manufacturing example  One  - Biofilm  Immobilized microorganisms Fluid phase Carrier  Manufacture and measurement of signal molecule degradation activity

Biofilm formation inhibitory microorganisms, Rhodoccocus qingshengii , which is known to produce lactonase , a kind of quorum-sensing inhibitory enzyme, was used as a microorganism suitable for the actual water treatment process. Rhodoccocus qingshengii collected sludge from the Okcheon Sewage Treatment Plant in Chungcheongbuk-do to isolate the biofilm suppressant microorganisms. Among the many microorganisms isolated, bioassay and enriched cultures were used to find the signal molecule-degrading microorganisms, and the microorganisms of the genus Rhodoccocus with excellent signal molecule-degrading activity were isolated.

Sodium alginate, a natural polymer produced by Sigma co., Was used as a fluidized bed carrier to immobilize the microorganisms to inhibit biofilm formation.Alginate is a representative material used for microbial encapsulation. . In order to maintain physical strength for a long time in the membrane biofilm reactor, preliminary tests were performed on various concentrations of alginate, and in the preparation of the present invention, the concentration of the alginate solution was 4 wt% based on the concentration of the final injection. Adjusted.

Rhodoccocus qingshengii is grown for 24 hours by shacked culture, and then centrifuged with 200 ml of shake culture to remove the culture components of the supernatant and remaining Rhodoccocus. Aggregates of qingshengii were washed with Tris-HCl 50 mM buffer (pH 7.0) and resuspended in ultrapure water. Thereafter, as shown in FIG. 3, the resuspension solution of the biofilm formation inhibiting microorganism and the alginate solution are mixed and sprayed onto a calcium chloride (CaCl ₂ ) solution, whereby a good cross-linking is carried out. Was made of a fluidized bed carrier. When preparing the fluidized bed carrier, the concentration of alginate was 4 wt% in the final spraying, crosslinked in 2 wt% calcium chloride (CaCl ₂ ) solution for 1 hour, and dried at room temperature for 20 hours to increase physical strength. .

Signal molecule (AHL) degradation activity of the biofilm formation inhibiting microorganism immobilized fluidized bed carrier prepared as described above was measured using N-octanoyl-L-homoserine lactone (OHL), one of the representative signal molecules found in the water treatment process. Put 30 mL of Tris-HCl 50mM buffer solution (pH 7.0) into the test tube, inject OHL to a concentration of 0.2 mm, and inhibit biofilm formation microorganisms ( Rhodoccocus qingshengii ) was added to the fixed fluidized bed carrier in a shaking incubator at 30 degrees temperature and reacted at 200rpm for 60 minutes. This confirmed that about 92% of the signal molecule was degraded for 60 minutes by the biofilm formation inhibitory enzyme (lactonase) produced from the biofilm formation inhibitory microorganism (FIG. 4).

Example  One Separator Bioreactor  Application to the device

The biofilm formation inhibiting microorganism immobilized fluidized bed carrier prepared by the above-mentioned method was applied to a laboratory scale membrane bioreactor apparatus. (See FIG. 5). Specifically, 1.6 L of activated sludge was filled in a cylindrical reactor, and an acidic stone was installed at the lower end to maintain an aeration of 1 L / min. A total of 60 microbial-immobilized microbial immobilized fluidized bed carriers were added into the reactor. It was. For the continuous process, synthetic wastewater using glucose as the main carbon source was introduced through the inlet pump. The chemical oxygen demand (COD) of the synthetic wastewater was about 560 ppm and the hydraulic residence time was operated at 5.3 hours (hr). The membrane for filtration installed in the reactor was an ultrafiltration membrane (Zeeweed 500, manufactured by GE-Zenon, pore size 0.04 mm) as an immersion hollow fiber membrane, and the flux of treated water passing through the membrane was operated at 28.7 L / m ² · hr. It was. In addition, some of the treated water was returned to the reactor through a level controller and a 3-way valve to maintain the reactor level. As the operation proceeds, a biofilm is formed on the surface of the membrane, which leads to a decrease in membrane permeability due to an increase in membrane contamination. The degree of biofilm contamination is expressed as a transmembrane pressure (TMP) value, and the interlayer differential pressure increases. The higher the degree of biofilm contamination is shown. As a result of the experiment, the transmembrane pressure was only 5 kPa after 77 hours of operation, and the intermembrane pressure reached 70 kPa only after 400 hours of operation (see FIG. 6).

Comparative example  One

Instead of biofilm formation inhibiting microorganism-immobilized fluidized bed carrier in Example 1, 60 hydrogel fluidized bed carriers (carrier prepared without immobilizing biofilm-forming inhibitory microorganisms in Production Example 1) were introduced into the reaction tank. Operation was carried out in the same manner as in Example 1 except for the above. As a result, the interlude differential pressure reached 70 kPa after 77 hours of operation (see FIG. 6).

Comparative example  2

Example 1 was operated in the same manner as in Example 1 except that the biofilm formation inhibiting microorganism immobilized fluidized bed carrier was not added to the membrane bioreactor. As a result, the interlude differential pressure reached 70 kPa after 43 hours of operation (see Fig. 6).

That is, in Example 1 and Comparative Examples 1 to 2, in the case of the membrane bioreactor device in which the biofilm formation inhibiting microorganism-immobilized fluidized bed carrier of the present invention is added (Example 1), a fluidized bed carrier in which microorganisms are not immobilized is added. Compared to the case (Comparative Example 1) and the case in which no fluidized bed carrier is added (Comparative Example 2), it can be seen that the membrane fouling phenomenon caused by the biofilm on the surface of the separator is remarkably alleviated, which is stably immobilized inside the fluidized bed carrier. In addition to the molecular biological effects by controlling the mechanism of inhibiting biofilm formation of the microorganisms, the membrane fouling removal effect by physical washing of the surface of the membrane by the carrier having fluidity is estimated to be synergistic.

Example  2 : Biofilm  Formation Inhibition Microbial Immobilized Fluidized Bed Carrier  Keep active

An experiment was conducted to determine whether the signal molecule decomposition activity of the biofilm formation inhibiting microorganism in the biofilm formation inhibiting microorganism immobilized fluidized bed carrier of the present invention was maintained for a long time. Specifically, in Example 1, 0, 1, 3, 5, 7, 10, 13, 15, 17, 20, 23, 25, 27 on the basis of the time when the biofilm formation inhibiting microorganism immobilized fluidized bed carrier is added to the reaction tank. On the 30th day, the biofilm formation inhibiting microorganism-immobilized fluidized bed carrier was taken out, and the outside of the fluidized bed carrier was washed several times with distilled water, and then the same experiment as in Preparation Example 1 was performed. The relative activity was measured based on the activity (100%) of the result of the biofilm formation inhibiting microorganism-immobilized fluidized bed carrier on day 0. As a result, it was confirmed that even after the operation for 20 days or more, the biofilm formation inhibiting microorganism immobilized fluidized bed carrier did not decrease the signal molecule degradation activity, but rather increased slightly compared to the initial (0 day) activity (FIG. 7).

Example  3 : Fluidized bed carrier  undergarment Biofilm  Confirmation of growth of microorganisms inhibiting formation

When the biofilm formation inhibiting microorganism-immobilized fluidized bed carrier of the present invention was operated in a long-term separation membrane bioreactor, a confirmation experiment was performed on the extent of growth of the biofilm formation inhibiting microorganism.

Specifically, in Example 1, while operating for 25 days from the time when the biofilm formation-suppressing microorganism immobilized fluidized bed carrier was put into the reactor, 10 fluidized bed carriers on which biofilm formation-inhibiting microorganisms were immobilized every 24 hours were taken out and flowed into distilled water several times. After the outside of the phase carrier was washed off, the wet weight (average of five replicates) was measured. As a result, after 25 days, the wet weight of the biofilm-forming microorganisms was gradually increased (FIG. 12).

Comparative example  3

Except for using the alginate fluidized bed carrier that did not immobilize the biofilm formation inhibiting microorganisms in Example 3, the experiment was carried out in the same manner as in Example 3, it was confirmed that there is little change in the wet weight (Fig. 8).

That is, from Examples 2 to 3 and Comparative Example 3, as a result of growth of the biofilm formation inhibiting microorganisms enclosed inside the biofilm formation inhibiting microorganism immobilized fluidized bed carrier of the present invention, the wet weight increases, thereby decomposing signal molecules. It can be seen that the activity does not decrease but increases slightly.

Claims (9)

A biofilm formation inhibiting microorganism immobilized fluidized bed carrier in which biofilm formation inhibiting microorganisms are immobilized therein and have fluidity due to aeration in water. The method according to claim 1,
The fluidized bed carrier includes a biogel to inhibit biofilm formation. The method according to claim 1,
The fluidized bed carrier comprises at least one member selected from the group consisting of alginate, PVA, polyethylene glycol, and polyurethane. The method according to claim 1,
The fluidized bed carrier has a three-dimensional network structure through the chemical cross-linking therein biofilm formation inhibiting microorganism immobilized fluidized bed carrier. The method according to claim 1,
The fluidized bed carrier is microbial immobilized microorganism immobilized fluid bed carrier, characterized in that consisting of a sphere. The method according to claim 1,
The biofilm formation inhibiting microorganism is a biofilm formation inhibiting microorganism immobilized fluidized bed carrier, characterized in that the recombinant microorganism or a natural microorganism capable of producing a biofilm formation inhibitory enzyme. The method according to claim 1,
The biofilm formation inhibiting microorganism is capable of producing a quorum detection inhibitory enzyme biofilm formation inhibiting microorganism immobilized fluid bed carrier. The method of claim 7,
The quorum-sensing inhibitory enzyme is lactonase or acylase. Separation water treatment apparatus comprising a reaction vessel for receiving the water to be treated, and the biofilm formation inhibiting microorganism immobilized fluid bed carrier of any one of claims 1 to 8 disposed inside the reactor and the membrane module for water treatment.

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