Microcapsule material capable of reducing pollution containing polycyclic aromatic hydrocarbon, and preparation method and application thereof

Abstract

The present invention discloses a microcapsule material capable of reducing pollution containing polycyclic aromatic hydrocarbons, and a preparation method and application thereof. The preparation method comprises the following steps: (1) culturing mycobacterium gilvum CP 13 in a bacteria culture liquid to obtain a bacterial broth, wherein the mycobacterium gilvum CP 13 was deposited in China General Microbiological Culture Collection Center (CGMCC) on Jul. 22, 2013 with a CGMCC number of CGMCC No. 7963; and (2) applying a calcium alginate and chitosan to encapsulate the bacterial broth in a microcapsule through layer-by-layer self-assembly to produce a microcapsule material capable of reducing pollution containing polycyclic aromatic hydrocarbons. The present invention produces a microcapsule material with the microorganic activity through layer-by-layer self-assembly, which has superior adaptability to the environment and good ability to reduce pollution containing polycyclic aromatic hydrocarbons. The microcapsule material of the present invention can be used in bioremediation of industrial wastewater containing polycyclic aromatic hydrocarbons and contaminated soil containing polycyclic aromatic hydrocarbons.

Claims

1. A method for preparing a microcapsule material capable of reducing pollution containing polycyclic aromatic hydrocarbons, the method comprising: (1) culturing mycobacterium gilvum CP 13 in a bacteria culture liquid to obtain a bacterial broth; and (2) encapsulating the bacterial broth in the microcapsules by applying a calcium alginate and chitosan to the bacterial broth through layer-by-layer self-assembly to produce a microcapsule material capable of reducing pollution containing polycyclic aromatic hydrocarbons.

2. The method according to claim 1, wherein the bacteria culture liquid is nutrient broth medium, and wherein the bacteria culture liquid further comprises beef extract 3 g/L, peptone 10 g/L, NaCl 5 g/L, agar 20 g/L, and has a pH in the range of 7.0-7.2.

3. The method according to claim 1, wherein the step (2) of encapsulating the bacterial broth in the microcapsule comprise adding chitosan into the bacterial broth so that chitosan is deposited on a surface of bacteria, adding the calcium alginate into the bacterial broth so that the calcium alginate is deposited on a surface of chitosan; and repeating these steps 2-3 times, thereby encapsulating the bacterial broth in the microcapsules.

4. The method according to claim 3, wherein concentrations of chitosan and the calcium alginate are each in a range of 0.1-2.0 g/L.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1. Effects of different initial concentrations of pyrene on the degradation of pyrene by the microcapsule material of the present invention. (a) 10 mg/L, (b) 30 mg/L, and (c) 50 mg/L. Treatment: () a blank of free bacteria, (.circle-solid.) a blank of the microcapsule material of the present invention, () free bacteria, (.box-tangle-solidup.) the microcapsule material of the present invention, () free bacteria, (.square-solid.) the microcapsule material of the present invention, () free bacteria, and (.diamond-solid.) the microcapsule material of the present invention.

(2) FIG. 2. Effects of different pH values (a) and different temperatures (b) on the degradation of pyrene by the microcapsule material of the present invention. Initial concentration of pyrene=10 mg/L. Treatment: (a) () free bacteria-pH 7, (.box-tangle-solidup.) the microcapsule material of the present invention-pH 7, () free bacteria-pH 3, (.square-solid.) the microcapsule material of the present invention-pH 3, () free bacteria-pH 10, (.diamond-solid.) the microcapsule material of the present invention-pH 10; (b) () free bacteria-30 C., (.box-tangle-solidup.) the microcapsule material of the present invention-30 C., () free bacteria-10 C., (.square-solid.) the microcapsule material of the present invention-10 C., () free bacteria-40 C., and (.diamond-solid.) the microcapsule material of the present invention-40 C.

(3) FIG. 3. Changes of the removal rate of pyrene in soil in the process of remediation of pyrene contaminated soil by the microcapsule material of the present invention. The initial concentration of pyrene=120 mg/kg. Treatment: () free bacteria, and () the microcapsule material of the present invention.

DETAILED DESCRIPTION OF THE EMBODIMENTS

(4) The present invention will be further described below in detail with reference to specific examples; however, the embodiments of the present invention are not limited thereto, and process parameters not specifically given can be determined by referring to conventional techniques.

EXAMPLE 1

(5) Preparation of the microcapsule material of the present invention capable of degrading polycyclic aromatic hydrocarbons

(6) A method for culturing mycobacterium gilvum CP13 strain of the present invention is as follows:

(7) Respectively performing streak inoculation of the CP13 strains in beef juice peptone medium, and culturing at 28 C.-30 C. for 48 h; then respectively inoculating them into a 500 mL flask, and culturing at 30 C. and 150 r/min for 12 h; and finally inoculating them into a 5 L seed tank in accordance with an inoculum concentration of 5%, and culturing at 180 r/min, pH 7.5 and a ventilatory capacity of 5 L/min for 24 h.

(8) Composition and dosage of the nutrient broth medium are as follows (g/L): Beef extract 3, peptone 10, NaCl 5, agar 20, and a pH adjusted to 7.0-7.2.

(9) The specific steps are as follows: (1) Culturing mycobacterium gilvum CP 13 in a bacteria culture liquid to obtain a bacterial broth, (2) adding 20 ml of 0.2 g/L polycation (chitosan) to the bacterial broth so that polycation is deposited on the surface of bacteria, and then adding the same volume and concentration of polyanion (calcium alginate) solution so that polyanion is further deposited on the surface of the chitosan. Repeating the operations above 2-3 times so that the bacterial broth can be encapsulated in the microcapsule. The bacteria-immobilized microcapsules are thus prepared. They are sealed in sterile bags and stored in a refrigerator at 4 C.

EXAMPLE 2

(10) Inoculating the microcapsule material prepared in Example 1 at a ratio of 0.1 g/20 ml to a MSM culture solution with the concentration of pyrene at 10 mg/L; taking a pyrene-containing MSM medium without the microcapsule material as a blank and placing it in a shaker at 30 C., culturing in darkness while shaking at 150 r/min, and then determining the residual amount of pyrene in the culture solution. The formulation of the MSM culture solution: 100 mg/L (NH.sub.4).sub.2SO.sub.4, 20 mg/L MgSO.sub.4.Math.7H.sub.2O, 10 mg/L CaCl.sub.2.Math.2H.sub.2O; trace elements: 1.2 mg/L FeSO.sub.4.Math.7H.sub.2O, 0.3 mg/L MnSO.sub.4.Math.H.sub.2O, 0.3 mg/L ZnSO.sub.4.Math.7H.sub.2O, 0.1 mg/L CoSO.sub.4.Math.7H.sub.2O, 0.1 mg/L (NH.sub.4).sub.6Mo.sub.7O.sub.24.Math.4H.sub.2O; phosphate buffer solution: 2.5 g/L K.sub.2HPO.sub.3, 0.77 g/L KH.sub.2PO.sub.3, pH 7.2-7.4.

(11) As can be seen from FIG. 1, in the experimental concentration range, the efficiency of removal of PYR by the microcapsule material is higher. At the initial pyrene concentrations of 10 mg/L, 30 mg/L and 50 mg/L, respectively, the 3d removal rates are increased from 60%, 40%, 20% by the free CP13 bacteria to 95%, 65%, 55% by the microcapsule material, respectively. In the case of high concentration (such as 50 mg/L), the microcapsule material is more advantageous.

EXAMPLE 3

(12) Inoculating the microcapsule material prepared in Example 1 at a ratio of 0.1 g/20 ml to a MSM culture solution with the concentration of pyrene at 10 mg/L; taking a pyrene-containing MSM medium without the microcapsule material as a blank and free bacteria as a control and placing them in a shaker at 30 C., culturing in darkness while shaking at 150 r/min, and then determining the residual amount of pyrene in the culture solution. As can be seen from FIG. 2, the microcapsule material of the present invention can effectively degrade polycyclic aromatic hydrocarbon pyrene under the conditions of pH 3-10, Especially in the acidic case of pH 3, the removal rate of free bacteria is only 15%, while the removal rate of the microcapsule material of the present invention can still be maintained at 95% or more. As can be seen, the microcapsule material of the present invention has a broader range of pH adaptation than free bacteria. Besides, the microcapsule material of the present invention can effectively degrade polycyclic aromatic hydrocarbon pyrene at a temperature of 10 C.-40 C. Similarly, in the case of ambient temperatures as low as 10 C., the 4d removal rate by the free bacteria is only about 17%, while the 4d removal rate by the microcapsule material of the present invention can reach 80%. At 40 C., the 4d removal rate of the microcapsule material of the present invention is maintained at 80%, much higher than 40% by the free bacteria within the same duration. It can be seen that the microcapsule material of the present invention has better thermal stability than the free bacteria.

(13) This example shows that the microcapsule material of the present invention capable of degrading polycyclic aromatic hydrocarbons can degrade pyrene well in a wide pH range of acidic, neutral and alkaline conditions and at 10 C.-40 C., ensuring its application at different pHs and in low and high temperature environments.

EXAMPLE 4

(14) Experiment on remediation of pyrene contaminated soil by the microcapsule material of the present invention

(15) Adding 400 g of not sterile soil, to which pyrene has been added at 120 mg of pyrene per kilogram of soil, to a 500 ml beaker, and adding 2.0 g of the microcapsule material of Example 1 to the soil so that the initial inoculum is 10.sup.7 CFU per gram of soil. Using free CP13 as a control at the same amount of 10.sup.7 CFU per gram of soil, and incubating in an incubator at 30 C. Sampling to determine the residual amount of pyrene in the soil after culturing for 6 d, 10 d, 20 d, 40 d.

(16) As indicated by the results in FIG. 3, after using the microcapsule material of the present invention to treat the soil containing pyrene at 120 mg of pyrene per kilogram of soil for 40 days, the removal rate of pyrene can reach 81%, while here the degradation rate of the control using free bacteria is only 42%.