JP2012219260A - Method for producing chitin from crustacean shell of crustacea or shell of shellfish using ionic liquid - Google Patents

Abstract

［式中、
R¹は、C₁〜C₄アルキルであり、
R²は、C₁〜C₄アルキル又はC₂〜C₄アルケニルであり、
Xは、ハロゲンである。］
で表されるイオン液体とを接触させてキチンを抽出するキチン抽出工程；及び
キチン抽出工程で得られたキチンを含有する抽出物とキレート剤とを接触させて、該抽出物から無機塩を除去する無機塩除去工程；
を含む、キチンの製造方法。
【選択図】なしProvided is a method for producing chitin, which can produce high-purity chitin from a crustacean shell or a shell of shellfish by a simple process.
A shell or shell containing chitin and an inorganic salt and formula I:

[Where:
R ¹ is C ₁ -C ₄ alkyl;
R ² is C ₁ -C ₄ alkyl or C ₂ -C ₄ alkenyl,
X is a halogen. ]
A chitin extraction step for extracting chitin by contacting with the ionic liquid represented by the formula; and an extract containing the chitin obtained in the chitin extraction step and a chelating agent are contacted to remove inorganic salts from the extract Removing inorganic salt;
A method for producing chitin, comprising:
[Selection figure] None

Description

  The present invention relates to a method for producing chitin from a crustacean shell or shellfish shell using an ionic liquid.

Chitin is a natural glucan having a structure in which N-acetyl-D-glucosamine is polymerized by β-1,4-glycosidic bonds. The average molecular weight of naturally occurring chitin is said to be about 10 ⁵ to 10 ⁶ .

  N-acetyl-D-glucosamine, a hydrolyzate of chitin, is commercially available as a health food. In addition, chitosan, which is a deacetylated product of chitin, is a cationic polysaccharide having a primary amino group. Therefore, various functional materials such as metal adsorbents are used as functional materials using amino groups and hydroxyl groups of sugar chain skeletons. Applications have been proposed (Patent Document 1). For this reason, the demand for chitin as a raw material for N-acetyl-D-glucosamine and chitosan is also increasing.

  Chitin is a constituent of the cell walls of arthropod shells, especially crustacean shells, molluscs, particularly shellfish shells, and fungal (mushrooms) and basidiomycetes (mushrooms). is there. For this reason, chitin is produced exclusively by isolation from these biological materials. On the other hand, crustacean shells, shellfish shells, and the like are also biomass wastes that are discharged in large quantities in the fishery or food processing industries. At the present time when wastes cannot be dumped into the ocean due to increasing awareness of environmental protection, the development of biomass waste treatment technology and effective utilization technology as described above is an urgent issue.

Crustacean shells and shellfish shells contain about 9 to 17% chitin, and also contain large amounts of protein and inorganic salts such as calcium carbonate (CaCO ₃ ). For this reason, in order to produce chitin from the crustacean shell or shellfish shell, it is necessary to purify and isolate the chitin from these impurities. As a method for producing chitin from the shell, a method is generally employed in which the shell is treated with hydrochloric acid to remove inorganic salts, and then the protein is hydrolyzed with an aqueous sodium hydroxide solution (Non-patent Document 1). However, this method has a problem that the process is complicated and a hydrolysis and deacetylation of β-1,4-glycoside bond proceeds because of the use of a strong acid and a strong base.

  The present inventor has found that 1-allyl-3-methylimidazolium bromide (AMIMBr), which is a kind of ionic liquid, has excellent solubility in chitin (Non-patent Document 2). An ionic liquid is a salt that is liquid at room temperature composed of an anion and a cation, and has a characteristic of extremely low vapor pressure. Patent documents 2 to 4 describe a method for producing chitin from a crustacean shell using a solvent containing an ionic liquid of imidazolium halide. Non-Patent Document 3 describes a method for producing chitin from crustacean shells using 1-ethyl-3-methylimidazolium acetate as an ionic liquid.

JP 2004-255302 A JP 2008-248217 A JP 2009-179913 JP JP 2010-104768 A

Chitin and Chitosan Study Group, Chitin and Chitosan Handbook, 1995, p. 1-30 Prasad, K. et al., Int. J. Biol. Macromol., 2009, 45, p. 181-189 Qin, Y. et al., Green Chem., 2010, Vol. 12, p. 968-971

When producing chitin from crustacean shells or shells of shellfish, the method using hydrochloric acid and sodium hydroxide is complicated, and the use of strong acids and strong bases results in hydrolysis of β-1,4-glycosidic bonds. And the problem that deacetylation proceeds. On the other hand, since the ionic liquids developed by the present inventors (Non-patent Document 2 and Patent Documents 2 to 4) dissolve chitin under mild conditions, the progress of hydrolysis and deacetylation can be suppressed. However, extraction of chitin from crustacean shells or shellfish shells using this ionic liquid is expected to contain a large amount of inorganic salts such as CaCO ₃ contained in the shell and shells, and a method for producing high-purity chitin Was not established. Further, in the method described in Non-Patent Document 3, the crustacean shell is limited to shrimp shell, and the ionic liquid used is the imidazolium halide described in Non-Patent Document 2 and Patent Documents 2 to 4. In comparison, there was a problem of being a special compound.

  Therefore, an object of the present invention is to provide a method for producing chitin, which can produce high-purity chitin from a crustacean shell or a shellfish shell by a simple process.

As a result of various studies on means for solving the above-mentioned problems, the present inventor extracted the chitin from the shell or shell using an ionic liquid, and then treated with a chelating agent to thereby contain CaCO contained in the chitin extract. _The present inventors have found that high-purity chitin can be produced by efficiently removing inorganic salts such as ₃ etc.

［式中、
R¹は、C₁〜C₄アルキルであり、
R²は、C₁〜C₄アルキル又はC₂〜C₄アルケニルであり、
Xは、ハロゲンである。］
で表されるイオン液体とを接触させてキチンを抽出するキチン抽出工程；及び
キチン抽出工程で得られたキチンを含有する抽出物とキレート剤とを接触させて、該抽出物から無機塩を除去する無機塩除去工程；
を含む、キチンの製造方法。
（2） キレート剤が、クエン酸、エチレンジアミン四酢酸及びニトリロ三酢酸から選択される、前記（1）の方法。
（3） キチン抽出工程において、80〜120℃の範囲の温度で6〜48時間キチンを抽出する、前記（1）又は（2）の方法。 That is, the gist of the present invention is as follows.
(1) Shell or shell containing chitin and inorganic salt and formula I:

[Where:
R ¹ is C ₁ -C ₄ alkyl;
R ² is C ₁ -C ₄ alkyl or C ₂ -C ₄ alkenyl,
X is a halogen. ]
A chitin extraction step for extracting chitin by contacting with the ionic liquid represented by the formula; and an extract containing the chitin obtained in the chitin extraction step and a chelating agent are contacted to remove inorganic salts from the extract Removing inorganic salt;
A method for producing chitin, comprising:
(2) The method according to (1) above, wherein the chelating agent is selected from citric acid, ethylenediaminetetraacetic acid and nitrilotriacetic acid.
(3) The method according to (1) or (2) above, wherein in the chitin extraction step, chitin is extracted for 6 to 48 hours at a temperature in the range of 80 to 120 ° C.

  According to the present invention, it is possible to provide a method for producing chitin that can produce high-purity chitin from a crustacean shell or a shellfish shell by a simple process.

It is a figure which shows the X-ray powder diffraction (XRD) spectrum of the chitin obtained by the method of this invention. A: XRD spectrum of sample 1; B: XRD spectrum of sample 2; C: XRD spectrum of commercially available chitin. It is a figure which shows the infrared absorption (IR) spectrum of the chitin obtained by the method of this invention. A: IR spectrum of sample 2; B: IR spectrum of commercially available chitin.

  The present invention relates to a method for producing chitin. Hereinafter, preferred embodiments of the present invention will be described in detail.

<1. Chitin extraction process>
The method of the present invention includes a chitin extraction step of extracting chitin by bringing a shell or shell containing chitin and an inorganic salt into contact with an ionic liquid.

In the present specification, “crustacea” means an arthropod having a crust, and “crustacea” means a shell-like outer skin. In the present specification, “shellfish” means a mollusk having a shell, and “shell” means a shell-like organ that protects the animal body formed around the animal body (soft body part). . The crust used in this step is preferably a crustacean crust selected from the group consisting of shrimp and crab. The shell used in this step is preferably a scallop shell. The above-mentioned shells and shells contain chitin, protein and inorganic salts such as calcium carbonate (CaCO ₃ ) as main components. Therefore, by using the above shell or shell as a chitin source, it becomes possible to produce chitin with high yield.

で表されるハロゲン化イミダゾリウムであることが好ましい。 In the present specification, the “ionic liquid” means a salt that is liquid at room temperature and is composed of an anion and a cation. An ionic liquid has a feature that it is nonvolatile because its vapor pressure is extremely low. The present inventor has found that a specific ionic liquid can dissolve poorly soluble polysaccharides such as cellulose and chitin (Patent Documents 2 to 4, Non-Patent Document 2). Therefore, the ionic liquid used in this step is of formula I:

It is preferable that it is imidazolium halide represented by these.

As used herein, “alkyl” means a straight or branched chain aliphatic hydrocarbon group containing the specified number of carbon atoms. For example, “C ₁ -C ₄ alkyl” means a straight or branched hydrocarbon chain containing at least 1 and at most 4 carbon atoms. Suitable alkyls include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl and tert-butyl.

  In the present specification, “alkenyl” means a group in which one or more C—C single bonds of the alkyl are substituted with double bonds. Suitable alkenyls include, but are not limited to, vinyl, 1-propenyl, allyl, 1-methylethenyl (isopropenyl), 1-butenyl, 2-butenyl, 3-butenyl, 1-methyl-2-propenyl, 2 Mention may be made of -methyl-2-propenyl, 1-methyl-1-propenyl and 2-methyl-1-propenyl.

In the compound represented by Formula I, R ¹ is preferably selected from the group consisting of C ₁ -C ₄ alkyl and C ₂ -C ₄ alkenyl, more preferably C ₁ -C ₄ alkyl, Particularly preferred is methyl or ethyl.

In the compound represented by formula I, R ² is preferably selected from the group consisting of C ₁ -C ₄ alkyl and C ₂ -C ₄ alkenyl, and more preferably C ₁ -C ₄ alkyl or allyl. Preferably, it is allyl.

  In the compounds of the formula I, X means halogen. X is preferably selected from the group consisting of fluorine, chlorine, bromine and iodine, more preferably bromine.

In the compounds of the formula I, it is particularly preferred that R ¹ is C ₁ -C ₄ alkyl, R ² is C ₁ -C ₄ alkyl or C ₂ -C ₄ alkenyl and X is halogen.

  Particularly preferably, the compound of the formula I is 1-allyl-3-methylimidazolium bromide (AMIMBr) or 1-ethyl-3-methylimidazolium bromide.

  In this step, the ionic liquid may be used alone or in the form of an ionic liquid mixture containing two or more ionic liquids, and one or more ionic liquids and one or more solvents are used. You may use in the form of the solvent mixture to contain. Any form is included in the “ionic liquid” in this step. Here, the solvent that can form the solvent mixture is not limited, but for example, methanol, ethanol, acetonitrile, dimethyl sulfoxide, N, N-dimethylacetamide, N, N, -dimethylformamide and 1-methyl-2 -Pyrrolidone can be mentioned. However, the ionic liquid used in this step is preferably used in a single form. By using the ionic liquid, chitin contained in the shell and shell can be efficiently dissolved.

  In this step, the weight ratio between the shell or shell and the ionic liquid is preferably in the range of 1: 2 to 1: 100, more preferably 1: 5 to 1:20. By bringing the shell or shell and the ionic liquid into contact with each other at the above weight ratio, the inorganic salt can be removed from the extract.

  In this step, the temperature for extracting chitin is preferably in the range of 80 to 120 ° C, and more preferably in the range of 80 to 100 ° C. The time for extracting chitin is preferably in the range of 6 to 48 hours, and more preferably in the range of 6 to 24 hours.

  By carrying out this step under the above conditions, chitin can be efficiently extracted from the shell or shell.

<2. Inorganic salt removal process>
The method of the present invention includes an inorganic salt removing step of contacting an extract containing chitin obtained in the chitin extraction step with a chelating agent to remove inorganic salts from the extract. The extract obtained by the chitin extraction process contains not only chitin but also inorganic salts contained in large amounts in the shell and shell. Here, examples of the inorganic salt that can be contained in the extract include metal ion salts such as calcium carbonate (CaCO ₃ ). Since the above inorganic salts are generally insoluble, they usually behave together with chitin by conventional separation means such as centrifugation and filtration and cannot be easily separated. In addition, when an inorganic salt is solubilized by a strong acid treatment, hydrolysis and deacetylation of a β-1,4-glycoside bond proceeds, leading to a decrease in chitin yield.

  When the present inventors add an extract containing chitin obtained in the chitin extraction step to an aqueous citric acid solution that is a chelating agent, metal ions are chelated and solubilized in the form of complex ions. It was found that it can be easily separated. The inventor has also found that chitin obtained by this method has a low protein content as well as inorganic salts. Therefore, high-purity chitin can be produced by carrying out this step.

The chelating agent used in this step is not particularly limited, and a normal chelating agent capable of chelating metal ions such as Ca ²⁺ can be used. Specifically, the chelating agent used in this step is preferably citric acid, ethylenediaminetetraacetic acid or nitrilotriacetic acid, and more preferably citric acid. Citric acid is an inexpensive compound that is also commonly used as a food additive, and is particularly preferred because it selectively chelates with Ca ^2+, which is particularly abundant in shells and shells.

  This step is preferably carried out by bringing the extract containing chitin obtained in the chitin extraction step into contact with the chelating agent in water or a mixed solvent of water and a water-miscible solvent. The water-miscible solvent used in this step is preferably methanol, ethanol, acetonitrile, dimethyl sulfoxide, N, N-dimethylacetamide, N, N, -dimethylformamide or 1-methyl-2-pyrrolidone. The mixing ratio of water and the water-miscible solvent is preferably in the range of 10: 1 to 1000: 1, and more preferably 100: 1 to 1000: 1. This step is particularly preferably carried out by bringing an extract containing chitin obtained in the chitin extraction step into contact with a chelating agent in water. By bringing the extract and the chelating agent into contact with each other in water or the above mixed solvent, metal ions derived from inorganic salts can be solubilized in water or the mixed solvent and separated from chitin.

  In this step, the weight ratio of the extract to the chelating agent is preferably in the range of 1: 1 to 1:50, preferably 1: 5 to 1:10, based on the total weight of the extract. More preferred. Moreover, when based on the weight of chitin contained in the extract, the range is preferably 1:10 to 1: 500, more preferably 1:50 to 1: 100. By bringing the chitin extract and the chelating agent into contact with each other at the above weight ratio, high-purity chitin can be obtained.

  In this step, the temperature at which the extract is brought into contact with the chelating agent is preferably in the range of 10 to 50 ° C, more preferably in the range of 20 to 30 ° C. The time for contacting the extract with the chelating agent is preferably in the range of 1 to 24 hours, and more preferably in the range of 3 to 10 hours.

The chitin obtained by contacting the extract with a chelating agent can be separated from the reaction mixture by conventional means such as filtration or centrifugation, and washed with water if desired.
By carrying out this step under the above conditions, high-purity chitin can be produced.

  Hereinafter, the present invention will be described in more detail with reference to Examples and Comparative Examples.

<Example 1>
[Direct extraction and purification of chitin]
As crab shells, dried crab shells (manufactured by Tamagoya Co., Ltd.) marketed for fertilizers were used. The crab shell was boiled with water and then screened to obtain a small piece of crab shell of about 3 to 5 mm square. A small piece of 2.0 g of crab shell was added to 20.0 g of 1-allyl-3-methylimidazolium bromide (AMIMBr) and heated and stirred at 100 ° C. for 24 hours to extract chitin. Thereafter, the obtained reaction solution was centrifuged to obtain a supernatant containing crude chitin (Sample 1-1). This supernatant was added to 500 ml of 1.5 wt / vol% citric acid aqueous solution and stirred at room temperature for 3 hours to reprecipitate chitin and remove inorganic salts (decalcification). The resulting precipitate was filtered off, washed with water, and then dried to obtain 0.22 g of chitin (sample 1-2) (yield (weight ratio): about 11%).

[Identification of chitin]
Samples 1-1 and 1-2 obtained were analyzed by X-ray powder diffraction (XRD). The results are shown in Figure 1.
As shown in FIG. 1, in the XRD spectrum (A) of Sample 1-1, although a diffraction peak derived from chitin (spectrum C) was slightly observed, a diffraction peak of 2θ = 30 ° or more derived from CaCO ₃ Was observed more strongly. From this, it was clarified that crude chitin can be obtained from crab shell by extraction with AMIMBr, but a large amount of CaCO ₃ contained in crab shell is contained in this crude chitin.

In contrast, in the XRD spectrum (B) of Sample 1-2 after decalcification with an aqueous citric acid solution, the diffraction peak of 2θ = 30 ° or more derived from CaCO ₃ disappears, and the diffraction peak derived from chitin (spectrum C) Was strongly observed. From this, it was clarified that when a citric acid aqueous solution was added to the supernatant containing Sample 1-1 and decalcified, inorganic salts such as CaCO ₃ were removed, and high-purity chitin could be obtained.

[Determination of degree of deacetylation]
Sample 1-2 thus obtained was analyzed by infrared absorption spectroscopy (IR). The result is shown in figure 2.
As shown in FIG. 2, the IR spectrum (A) of Sample 1-2 shows the same spectral pattern as the IR spectrum (B) of commercially available chitin, confirming that Sample 1-2 is high-purity chitin. .

Here, the degree of deacetylation of chitin according to the method of MIYA et al. (Katsumi MIYA, Reikichi Iwamoto, Koji Ota, Seiichi Mima, 1985, Vol. 42, p. 181-189) Was calculated. In spectrum A of FIG. 2, from the peak area ratio of the absorption peak at 1554 cm ⁻¹ derived from the NH bending angle of the amide and the absorption peak at 897 cm ⁻¹ derived from the CN stretching of the amine or amide, the sample 1- The degree of deacetylation of 2 chitin was determined to be less than 5%.

[Determination of protein content]
The protein content of the obtained sample 1-2 was determined according to the method of Lowry et al. (Lowry, OH et al., J. Biol. Chem., 1951, 193, p. 265-275). To 1.0 g of Sample 1-2, 200 ml of 5 wt / vol% sodium hydroxide aqueous solution was added and stirred at 95 ° C. for 2.5 hours. Thereafter, the supernatant was obtained by centrifugation. To 1.0 ml of supernatant, add 5 ml of Reagent A (45: 5 mixture of 0.10 N NaOH aqueous solution containing 2% NaCO and 1% sodium tartrate aqueous solution containing 0.5% CuSO ₄ · 5H ₂ O) And left for 10 minutes. Next, 0.5 ml of 1N foreign reagent was added and allowed to stand for 30 minutes, and then the absorbance at 750 nm was measured. The obtained absorbance was applied to a calibration curve to determine the protein content. As a result, it was revealed that the protein content of Sample 1-2 was less than 0.1% by weight based on the total weight.

[Determination of weight average molecular weight]
The weight average molecular weight was determined by the viscosity method according to the method of Martin et al. (Martin, P. et al., Carbohydr. Polym., 2002, Vol. 50, p. 363-370). 0.002 g, 0.004 g, and 0.006 g of Sample 1-2 were dissolved in 10 ml of 5 wt% dimethylacetamide (DMAc) / LiCl solution, stirred for 6 hours at room temperature, and then the viscosity was measured. The obtained viscosity was applied to the Mark-Houwink-Sakurada equation to calculate the weight average molecular weight. As a result, the weight average molecular weight M _w was determined to be 3.0 × 10 ⁵ .

<Example 2>
In the procedure for direct extraction and purification of chitin in Example 1, chitin was extracted and purified from crab shells under the same conditions as above except that AMIMBr was added to change the time for extracting chitin. The degree of deacetylation and the weight average molecular weight M _w of the obtained chitin were determined in the same manner as in Example 1. The results are shown in Table 1.

  As shown in Table 1, the chitin yield improved as the extraction time was increased, but the degree of deacetylation and the weight average molecular weight showed the same or substantially the same value. Moreover, the deacetylation degree and weight average molecular weight of chitin of Samples 2-1 to 2-5 were in good agreement with the values of the commercially available chitin of the comparative example.

  The method of the present invention makes it possible to produce high-purity chitin from a crustacean shell or shellfish shell by a simple process.

Claims (3)

Shells or shells containing chitin and inorganic salts and formula I:

[Where:
R ¹ is C ₁ -C ₄ alkyl;
R ² is C ₁ -C ₄ alkyl or C ₂ -C ₄ alkenyl,
X is a halogen. ]
A chitin extraction step for extracting chitin by contacting with the ionic liquid represented by the formula; and an extract containing the chitin obtained in the chitin extraction step and a chelating agent are contacted to remove inorganic salts from the extract Removing inorganic salt;
A method for producing chitin, comprising:   The process of claim 1, wherein the chelating agent is selected from citric acid, ethylenediaminetetraacetic acid and nitrilotriacetic acid.   The method according to claim 1 or 2, wherein in the chitin extraction step, chitin is extracted at a temperature in the range of 80 to 100 ° C for 6 to 24 hours.

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