JP2017038537A - Production method of algaenan composition - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide production methods of efficiently and extremely simply obtaining high purity algaenan compositions from biomass derived from microalgae.SOLUTION: The present invention provides a production method of an algaenan composition wherein floating separation of algaenan from slurry comprising biomass-ground products derived from microalgae is performed. Preferably the biomass-ground products derived from microalgae is obtained by a wet grinding treatment, and preferably the specific gravity of the slurry solvent is 0.9-1.2.SELECTED DRAWING: None

Description

  The present invention relates to a production method for obtaining a highly pure arginane composition from microalga-derived biomass efficiently and very simply.

Hydrocarbons produced by algae are considered promising as biofuels, and research for industrialization is underway. Among the algae, microalgae belonging to the genus Botryococcus are attracting attention as algae from which liquid fuel can be efficiently obtained because they produce hydrocarbons having properties equivalent to heavy oil. The hydrocarbons produced from the microalgae of the genus Botryococcus are divided into four groups, Race-A, Race-B, Race-L and Race-S, based on the structural features, and in particular, C _n H _2n The strain of the Race-B group that produces hydrocarbons having a triterpene structure represented by ₋₁₀ (n = 30 to 37) often produces 30 to 40% by mass of hydrocarbons (for example, non-patent literature) 1).

  A microalga of the genus Botryococcus forms an aggregate (colony) of several to several hundred cells during the growth process, and is a biopolymer (arginane) that is a polymerization compound such as hydrocarbons produced by itself. The colony structure is maintained. The hydrocarbons produced as described above are retained and accumulated up to about 30 to 40% by mass in this arginane. The hydrocarbons accumulated in this way are used as raw materials for biofuels and biorefinery through processes such as solvent extraction, but a large amount of residue from which hydrocarbons have been removed becomes waste, and algae-derived biofuels It became a foothold of industrialization (for example, refer nonpatent literature 2).

  A large amount of residue obtained by removing hydrocarbons from the microalgae of the genus Botryococcus contains about 50 to 60% by mass of arginane, which is a polymerized product of the hydrocarbons described above. In order to increase the purity, conventionally, the residue was used as a raw material, and it was obtained through complicated operations such as removal of elution with various solvents, saponification treatment with potassium hydroxide, phosphoric acid treatment, and washing treatment with various solvents ( For example, refer nonpatent literature 3).

  On the other hand, the structure of alginane has been estimated by instrumental analysis, and although it has a slightly different structure depending on the species of microalgae of the genus Botryococcus, since a double bond exists in the hydrocarbon chain, rubber-like, It is expected to have a sponge-like elasticity (for example, Non-Patent Document 4) and is expected to be used in various industrial fields. However, as described above, complicated purification operations are required, and alginane is an acid, Since it is highly resistant to various chemicals such as alkali and does not dissolve in organic solvents, it is currently not effectively used, and development of a production method for increasing the purity of arginane efficiently and easily is desired. It was.

Microbiol. Cult. Coll. 26 (1). 1-10 (2010) p. 4 2011 Rural and Mountainous Village Sixth Industrialization Measures Project “Survey Report on Practical Use of Algal Biomass Farms in Rural and Mountainous Villages” p. 7 Phytochemistry 22 (2). 389-97 (1983) p. 395 Microbiol. Cult. Coll. 26 (1). 1-10 (2010) p. 5-7

  An object of the present invention is to provide a production method for obtaining a highly pure arginane composition from microalga-derived biomass efficiently and very simply.

  As a result of intensive studies to solve the above problems, the present inventors have found that the microalgae-derived biomass pulverized product can be suspended and separated very easily in the slurry due to the difference in specific gravity between the slurry solvent and the pulverized product. I found. The present inventors have further researched and found that a high-purity arginane composition can be obtained efficiently and simply by subjecting arginane to floating separation from a slurry containing a pulverized biomass of microalgae. Completed the invention.

That is, the gist of the present invention is the following (1) to (7).
(1) A method for producing an arginane composition comprising subjecting arginane to a floating separation treatment from a slurry containing a pulverized product of biomass derived from microalgae.
(2) The method for producing an arginane composition according to (1), wherein the microalgae-derived biomass pulverized product is obtained by a wet pulverization treatment.
(3) The method for producing an arginane composition according to (1) or (2), wherein the slurry solvent has a specific gravity of 0.9 to 1.2.
(4) The method for producing an arginane composition according to any one of (1) to (3), wherein the microalga is derived from the genus Botryococcus.
(5) The method for producing an arginane composition according to any one of (1) to (4), wherein the floating separation treatment is a centrifugal separation treatment.
(6) The method for producing an arginane composition according to any one of (1) to (4), wherein the floating separation treatment is a stationary treatment.
(7) An arginane composition obtained by the production method according to any one of (1) to (6).

  According to the present invention, a high-purity arginane composition can be obtained efficiently and simply by subjecting arginane to a floating separation treatment from a slurry containing a pulverized product of microalgae-derived biomass.

  Hereinafter, the present invention will be described in detail.

  The microalga-derived biomass of the present invention refers to biomass derived from microalgae that mainly produces biofuel, and specific examples include biomass derived from green algae and diatoms. More specifically, Botryococcus braunii, Chlorella spp., Cryptothecodinium chonii, Cylindrotheca spp. Dunaliella primorecta, Isochrysis spp. , Monallanthus salina, Nannochloris spp. , Nannochloropsis spp. , Neochloris spp. , Nitzschia spp. , Phaeodactylum tricornutum, Schizochytrium spp. And biomass derived from Tetraselmis suieia.

  Among these, biomass derived from the genus Botryococcus that produces hydrocarbons equivalent to heavy oil is preferable.

  Biomass derived from microalgae may be any of microalgae itself, dried microalgae, microalgae aggregated and recovered with various coagulants, and dried microalgae aggregated and recovered, and organic solvents from them. It is preferably an extraction residue after the hydrocarbons are extracted by the above, and more preferably an extraction residue after the hydrocarbons are extracted from the aforementioned Botryococcus microalgae with an organic solvent.

  Specifically, the microalgae is one or more selected from a group of organic solvents consisting of hexane, chloroform, methanol, ethanol, diethyl ether and acetone, or a mixed solvent of hexane / acetone, a mixed solvent of chloroform / methanol, ethanol. An extraction residue obtained by dispersing in a mixture of organic solvents exemplified by a mixed solvent of / diethyl ether and extracting hydrocarbons in microalgae can be used as biomass derived from microalgae.

  In the present invention, the microalgae-derived biomass can be used as it is as a raw material for the mechanical pulverization process described later, but it is preferably processed in advance into a shape such as a lump in order to further increase the purity of the alginane.

  The microalga-derived biomass can be made into a microalga-derived biomass pulverized product by mechanical pulverization. The mechanical pulverization treatment can be performed by a wet pulverization treatment in which a solvent is added in addition to performing a mechanical pulverization treatment in a dry state, but a wet pulverization treatment is preferable.

  The wet pulverization process is a mechanical pulverization process in the presence of a solvent. Among mechanical pulverization processes, it is efficient to combine any one or more of ball milling, bead milling, and homogenizer processing. It is preferable from the viewpoint of separation of arginane.

  The solvent used in the wet pulverization treatment is not particularly limited, and examples thereof include water, saline, alkaline aqueous solution, or a mixture thereof.

  In the ball mill treatment and the bead mill treatment, it is preferable to use 0.5 to 2 volumes of media (for example, balls and beads) with respect to 1 volume of biomass from microalgae. The diameter of the medium is not particularly limited, but is preferably 0.03 to 100 mm, more preferably 0.1 to 50 mm, and even more preferably 1 to 20 mm. The material of the media is not particularly limited, but is preferably higher in hardness than microalgae-derived biomass, and more preferably stainless steel, alumina, or zirconia. The grinding container (pot, drum) is preferably the same as the ball.

  The homogenizer treatment includes, for example, a mechanical tearing (rotor / stator) method using a fixed blade and a rotating blade, such as a polytron homogenizer, and a colloid mill method that allows a high-speed rotation between a disk rotor and a stator to pass through at high pressure. Can be used.

  Prior to the wet pulverization treatment, in order to quickly absorb the solvent, the microalgae-derived biomass may be previously immersed in the solvent. The amount of the solvent at the time of wet pulverization or pre-immersion before the wet pulverization may be adjusted while confirming the fluidity of the microalgae-derived biomass in the solvent. 3 to 20 parts by mass is preferable, and 5 to 10 parts by mass is more preferable. High grinding efficiency can be obtained while ensuring fluidity by setting it as 3-20 mass parts.

  The slurry containing the microalga-derived biomass pulverized product in the present invention refers to a slurry obtained by dispersing the above pulverized product in a solvent. The solvent is not particularly limited, and examples thereof include water, saline, alkaline aqueous solution, or a mixture thereof.

  From the viewpoint of promoting the suspension of arginane, the specific gravity of the solvent is preferably 0.9 to 1.2, more preferably 0.95 to 1.1, and even more preferably 1.0 to 1.07. Among them, 0-10 mass% salt solution (the specific gravity of the solvent is equivalent to 1.0 to 1.07) allows rapid separation of arginane from other water-soluble components and insoluble components in the floating separation treatment described below. This is preferable because it occurs. Further, from the viewpoint of the efficiency of separation and purification, the solvent used for the above-mentioned wet pulverization treatment and the solvent used for the slurry are more preferably the same.

Next, the floating separation process will be described.
The floating separation treatment in the present invention is a step of floating and separating arginane from a slurry containing the above-described pulverized product of microalgae using the specific gravity difference between the solvent and arginane. The floating separation treatment is not particularly limited as long as it is a treatment that can float and separate arginane, but centrifugal separation treatment and stationary treatment are also industrially suitable.

  Centrifugation processing is separation processing using centrifugal force, and for example, three-layer separation such as a cylindrical (sharpless) method, a separation plate (disk) method, and a decanter method is preferable.

  In the stationary treatment, arginane is floated and separated in a stationary state from a slurry containing a pulverized biomass of microalgae. In the stationary treatment, it is preferable to use a tank or the like in which an observation window for visually recognizing the solid-liquid interface is installed and a drainage port is installed at the bottom.

  The arginane composition obtained by the aforementioned floating separation treatment can be subjected to steps such as washing, dehydration, drying, and pulverization. Furthermore, in order to improve the usability, it is preferable to perform various water-soluble components and solvent-soluble component removal operations before obtaining a dry powder.

  Examples of the operation for removing various water-soluble components include acid treatment, alkali treatment, hot water treatment and the like. Examples of the chemical used for the operation for removing the water-soluble component include acids such as hydrochloric acid, sulfuric acid and trichloroacetic acid, and alkalis such as sodium hydroxide and potassium hydroxide. Moreover, as temperature of a hot water process, 20-110 degreeC is preferable, 25-80 degreeC is more preferable, and 30-50 degreeC is still more preferable. The acid treatment, alkali treatment and hot water treatment can be performed in combination.

  Examples of the operation for removing various solvent-soluble components include extraction treatment such as immersion, stirring, reflux, and Soxhlet extraction treatment, and subsequent separation treatment such as centrifugation and filtration treatment. Solvents used for removal of various solvent-soluble components include alcohols such as methanol, ethanol and isopropanol, halogen solvents containing chloroform and tetrachloroethane, highly polar solvents such as ethyl acetate and methyl ethyl ketone, toluene and hexane Low polar solvents such as dimethylformamide, dimethyl sulfoxide, N-methylpyrrolidone, and the like, and some of these may be used alone or as a mixed solvent. it can.

  Next, the present invention will be described in more detail by way of examples. In addition, this invention is not limited to an Example.

  The measuring method of the arginane purity (content) in the Example of this invention is shown below.

1. Arginane content (purity)
It calculated | required from the weight decreasing rate in 400 degreeC or more on the following temperature conditions using the differential thermothermal weight simultaneous measuring apparatus (SII nanotechnology Co., Ltd. make, TG / DTA7200) with respect to the arginane composition.
<Measurement conditions>
Start-up temperature: 25 ° C
Final temperature: 800 ° C
Temperature rising rate: + 10 ° C./min (maintained at 100 ° C. for 30 minutes)
Temperature calibration standard sample (indium (156.6 ° C), zinc (419.4 ° C)

<Reference Example 1>
To 500 g of dried Botryococcus microalgae, 5 L of hexane was added, and the mixture was allowed to stand at room temperature for 24 hours to extract hydrocarbons and the like. The same operation was further repeated twice and then air-dried in a draft at room temperature to obtain 350 g of a dried extract residue. The dried residue of the extraction residue was pulverized with a blender and passed through a sieve having a pore size of 5 mm to obtain 320 g of a dried extraction residue powder (Arginane crude product, Arginane purity 33%).

<Comparative Example 1>
Arginane was prepared according to the method described in Non-Patent Document 3. That is, 20 g of the extraction residue dry powder A obtained in Reference Example 1 was washed with 100 mL of acetone by stirring at room temperature for 3 hours, and then insoluble by centrifugation (model 7930, 10,000 G × 20 minutes, manufactured by Kubota Corporation). Subsequently, 100 ml of methanol, chloroform-methanol (vol / vol = 2: 1) and diethyl ether were washed in the same manner, and then air-dried at room temperature in a fume hood to obtain 18 g of a dry residue. Next, saponification treatment (stirring at room temperature for 6 hours) was performed with 100 mL of 6 mass% potassium hydroxide-90% methanol solution. Centrifugation collect | recovered the insoluble content, and 15 g of residue was obtained through water washing | cleaning, ethanol washing | cleaning, acetone washing | cleaning, and diethyl ether washing | cleaning. The residue was further immersed in 100 mL of phosphoric acid and allowed to stand for 13 days under heating at 55 ° C. An excess amount of pentane was added, the insoluble matter was recovered by centrifugation, and washed with water until neutralized. Finally, after washing with 100 mL of water, methanol, acetone, and diethyl ether, 2.0 g of arginane composition (arginane purity 72%) was obtained.

<Example 1>
50 g of extraction residue dry powder A obtained in Reference Example 1 is placed in a 500 mL magnetic pot, and 50 mL of ethanol, 250 mL of 5% by weight saline, and zirconia sphere (mixture of 5 mm diameter 50 mL volume, 10 mm diameter 50 mL volume) are put therein and sealed. Then, a pulverization process was performed at a room temperature at a rotor rotational speed of 250 rpm × 48 hours using a ball mill apparatus (manufactured by Nippon Ceramic Science Co., Ltd., ANZ-50S). The treatment liquid was passed through a sieve having a pore diameter of 1 mm to remove uncrushed materials and zirconia spheres, and then transferred to a transparent container (500 mL separatory funnel) and allowed to stand at room temperature for 2 hours. The lower layer (insoluble content, liquid content) was discarded, and the floating fraction was further washed twice with 250 mL of 5% by mass saline solution, and then the remaining salt was removed by washing with water. The obtained floating fraction was dried in a vacuum dryer set at 40 ° C. for 16 hours and pulverized in a mortar to obtain about 5.0 g of arginane (purity 73%).

<Example 2>
50 g of extraction residue dry powder A obtained in Reference Example 1 is placed in a 500 mL magnetic pot, and 50 mL of ethanol, 250 mL of 5% by weight saline, and zirconia sphere (mixture of 5 mm diameter 50 mL volume, 10 mm diameter 50 mL volume) are put therein and sealed. Then, a pulverization process was performed at a room temperature at a rotor rotational speed of 250 rpm × 48 hours using a ball mill apparatus (manufactured by Nippon Ceramic Science Co., Ltd., ANZ-50S). The treatment liquid was passed through a sieve having a pore diameter of 1 mm to remove uncrushed materials and zirconia spheres, and then transferred to a transparent container (500 mL separatory funnel) and allowed to stand at room temperature for 2 hours. The lower layer (insoluble content, liquid content) was discarded, and the floating fraction was further washed twice with 250 mL of 5% by mass saline. The floating fraction was transferred to a 500 mL beaker, 250 mL of 1N sodium hydroxide-5 mass% brine was added, and alkali treatment was performed with stirring at room temperature for 15 hours. Similarly, the floating fraction was separated using a separatory funnel, and the remaining sodium chloride was removed by washing with 5% by mass sodium chloride water. The obtained floating fraction was dried and pulverized in a mortar to obtain about 3.8 g of arginane (purity 86%).

<Example 3>
50 g of extraction residue dry powder A obtained in Reference Example 1 is placed in a 500 mL magnetic pot, and 50 mL of ethanol, 250 mL of 5% by weight saline, and zirconia sphere (mixture of 5 mm diameter 50 mL volume, 10 mm diameter 50 mL volume) are put therein and sealed. Then, a pulverization process was performed at a room temperature at a rotor rotational speed of 250 rpm × 48 hours using a ball mill apparatus (manufactured by Nippon Ceramic Science Co., Ltd., ANZ-50S). The treatment liquid was passed through a sieve having a pore diameter of 1 mm to remove uncrushed materials and zirconia spheres, and then transferred to a transparent container (500 mL separatory funnel) and allowed to stand at room temperature for 2 hours. The lower layer (insoluble content, liquid content) was discarded, and the floating fraction was further washed twice with 250 mL of 5% by mass saline. The floating fraction was transferred to a 500 mL beaker, 250 mL of 1N sodium hydroxide-5 mass% brine was added, and alkali treatment was performed with stirring at room temperature for 15 hours. Similarly, the floating fraction was separated using a separatory funnel, washed with 5% by mass saline solution, transferred to a 500 mL beaker, added with 250 mL of 1N hydrochloric acid-5% by mass brine, The acid treatment was performed with stirring for a period of time. Similarly, the floating fraction was separated using a separatory funnel, and the remaining sodium chloride was removed by washing with 5% by mass sodium chloride water. The obtained floating fraction was dried and pulverized in a mortar to obtain approximately 3.4 g of arginane (purity 91%).

<Example 4>
50 g of extraction residue dry powder A obtained in Reference Example 1 is placed in a 500 mL magnetic pot, and 50 mL of ethanol, 250 mL of 5% by weight saline, and zirconia sphere (mixture of 5 mm diameter 50 mL volume, 10 mm diameter 50 mL volume) are put therein and sealed. Then, a pulverization process was performed at a room temperature at a rotor rotational speed of 250 rpm × 48 hours using a ball mill apparatus (manufactured by Nippon Ceramic Science Co., Ltd., ANZ-50S). The treatment liquid was passed through a sieve having a pore diameter of 1 mm to remove uncrushed materials and zirconia spheres, and then transferred to a transparent container (500 mL separatory funnel) and allowed to stand at room temperature for 2 hours. The lower layer (insoluble content, liquid content) was discarded, and the floating fraction was further washed twice with 250 mL of 5% by mass saline solution, and then the remaining salt was removed by washing with water. The obtained floating fraction was transferred to a 500 mL beaker, 250 mL of ethanol was added, and the mixture was washed with ethanol while stirring at room temperature for 1 hour. Insoluble matter was recovered by centrifugation (Model 7930, 10,000 G × 20 minutes, manufactured by Kubota Seisakusho), and further washed twice with 250 mL of ethanol. The obtained precipitate was dried and pulverized in a mortar to obtain about 4.1 g (purity 85%) of arginane.

<Example 5>
50 g of extraction residue dry powder A obtained in Reference Example 1 is placed in a 500 mL magnetic pot, and 50 mL of ethanol, 250 mL of 5% by weight saline, and zirconia sphere (mixture of 5 mm diameter 50 mL volume, 10 mm diameter 50 mL volume) are put therein and sealed. Then, a pulverization process was performed at a room temperature at a rotor rotational speed of 250 rpm × 48 hours using a ball mill apparatus (manufactured by Nippon Ceramic Science Co., Ltd., ANZ-50S). The treatment liquid was passed through a sieve having a pore diameter of 1 mm to remove uncrushed materials and zirconia spheres, and then transferred to a transparent container (500 mL separatory funnel) and allowed to stand at room temperature for 2 hours. The lower layer (insoluble content, liquid content) was discarded, and the floating fraction was further washed twice with 250 mL of 5% by mass saline solution, and then the remaining salt was removed by washing with water. The obtained floating fraction was transferred to a 500 mL beaker, 250 mL of ethanol was added, and the mixture was washed with ethanol while stirring at room temperature for 1 hour. Insoluble matter was recovered by centrifugation (Model 7930, 10,000 G × 20 minutes, manufactured by Kubota Seisakusho), and further washed twice with 250 mL of ethanol. The resulting precipitate was transferred to a 100 mL beaker, added with 50 mL of chloroform, and washed with chloroform while stirring at room temperature for 1 hour. Corrugated filter paper (No. 5C) was set in the funnel, and the suspended matter was collected, and further washed while dropping 100 mL of chloroform uniformly. The obtained recovered material was dried and pulverized in a mortar to obtain approximately 3.7 g of arginane (purity 89%).

<Reference Example 2>
5 kg of hexane was added to 1 kg of dried Botryococcus microalgae and allowed to stand at room temperature for 24 hours to extract hydrocarbons and the like. The same operation was further repeated twice to obtain a dried extract residue. The operation so far was repeated four more times to obtain 3.4 kg of dried extract residue. The dried extract residue was pulverized with a blender and passed through a sieve with a pore size of 5 mm to obtain 3.2 kg of dried extract residue powder B (Arginane crude product, Arginane purity 32%).

<Example 6>
2 kg of the extraction residue dry powder B obtained in Reference Example 2 is put into a ball mill apparatus (AB200, manufactured by Chuo Kako), and 30 L of water and zirconia spheres (mixture of 10 mm diameter and 10 L diameter, 50 mm diameter and 5 L volume) are put therein and sealed. Below, the grinding | pulverization process of rotor rotation speed 30rpm x 48 hours was performed. The treatment liquid was dispensed onto a sieve having a pore diameter of 1 mm, and after removing uncrushed materials and zirconia spheres, the treatment liquid was introduced into a shear press centrifuge (manufactured by Kansai centrifuge Seisakusho, model NO-6-A, 15000 rpm). The supernatant was discarded, and the upper layer of the residue in the cylinder was recovered, transferred to a 45 L plastic bucket, and suspended again in 20 L of water. Similarly, it was introduced into a sharp press centrifuge, the supernatant was discarded, and the upper layer portion of the residue in the cylinder was recovered. The same operation was repeated once more. The obtained recovered material was dried and pulverized with a blender to obtain about 390 g (purity 67%) of arginane.

Table 1 shows the results obtained in Examples 1 to 6 and Comparative Example 1.

As shown in Table 1, in Examples 1-6, despite the extremely simple method of floating and separating arginane from a slurry containing a pulverized biomass of microalgae, it is equal to or higher than that of Comparative Example 1 which is a conventional method. The high-purity arginane composition was obtained with good yield. In particular, in Examples 2 to 5 subjected to further purification, a higher purity arginane composition could be obtained. Furthermore, in Example 6, it was found that an arginane composition having a purity level equivalent to that of the conventional method can be obtained even with an industrially applicable apparatus.

Claims (7)

  A method for producing an arginane composition comprising subjecting arginane to a floating separation treatment from a slurry containing a pulverized product of biomass derived from microalgae.   The method for producing an arginane composition according to claim 1, wherein the pulverized biomass of microalgae is obtained by a wet pulverization treatment.   3. The method for producing an arginane composition according to claim 1, wherein the slurry solvent has a specific gravity of 0.9 to 1.2.   The method for producing an arginane composition according to any one of claims 1 to 3, wherein the microalga is derived from the genus Botryococcus.   The said floating separation process is a centrifugation process, The manufacturing method of the arginane composition of any one of Claims 1-4 characterized by the above-mentioned.   The said floating separation process is a stationary process, The manufacturing method of the arginane composition of any one of Claims 1-4 characterized by the above-mentioned. The arginane composition obtained by the manufacturing method as described in any one of Claims 1-6.