JP2009191008A - Method for extracting functional component derived from brown algae - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for extracting functional components derived from brown algae, which solves problems in a conventional method for extracting functional components derived from brown algae and efficiently extracts functional components derived from brown algae being useful components contained in brown algae, further in a greatly shortened time. <P>SOLUTION: The method for extracting functional components derived from brown algae includes extracting brown algae with a supercritical fluid. Preferably the functional components derived from brown algae are fucoxanthin and fucosterol. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to a method for extracting brown algae-derived functional components from marine plant resources.

  Brown algae have functional components derived from brown algae such as fucoxanthin, which is one of carotenoids whose usefulness is recognized, and fucosterol, which is one of sterols.

  Among the functional components derived from the brown algae, fucoxanthin is a known substance having the following structure, and its effects include an antitumor effect (Patent Document 1) and a nerve cell protective effect (Patent Document 2) due to ingestion in the body, Recently, a blood glucose level increase suppressing effect (Patent Document 3) and the like have been reported.

  As described above, since fucoxanthin is effective when ingested, it is desirable to use a fucoxanthin extraction solvent that is harmless to the human body. However, all fucoxanthins used in the above reports are extracted with an organic solvent. Therefore, a technique (Patent Document 4 and Patent Document 5) using ethanol or hydrous ethanol from brown algae at the stage of extraction before purifying fucoxanthin has also been reported.

  However, from the viewpoint of evaporation of the solvent and extraction efficiency, it is desirable that the extraction process and the amount of the solvent used be as small as possible, and it was necessary to omit further steps and to reduce the amount of the solvent compared to the above technique.

  In addition, since the active component of fucoxanthin is easily decomposed by light, it is desirable that the extraction be performed quickly under light shielding. However, the conventional method has a problem in that it takes time to extract fucoxanthin, such as the condition of being shielded from light, the complexity of having to go through several extraction steps, and extraction by soaking for a long time.

  On the other hand, among the functional components derived from brown algae, fucosterol is a known substance having the following structure, and its effects include anti-arteriosclerosis action (Patent Document 6), cholesterol reduction action, and thrombus prevention action by ingestion in the body. Etc. are generally advocated. Fucosterol is used as a functional food or health food derived from natural products without side effects.

However, organic solvents such as aliphatic hydrocarbons such as hexane, esters such as ethyl acetate, ketones such as acetone, or a mixture thereof are used for the extraction of fucosterol, and further 1 to 20 days until the extraction. As is the case with the fucoxanthin, there are problems with the extraction solvent and the extraction time.
Japanese Patent Laid-Open No. 10-158156 JP 2001-335480 A JP 2007-297370 A JP 2004-75634 A JP 2004-35528 A JP 2005-104887 A

  Therefore, the present invention solves the above-described problems in the conventional method, efficiently and functionally improves functional components derived from brown algae such as fucoxanthin and fucosterol, which are useful components contained in brown algae. It is an object of the present invention to provide a method for extracting by shortening.

  As a result of intensive studies to solve the above problems, the present inventors have found that a functional component derived from brown algae can be efficiently extracted from brown algae by utilizing a supercritical fluid, and completed the present invention. .

  That is, this invention is a brown algae origin functional component obtained by the extraction method of the brown algae origin functional component characterized by extracting brown algae with a supercritical fluid, and the said extraction method.

  Since the extraction method of the functional component derived from brown algae of the present invention can greatly shorten the extraction time compared with the conventional method, the exposure time of the sample, that is, the light of the functional component derived from brown algae is also simplified because of its simplicity. Degradation due to can be minimized.

  Further, the above extraction method is characterized by relatively little contamination with chlorophyll, which is a contaminant during extraction, not found in conventional methods, and is easy to use for subsequent separation and purification.

  Therefore, the brown algae-derived functional component extracted by the brown algae-derived functional component extraction method of the present invention can be suitably used for health foods and pharmaceuticals.

  In the present specification, the functional component derived from brown algae is contained in brown algae and refers to pigments such as carotenoid fucoxanthin, fat-soluble components such as fucosterol, polysaccharides, etc., preferably fucoxanthin and It means fucosterol. The functional component derived from brown algae can be measured according to a conventional method. For example, fucoxanthin can be measured by HPLC, and fucosterol can be measured by GC-MS.

  The method for extracting functional components derived from brown algae of the present invention (hereinafter referred to as “the present invention extraction method”) is performed by supercritical fluid extraction using brown algae as an extraction raw material and using this as a supercritical fluid as an extraction solvent. It is.

  In the extraction method of the present invention, the brown algae used as the extraction raw material can be widely adopted as long as they contain functional components derived from brown algae. Examples of such brown algae include brown algae belonging to the order of Chordaceae, Fucales, and Laminariae. Among these brown algae, there are Chronaceae or Chronaceae, Spermatochaceae, Sargassaceae, Laminariaceae, and Alaeceae (Alaria). Preferably, among the brown algae belonging to the above family, the mosquitoes of the family Nymphalidae (Cladosiphon okamuranus) or the moss family (Nemacystus decipiens), the kingfisher family (Sargassum fulhum) The Alame (Eisenia bicyclis (Kjellman ) Setchell), Laminaria japonica (Laminaria japonica), Onikonbu (Laminaria diabolica), Rishirikonbu (Laminaria ochotensis), Hosomekonbu (Laminaria religiosa), Mitsuishikonbu (Laminaria angustata), Nagakonbu (Laminaria longissima) or Gagomekonbu (Kjellmaniella crassiifolia), of Chigaiso Department of seaweed (Undaria pinnatifida) is preferable, and particularly, the Okinawa Mosque of the Nagamatsue family is more preferable because it contains a functional component derived from brown algae at a high concentration. The brown algae used as the raw material for extraction is the release of zoospores from matured bodies, the generation of discoids or filaments from the zoospores, the formation of a solid body by the implantation of a disklike body or a filamentous body, Of the growth cycle of growth into a plate, it may be in any growth stage of a plate or filamentous body to a mature body, but since it contains abundant functional components derived from brown algae, it is a plate or filament. The body is preferred. In addition, although a board | plate-like body or a filamentous body is located in the middle of the zoospore and a direct solid in the said growth cycle, and may be called with another name, in the present invention, all of these are included. Furthermore, these extraction raw materials are preferably pulverized to an appropriate size as necessary before extraction with a supercritical fluid described later, in order to make the density in the container used for extraction as small as possible, In particular, it is preferable to use a powder form.

  In the extraction method of the present invention, as the supercritical fluid used as the extraction solvent, for example, a higher pressure tends to be preferable, and therefore, 7.38 MPa or more, preferably 10 MPa or more, particularly preferably 30 MPa or more, Examples thereof include carbon dioxide in a supercritical state at a temperature of 25 to 80 ° C., preferably 30 to 80 ° C., particularly preferably 70 to 80 ° C.

  Supercritical fluid extraction of functional components derived from brown algae using the above-mentioned brown algae and supercritical fluid is a pump for generating a supercritical fluid, and contacting the supercritical fluid and brown algae for extraction. It is carried out in a supercritical extraction apparatus comprising a container, a container for storing the extract, and an oven for heating and keeping the entire system. As such a supercritical fluid extraction device, a general device commercially available from each company can be used.

  Although the extraction conditions such as the flow rate and extraction time of the supercritical fluid in the supercritical fluid extraction change depending on the size of the container and the like, it cannot be generally stated. For example, if the container has an internal volume of 10 ml, the flow rate is modified as described later. 1 to 2 ml / min in combination with the yarn, and the extraction time is preferably 5 to 30 minutes, more preferably 5 to 20 minutes.

  In addition, it is preferable to add a modifier to the supercritical fluid during the supercritical fluid extraction. Examples of such a modifier include ethanol and / or water, ethanol and water are preferable, and ethanol alone is more preferable. When ethanol and water are used as a modifier, the ethanol concentration is 1 to 100% by mass (hereinafter simply referred to as “%”), preferably 50 to 100%. By adding this modifier to the supercritical fluid, the extraction efficiency is improved and the yield of functional components derived from brown algae is increased. Further, the amount of the modifier is preferably 1 to 50%, particularly preferably 2 to 30%, of the total flow rate of the supercritical fluid and the modifier combined.

  By the supercritical extraction, an extract containing brown algae-derived functional components is extracted from brown algae. Among the functional components derived from brown algae, for example, fucoxanthin is about 0.5 to 100 μg per gram of the extract, and fucosterol is 0.1 to 2 per gram of the extract. Among the functional components derived from brown algae, for example, fucoxanthin is about 10 to 2060 μg per gram of extract, and fucosterol is 1 per gram of extract. About 0.0 to 5.0 mg is contained. Moreover, this extract has few impurities, such as chlorophyll, compared with the conventional method. Furthermore, the functional component derived from brown algae may be separated and purified from this extract by a general purification method such as chromatography or HPLC.

  The functional components derived from brown algae thus obtained include various health foods that are expected to have the above-described cholesterol-reducing action and thrombus-preventing action, pharmaceuticals for the purpose of antitumor effect, nerve cell protecting effect, blood sugar level increase inhibiting effect, etc. Can be used for

  Although the Example regarding extraction of a functional component derived from brown algae is shown below, this invention is not limited to these Examples at all.

Example 1
Supercritical fluid extraction of mature Okinawa mozuku (1):
(1) Sample A matured body of Okinawa moss (Cladosiphon okamuranus) was freeze-dried, stored frozen and milled (powder).

(2) Supercritical fluid extraction About 1 g (dry weight) of sample per specimen is placed in an extraction container having an internal volume of 10 ml, and placed in a predetermined position of a supercritical extraction system (SCF series: manufactured by JASCO Corporation). After the column oven reached 40 ° C., extraction was performed at a set pressure (10-45 MPa) with supercritical carbon dioxide and modifier at a total flow rate of 2 ml / min for 20 minutes (total solvent amount 40 ml). In addition, ethanol was used as a modifier, and the flow rate was set to 0.1 ml / min.

(3) Measurement of fucoxanthin The extract obtained in (2) above was dried and concentrated on a rotary evaporator, then dissolved in ethanol to prepare a sample for HPLC analysis, which was quantified under the following conditions. Table 1 shows the amount of fucoxanthin in the extract obtained by extracting the pressure conditions at the time of supercritical extraction at 10, 30 or 45 MPa.
<HPLC analysis conditions>
Column: Wakosil-II 5C18 HG (φ4.6 × 150 mm)
Eluent A: Water Eluent B: Acetonitrile Flow rate: 1.0 ml / min Detection wavelength: 440 nm

(4) Measurement of fucosterol The extract obtained in (2) above was dried and concentrated on a rotary evaporator, and then used as a sample for GC-MS analysis, which was quantified under the following conditions. Table 2 shows the amount of fucosterol in the extract obtained by extracting the pressure conditions at the time of supercritical extraction at 10, 30 or 45 MPa.
<GC-MS analysis conditions>
Column: DB-5MB (φ0.25 × 30 m, 0.25 μm film)
Column temperature: 250 ° C. (2 minutes) − (temperature rise at 3.0 ° C./min)−320° C. (3 minutes)
Ion source temperature: 200 ° C
Interface temperature: 320 ° C
Ionization method: EI
Detection mode: SIM

(5) Results From the above results, it was found that both fucoxanthin and fucosterol are extracted most when the extraction pressure is 45 MPa.

Example 2
Supercritical fluid extraction of mature Okinawa mozuku (2):
Of the supercritical fluid extraction conditions of Example 1, the extraction pressure was set to 30 MPa, and the temperature conditions were 30, 40, or 75 ° C. About the obtained extract, the fucoxanthin amount and the fucosterol amount were measured in the same manner as in Example 1. The results are shown in Table 3.

  From the above results, it was found that both fucoxanthin and fucosterol were extracted most when the extraction temperature was 75 ° C.

Example 3
Effect of modifier in supercritical fluid extraction (1):
(1) Sample The sample similar to Example 1 was used for extraction.

(2) Supercritical fluid extraction About 1 g (dry weight) of sample per specimen is placed in an extraction container having an internal volume of 10 ml, and placed in a predetermined position of a supercritical extraction system (SCF series: manufactured by JASCO Corporation). After the column oven reaches 40 ° C., the supercritical carbon dioxide and modifier (ethanol) are brought to a total flow rate of 2 ml / min at a pressure of 30 MPa, and the modifier flow rate is 0, 2, 5, 10 or 30% of the total flow rate. As a result, extraction was performed for 20 minutes.

(3) Measurement The amount of fucoxanthin and the amount of fucosterol was measured in the same manner as in Example 1 for the extract obtained by extracting the modifier flow rate with 0, 2, 5, 10 or 30% of the total flow rate. The results are shown in Table 4.

  From the above results, it was found that in the supercritical fluid extraction, when a modifier is used together with the supercritical fluid, fucoxanthin and fucostol are extracted most when the flow rate is 2 to 10% of the total flow rate.

Example 4
Effect of modifier in supercritical fluid extraction (2):
Of the supercritical fluid extraction conditions of Example 3, the flow rate of the modifier is fixed at 5%, the modifier is a mixture of ethanol and water and ethanol, and the ratio is ethanol concentration 50, 70, 90 or 100%. Extracted. About the obtained extract, the fucoxanthin amount and the fucosterol amount were measured in the same manner as in Example 1. The results are shown in Table 5.

  From the above results, it was found that as the concentration of ethanol contained in the modifier is higher, more fucoxanthin and fucosterol are extracted.

Example 5
Supercritical fluid extraction of mature Okinawa mozuku (3):
(1) Sample The sample similar to Example 1 was used for extraction.

(2) Supercritical fluid extraction About 1 g (dry weight) of sample per specimen is placed in an extraction container having an internal volume of 10 ml, and placed in a predetermined position of a supercritical extraction system (SCF series: manufactured by JASCO Corporation). From the results of Examples 1 to 4, the supercritical fluid extraction conditions were as follows: the extraction pressure was 45 MPa, the extraction temperature was 75 ° C., and 100% ethanol as a modifier was 5% (0.1 ml / min) of the total flow rate (2 ml / min). Min) and extraction was performed for 20 minutes.

(3) Measurement The amount of fucoxanthin and the amount of fucosterol were measured in the same manner as in Example 1 for the extract obtained by the above extraction. The amount of chlorophyll in the extract was measured with a spectrophotometer at 665 nm. In addition, in this measurement, since the measurement result of the amount of chlorophyll was not compared with the standard sample, the amount of chlorophyll was shown as chlorophyll / fucoxanthin.

Comparative Example 1
Ethanol extraction of mature Okinawa mozuku (1):
40 ml of ethanol was added to about 1 g (dry weight) of the same sample as in Example 1 and stirred for 1 minute. Thereafter, the mixture was centrifuged at 3000 rpm for 10 minutes, allowed to stand for 1 hour, further stirred for 1 minute, and the supernatant was collected. The solvent was distilled off from this supernatant to obtain an extract. For this extract, the fucoxanthin content, fucosterol content and chlorophyll content were measured in the same manner as in Example 5.

Comparative Example 2
Ethanol extraction of mature Okinawa mozuku (2):
40 ml of ethanol was added to about 1 g (dry weight) of the same sample as in Example 1, stirred and allowed to stand for 24 hours, and the supernatant was collected. The solvent was distilled off from this supernatant to obtain an extract. For this extract, the fucoxanthin content, fucosterol content and chlorophyll content were measured in the same manner as in Example 5.

Test example 1
Comparison of fucoxanthin content in all extracts:
Table 6 shows the amounts of fucoxanthin, fucosterol and chlorophyll (chlorophyll / fucoxanthin) measured in Example 5 (supercritical fluid extraction method), Comparative Example 1 (conventional method) and Comparative Example 2 (conventional method). Indicated.

  From the above results, in the supercritical fluid extraction method (Example 5), more fucoxanthin and fucosterol were obtained in spite of a shorter time and easier extraction than the conventional methods (Comparative Examples 1 and 2). In addition, it was found that the supercritical fluid extraction method has a lower ratio of fucoxanthin and chlorophyll extracted together with fucoxanthin than that of the conventional method, and extraction of impurities is small.

Example 6
Supercritical fluid extraction of Okinawa Mozuku board (1):
(1) Sample A sample obtained by freeze-drying a plate of Okinawa mozuku (Cladosiphon okamuranus), storing it frozen and milling (powder) was used as a sample.

(2) Supercritical fluid extraction About 1 g (dry weight) of sample per specimen is placed in an extraction container having an internal volume of 10 ml, and placed in a predetermined position of a supercritical extraction system (SCF series: manufactured by JASCO Corporation). After the column oven reached 40 ° C., extraction was performed at a set pressure (10-45 MPa) with supercritical carbon dioxide and modifier at a total flow rate of 2 ml / min for 20 minutes (total solvent amount 40 ml). In addition, ethanol was used as a modifier, and the flow rate was set to 0.1 ml / min.

(3) Measurement of fucoxanthin The extract obtained in (2) above was dried and concentrated on a rotary evaporator, then dissolved in ethanol to prepare a sample for HPLC analysis, which was quantified under the following conditions. Table 7 shows the amount of fucoxanthin in the extract obtained by extracting the pressure conditions at the time of supercritical extraction at 10, 30 or 45 MPa.
<HPLC analysis conditions>
Column: Wakosil-II 5C18 HG (φ4.6 × 150 mm)
Eluent A: Water Eluent B: Acetonitrile Flow rate: 1.0 ml / min Detection wavelength: 440 nm

(4) Measurement of fucosterol The extract obtained in (2) above was dried and concentrated on a rotary evaporator, and then used as a sample for GC-MS analysis, which was quantified under the following conditions. Table 8 shows the amount of fucosterol in the extract obtained by extracting the pressure conditions at the time of supercritical extraction at 10, 30 or 45 MPa.
<GC-MS analysis conditions>
Column: DB-5MB (φ0.25 × 30 m, 0.25 μm film)
Column temperature: 250 ° C. (2 minutes) − (temperature rise at 3.0 ° C./min)−320° C. (3 minutes)
Ion source temperature: 200 ° C
Interface temperature: 320 ° C
Ionization method: EI
Detection mode: SIM

(5) Results From the above results, it was clarified that fucoxanthin and fucoxanthin were extracted from the plate of Okinawa mozuku as well as the mature body. It was also found that fucoxanthin was extracted most when the extraction pressure was 45 MPa and fucosterol was extracted when the extraction pressure was 30 MPa.

Example 7
Supercritical fluid extraction of Okinawa Mozuku board (2):
Of the supercritical fluid extraction conditions of Example 6, the extraction pressure was set to 30 MPa, and the temperature conditions were 30, 40, or 75 ° C. About the obtained extract, the fucoxanthin amount and the fucosterol amount were measured in the same manner as in Example 6. The results are shown in Table 9.

  From the above results, it was found that both fucoxanthin and fucosterol were extracted most when the extraction temperature was 75 ° C.

Example 8
Effect of modifier in supercritical fluid extraction (4):
(1) Sample The sample similar to Example 6 was used for extraction.

(2) Supercritical fluid extraction About 1 g (dry weight) of sample per specimen is placed in an extraction container having an internal volume of 10 ml, and placed in a predetermined position of a supercritical extraction system (SCF series: manufactured by JASCO Corporation). After the column oven reaches 40 ° C., the supercritical carbon dioxide and the modifier (ethanol) are brought to a total flow rate of 2 ml / min at a pressure of 30 MPa, and the modifier flow rate is 0, 2, 5 or 30% of the total flow rate. Extraction was performed by performing extraction for a minute.

(3) Measurement The amount of fucoxanthin and the amount of fucosterol were measured in the same manner as in Example 6 for the extract obtained by extracting the modifier flow rate as 0, 2, 5 or 30% of the total flow rate. The results are shown in Table 10.

  From the above results, it was found that in the supercritical fluid extraction, when a modifier is used together with the supercritical fluid, fucoxanthin and fucosterol are extracted most when the flow rate is 2 to 30% of the total flow rate.

Example 9
Effect of modifier in supercritical fluid extraction (5):
Of the supercritical fluid extraction conditions of Example 8, the flow rate of the modifier was fixed at 5%, and the modifier was extracted from ethanol alone using a mixture of water and ethanol, and the ratio was extracted with an ethanol concentration of 50, 90, or 100%. . About the obtained extract, the fucoxanthin amount and the fucosterol amount were measured in the same manner as in Example 6. The results are shown in Table 11.

  From the above results, it was found that as the concentration of ethanol contained in the modifier is higher, more fucoxanthin and fucosterol are extracted.

Example 10
Supercritical fluid extraction from various brown algae:
Arame (Eisenia bicyclis (Kjellman) Setchell), Kombu (unknown type), Hizuki (Fujii fusiformis), Hawksawara (Sargassum fulvalum C. Agardh) or Wakame (Undariamil) Then, a small piece or powder was used as a sample, and supercritical fluid extraction was performed in the same manner as in Example 6. About the obtained extract, the fucoxanthin amount and the fucosterol amount were measured in the same manner as in Example 6. The results are shown in Table 12.

  From the above results, it was revealed that fucoxanthin and fucosterol were extracted from any mature body of brown algae.

  According to the extraction method of the present invention, functional components derived from brown algae such as fucoxanthin and fucosterol can be efficiently extracted from brown algae in a short time.

Therefore, the functional component derived from brown algae extracted by the extraction method of the present invention can be suitably used for health foods, pharmaceuticals and the like.

more than

Claims (5)

  A method for extracting functional components derived from brown algae, characterized by extracting brown algae with a supercritical fluid.   The method for extracting a brown algae-derived functional component according to claim 1, wherein the functional component derived from a brown algae is fucoxanthin and fucosterol.   The method for extracting a functional component derived from brown algae according to claim 1 or 2, wherein the brown algae is a plate-like body or a filamentous body of brown algae.   The method for extracting a functional component derived from brown algae according to any one of claims 1 to 3, wherein ethanol and / or water is further added to the supercritical fluid as a modifier.   A functional component derived from brown algae obtained by the extraction method according to claim 1.