TW200408707A - New use of porphyra dentata for producing high od phycoerythrin - Google Patents

Abstract

The invention relates to a new use of Porphyra dentata for producing high OD phycoerythrin. It has been found that Porphyra dentata has three generations in the life cycle: sexual reproduction, asexual reproduction and vegetative propagation. Then, carpospore filamentous plants thereof do not contain gel and can be maintained under some controlled conditions. The invention takes advantage of these findings to provide a new use for producing high OD phycoerythrin from carpospore filamentous phase of Porphyra dentata.

Description

200408707 V. Description of the invention (1) 5-1 Field of the invention: The present invention relates to a new use for discovering Porphyra dentata, especially for the discovery that Porphyra lutea can obtain algae red with high absorbance (0D) ratio A new use of vegetarian. 5-2 Background of the Invention: Regardless of phycoerythrin, phycocyanin or other natural protein pigments, it is usually practical and safe, and has a certain degree of stability to heat, pH, etc., and can be applied to food and cosmetics. At the same time, it can also be used as a fluorescent pigment for immunological antibody labeling in clinical diagnosis and cytobiochemical research. At present, phycocyanin and phycoerythrin have been developed and applied in the market. Since the raw material sources of phycocyanin production are mostly blue-green algae that can be mass-produced and cultivated, such as Spirul ina and Microcystis aeruginosa (Microcystis), and its cultivation and production methods have many patents. However, phycoerythrin is produced due to lack of raw materials or unfavorable pigment production of raw materials, but the output is small and expensive. For the global demand for tens of thousands of pounds of edible red pigment each year, a natural, safe, stable, and special fluorescent red pigment is necessary. After mass production, it is estimated that the price will fall, and its application range and market supply will be expanded. . At present, most of the phycoerythrin protein is isolated from the large leaflets of red algae.

200408707 V. Description of the invention (2) Such as Porphyra, Ceramium, etc., a small part is extracted from Porphyridium, which can be controlled in large quantities. Although there are a large number of wild red algae resources and cultivated laver as raw materials for phycoerythrin, most red algae are rich in colloids (agar gum, carrageenan, red algae gum), which makes it difficult to extract pigments, especially dry. The latter algae materials are even more so, coupled with the seasonal differences in quality and quantity of the raw materials of wild algae species are difficult for humans to grasp. At present, the culture of Porphyridium has significant difficulties in collecting cells. The collection of single-celled algal bodies has always been an energy-consuming and time-consuming operation, which affects the production cost and the soluble polysaccharides secreted by the algae cells during culture. In addition to hindering the collection of cells, it also affects the extraction of pigments. In order to solve the above-mentioned problems, U.S. Patent No. 5,358,858 proposes a method for producing phycoerythrin from the filaments of Bangia atropurpurea and Porphyra angusta, using the life of hair vegetables and laver The filamentous sporophyte stage in history does not contain the characteristics of various colloids. Using controlled light, temperature, and nutritional conditions, the filamentous stage is continued, and it is further used as a material to extract phycoerythrin. The steps are as follows: : 1. Cultivation and reproduction of filaments. Collect mature hair dish or laver gamete from the field, clean the algae with sterilized sea water and a writing brush, and dry it and put it into the modified SWM-III (as shown in Table 1). (Shown) In the culture orphan, the bottom of the culture orphan is covered with a cover glass. After the spores are released and attached to the cover glass,

200408707 V. Description of the invention (3) At room temperature, the illuminance of 1 0 0 0 1 u X ~ 4 0 0 0 1 u X and the incubator of 10 to 16 hours of light per day germinate and grow, and the final development is different. Branchy filamentous sporophytes. Table 1 swm- m

NaNO3 8.5 g / 100 ml 2 m 1 Na2HP04 0.595 g / 100 ml 2 m 1 Na2EDTA 0.5 g / 100 ml 2 m 1 0 · 01625 g / 100 ml 2 m 1 FeCl3 (or FeCl3 7H20) 0.02885 g / 100 ml 2m 1 * PI-meta1 2 m 1 Vitamins 2ml Soil extract 50ml Tris (10 cc / 1) 5 0 0 rag Liver extract 1 0 mg Sea Water up to pH = 7. 5 11 (When adjusting the pH: First adjust to pH < = 7 with concentrated HC1, then to p Η 7. 5 with appropriate concentration of NaOH, so as to avoid white precipitation during disinfection) 12.368 g 1.385 g 0.109 g then distilled water 21 * ΡI-meta1 Η3B03 MnCl2 ZnCl2

Page 8 200408707 V. Explanation of the invention (4)

CoC I26H20 4.4 7 9 mg

CuCl2 2H20 0. 0 3 4 mg ** S-3 Vitamins

Thiamin HC1 0.5 g

Capantothenate 0.1 g

Nicotinic acid 0.1 g P-am i nobenzo i c acid 10 mg redistilled water 21 Refrigerated storage)

Biotin 1 mg

Inositol 5 g

Folic acid 2 mg

Thymine 3 mg B12 1 mg (All St o c k solutions are stored in brown bottles

2. After the filaments were scraped off, they were transferred to a triangular flask containing a culture solution, and cultured under the above-mentioned growth environment conditions to produce a cluster of filaments. Broken the jujube of the filaments, moved to a larger growth volume bottle and continued to develop into more jujubes, and therefore gradually expanded the culture volume. In a larger volume of light culture tank, it should be pumped (per minute 300 ml of clean air). By filtering with 100 ~ 400 mesh cloth, the filtered culture medium can be recovered and reused without any hindrance to subsequent cultivation. 3. After the algal body is harvested, the filamentous structure can be quickly dried by vacuum or warm air, and then ground into powder, and then fully stirred and extracted with water or phosphate solution.

Page 9 200408707 V. Description of the invention (5) The clarified deep red algae protein solution can be obtained by centrifugation and centrifugation, and the algal pigment can be concentrated and purified. Some 20% ammonium sulfate solution can be used to remove some of the impurity protein precipitates. A 60% to 65% ammonium sulfate solution precipitates a pigment protein, which is a crude red pigment with OD 5 65 / 〇D280 = 1.4 to 1.6, which is a food-grade, cosmetic-useable pigment. 4. The precipitated protein is further purified by gel filtration chromatography (ge 1 filtration), and separated using Sephadex G200 resin. The first procedure can obtain a phycoerythrin product with an OD ratio of 3. 3 ~ 3.7. Repeat A phycoerythrin with an OD ratio of 5.1 to 5.2 can be obtained, and the result obtained by gel electrophoresis (SDS) electrophoresis shows a purity of about 99%, which can be used as a reagent for immunoassay. Although the above method can avoid the complicated procedures of heating and separating gel value in the traditional pigment extraction method of laver and seaweed, or the single cell plant extraction method such as Porphyridium due to its small volume, it is difficult to collect and the polysaccharides it secretes. Problems affecting the extraction of pigments, but the deep red algae protein solution formed directly by the hair dishes and laver laver filaments, the 0D565 / 0 D28Q only 1. 4 ~ 1.6, still need to be concentrated and purified To obtain phycoerythrin with a high OD value, we need to find the filaments of other algae species. The phycoerythrin solution formed by it for the first time has a phycoerythrin with a high OD value. 5-3 Purpose and Summary of the Invention: In view of the above background of the invention, it is known to use hair dish and laver

Page 10 200408707 V. Description of the invention (6) The problem that the absorbance ratio of the deep red algae protein solution formed directly by the filaments is low. The present invention finds that a new application of phycoerythrin having a high absorbance ratio can be obtained from Porphyra longae, which avoids the above situation. An object of the present invention is to discover a new use of Porphyra latifolia to obtain phycoerythrin. With its high weight ratio containing phycocyanin per unit weight, the goal of reducing unit costs is achieved. Another object of the present invention is to discover a new use of P. longifolia for obtaining phycoerythrin. By extracting phycoerythin with a high absorbance ratio, the purpose of reducing the phycoerythrin purification step is achieved. According to the above-mentioned object, the present invention finds a new use of algae species, whose life history has three generations of sexual reproduction, asexual reproduction and vegetative reproduction, and uses its spore filaments to produce algae red with high absorbance ratio Vegetarian. According to the above concept, the spore filaments are fruit spores (c a r ρ 〇 s ρ 〇 r e).

200408707 V. Description of the invention (7) According to the above idea, the phycoerythrin produced by the filamentous body of Porphyra longifolia fruit spores has a high absorbance ratio, comprising the following steps: culturing the gametophyte containing mature fruit sporangia of Porphyra longifolia with culture medium And obtain fruit spores; culture the fruit spores at 15 ~ 30 ° C under the environment of 500 lux ~ 6 0 0 lux with an illuminance and light-dark ratio of 10:14 or more, and grow them into filaments Collecting the filaments; adding an acid test buffer and stirring: stirring and centrifuging to obtain a dark pigmented protein solution; and removing impurities in the dark pigmented protein solution by salting out and precipitating pigmented protein. According to the above concept, the culture medium is SWM-III culture medium. According to the above concept, the swm-m culture solution is a swm-m culture solution without organic matter. According to the above concept, a method in which the fruit spore filaments are cultured is a spin-floating culture method. According to the above idea, the culture environment is 20 ° C, and the illumination and light-dark ratio of 20000 lux is 1 2: 12. According to the above concept, the step of collecting the filaments further comprises: collecting the filaments by a mesh collection method; drying the filaments; and grinding the filaments into a powder.

200408707 V. Description of the invention (8) According to the above-mentioned idea, wherein the net cloth collection method uses 20 to 4 0 mesh netting ° According to the above idea, wherein the method of drying the filament is a vacuum method. According to the above concept, a condition in which the filament is dried is a warm air method. According to the above conception, the conditions for centrifugation are a rotation speed of 6 0 0 r p m, a time of 10 m i η, and a temperature of 4 ° C. According to the above concept, the acid-base buffer solution is a potassium phosphate solution to maintain the solution's pp value between 5 and 10. According to the above concept, the salting-out method removes impurities by adding 20% ammonium sulfate solution. According to the above conception, in which the pigment protein is precipitated by the salting out method, a 60% to 65% ammonium sulfate solution is added. According to the above concept, the method further comprises purifying and separating phycoerythrin by ultrafiltration. According to the above concept, it further includes purification and separation by gel filtration chromatography.

Page 13 200408707 V. Description of the invention (9) Phycoerythrin. According to the above concept, the gel filtration chromatography method is a gel filtration chromatography method using Sephadex G 2000. According to the above-mentioned equipment for sexual reproduction and asexual fruit spores (carpospore), including: a first (Porphyra dentata); a second culture tank, 10:14 or more and 15 ~ 30 to grow it into Filaments; a grinder for grinding the filaments after grinding the filaments, for the purpose of obtaining the high-performance spectrogram of phycoerythrin from the dark pigmented egg according to the above concept, as described in the patent specification, the present invention A kind of phycoerythrin-culture tank with high absorptivity ratio was produced by using the filaments of three generations of alternate algae species with life history and vegetative reproduction. The fruit spores have an environment of 5001Ux ~ 600iUXi illumination and light-dark ratio C for culturing the fruit spores, a collector 'for collecting the filaments; a research J to: f ί 器' Add acid-base buffer and j / coating pigment protein solution; a white solution precipitates phycoerythrin. The chromatogram of 56511111 obtained by extracting the fruit spore filaments of Porphyra spp. With liquid phase layer # 565 in the figure 7 is based on the above concept, and the culture medium is SWM-dish culture medium according to the above Idea, where

Dl culture medium is deorganized

200408707 V. Description of the Invention (ίο) 5 W Μ-ΠΙ broth. According to the above concept, a method in which the fruit spore filaments are cultured is a spin-floating culture method. According to the above idea, the culture environment is 20 ° C, and the illumination and light-dark ratio of 20000 lux is 1 2: 12.

According to the above concept, the collector uses a mesh cloth to collect the filaments. According to the above concept, the mesh fabric is a mesh fabric of 20 to 400 mesh. According to the above concept, the grinder further includes a dryer. According to the above concept, the dryer is a vacuum dryer. According to the above concept, the dryer is a warm air dryer. According to the above concept, the precipitator is a precipitating phycocyanin using a salting-out method.

According to the above concept, wherein the salting out method precipitates phycoerythrin by adding 60 ° / 0 to 65% beryllium sulfate solution.

Page 15 200408707 V. Description of the invention (11) According to the above conception, it further includes a purifier, and the phycoerythrin is purified by gel filtration chromatography. According to the above concept, the gel filtration chromatography method is a gel filtration chromatography method using S p h a d e X G 2 0 0. I would like to describe the above according to its roots. This method has been purified by borrowing. A detailed explanation of the red envelope was issued in detail, except for the line. , Shi Zhuan, for example, siege. The actual description of Li Ru's description of it in detail will be extensively exemplified in the following details. Shi Keding is also limited to Jia Ming, who is less likely to accept the surrounding statements, and the statements outside A ’s hair. Hair Profile J = p Phycobiliprotein is a water-soluble fluorescent protein pigment from algae. Due to its special fluorescent properties, it is currently widely used in fluorescent reagents for flow cytometers. Among the various phycobiliproteins, phycoerythrin is one of them, and it is also the one with the highest fluorescence intensity in natural materials. Therefore, there are many fluorescent detection methods using such pigments. UV absorption and fluorescence emission chromatograms obtained by a chromatograph (HPLC). The chromatographic conditions are as follows: HPLC column: HYDROCELL DEAE NP10

Page 16

200408707 V. Description of Invention (12)

Column size: 50 氺 4.6mm

Buffer A: 10mM K-PBS pH 6.0

Buffer B: 10 mMk-PBS, 0.5 M NaCl pH 6.0

Gradient: 0% Buffer B-> 12min-&50; 50% Buffer B

Detection: 565nm

Flow rate: lml / min and high performance liquid chromatography is mainly composed of high precision high pressure pump, separation tube, detector and recorder. Observing the life history of the hairy cabbage (as shown in the second picture) and the life history of the laver (as shown in the third picture), we can know that the life history of the two algae is generally sexual reproduction and asexual reproduction. Alternating generations, sexual reproduction is the gametophyte (female gametophyte) and sperm (male gametophyte). When the female gametophyte is fertilized, it will develop sporangia. When the fruit spores mature, they will diffuse and become silk. Sporophyte. Asexual reproduction is the simple germination of monospores into small algae. When a certain period of time, monospores can be produced, and the monospores germinate into small algae. This cycle continues until the appropriate environmental conditions. The spores can germinate into large algae. At the same time, small algae can also be transformed into large algae, and then enter the sexual reproductive generation. The conventional technique is to take advantage of the fact that the filamentous sporophyte stage does not contain various gums. Under the controlled light, temperature, and nutritional conditions, it continues its filamentous stage, and further uses it as a material to extract phycoerythrin. Algae life history except for two generations of sexual reproduction and asexual reproduction

Page 17 200408707 V. Description of the invention (13) In addition to the alternation, another part of the algae has one more vegetative breeding generation, that is, the sporophyte (ie, the tetrasporium) of the owner's tetraspore. The process is after fertilization of the fruit generator. Formation of carposporephyte, carpospore, tetrasporophyte, tetrasporangium, and finally tetraspore, in which both fruit spores and tetraspores germinate into filaments Shape body 'But the characteristics of these two filaments are different, please refer to the sea surface (N ema 1 i 0 n) life history (as shown in the fourth figure). Similarly, the present invention uses the long-leaf laver with a vegetative breeding generation. The fruit spore filamentous stage does not contain the characteristics of various gums. Under controlled light, temperature, and nutritional conditions, the filamentous stage continues, and It is further used as a material to extract phycoerythrin 'with a high value of 01). The steps are as follows: 1. After the sporangia are dried, the ratio of the inner glass sheet to the bottom is 1 0. Finally, 2. The aggregate of the growing ring is larger. Cultivate and multiply, the mature long-leaf laver contains mature fruit-body ', clean the algae body with sterilized seawater and a writing brush, 2: in an incubator of SWM-m culture solution of organic matter, code = glass J-separated slices, Zori Private 2 After the mouth of the tussah silkworm, it was released at 15 ~ 30 / Dingguozi (CarP0SP〇re) and attached to the illuminance of '500001UX ~ 600001UX' at a temperature of 14 or more b, Ming The niece ua ± and the mother's day above 10 light in the incubator germinated and grew the branched filamentous body of the moon king. Grow, scrape, move to a large culture tank containing culture medium, and culture under two f pieces, which can grow into a cluster of filaments. Type II soil 2 to continue to develop into more aggregates of filaments In the light culture tank of the group, v, shape, it should be pumped to make the small aggregate = floating j

Page 18 200408707 V. Description of the invention (14) A large number of breeding cultures are carried out in the culture medium (referred to as rotary floating culture). By filtering with 20 ~ 400 mesh cloth, the culture solution can be recovered and reused without any hindrance to subsequent cultivation. 3. After the algal body is harvested, the filamentous structure can be quickly dried by vacuum or warm air, and then ground into powder by an automatic grinder. Take 2 · 4 g of powder, add 70 ml of 1 OmM potassium phosphate The solution or other acid-base buffer should be fully stirred to maintain the solution's P 5 value between 5 and 10, and then fully stirred, and then centrifuged at 6 0 rpm, 10 mi η, 4 ° C, can be A clear claret protein solution was obtained. The concentration and purification of algal pigment can be removed by 20% ammonium sulfate solution to remove part of the impurity protein precipitation, and then the pigment protein is precipitated in the solution by 60% ~ 65% ammonium sulfate solution, which is OD 56 5 / OD28G = 2.66's thick red pigment is a food-grade, cosmetic-useable pigment. The above-mentioned clarified deep red algae protein solution can also be obtained by directly grinding the wet algal body and adding potassium phosphate solution, and then centrifuging and obtaining. The concentration and purification of the algal pigment can be performed by ultrafiltration. 4 · The precipitated protein is further purified by gel filtration chromatography (ge 1 filtration), and separated by 120 cm long Sephadex G200 tree®. The first procedure can obtain a phycoerythrin product with an OD ratio of 4.5 or more, Repeatedly again to obtain an OD of 5.3 or more phycoerythrin, and the results obtained by gel electrophoresis (SDS) electrophoresis showed a purity of about 99%, which can be used as reagents for immunoassay. Fifth A to C are chromatograms of 280 nm, 5 65 nm, and 615 nm of pure phycoerythrin extracted from the hair dish by high performance liquid chromatography (HPLC) *

Page 19 200408707 V. Description of the invention (15) ^ ~ Spectrum U Γ: The figure shows the analytical spectrum of the pure quality extracted from P. annua. Actual J instrument === of 2 ^ m / 5 6 5 nm, 6l5nm < i which were subjected to high performance liquid chromatographyΪ = algae red extracted from the same table species / System d: The seven A to C diagrams are the chromatographic spectra of the monthly 丄 3ηΠ1 obtained from the 28/50 ", 5 6 5nm commercial performance liquid chromatograph on the date of issue. No. 4 of the weight (mg) of laver and the laverycin in the present invention, the unit weight of algae ... 4: "" Vein ", we can clearly know the ratio of bath red pigment which can produce radon. The second and third columns are from the algae species. Second, it contains the absorbance ratio (〇D 56 5 / 〇D28 °) and (0D615 / 0D 565). It means that the κ-introducer ^ indicates the ratio of erythrocyanin, and the higher (OD565 / 〇D28Q) ratio indicates that the algae such as' go :: and lacquer red have higher purity values, the more food or cosmetic pigments are used, the more High, its added value is purified, because it can be applied to medical treatment: 'the more favorable the back-end phycocyanin will absorb the fluorescein emitted by the phycoerythin = 1 reagent; and because the cosmetic pigment is used for fluorescent The poor color rendering, 0 h as the content of food or phycocyanin is less, the lower the color rendering effect / 0D 565), it can be clearly seen that the long-leaf laver, bean grave of the present invention. Similarly, our absorbance ratio. Therefore, using the present invention = extracting phycoerythrin has high ^ bright long-leaf laver filaments to extract

200408707 V. Explanation of the invention (16) Taking phycoerythrin, it has the advantage of higher phycoerythrin content and higher absorbance than the filaments of the common hairy dish and laver. Table 2 Analysis of algae pink pigment of each algae species Algae species B a Pa Pd RPE mg / g Algal powder 53.5 38. 89 44.98 Absorption ratio A565 / A280 1.40 1. 54 1.96 Absorption ratio A615 / A565 0.19 0.53 0.21

RPE: phycoerythrin, which is hydrophilic and forms a stable aqueous solution with water. Its maximum UV absorption wavelength and fluorescence emission wavelength are 566nm and 5 7 5 nm, respectively.

Ba: Bangia atropurpurea

Pa ·· Porphyra angusta

Pd: Porphyra dentata

In summary, the embodiments of the present invention can achieve the expected effect, and the specific structure disclosed is not only not seen in similar products, nor disclosed before the application, and has fully complied with the provisions of the Patent Law and Requirement, apply for an invention patent in accordance with the law, and beg for examination and grant the patent,

P.21 200408707 V. Explanation of the invention (17) is a real sense. Although the present invention has been described by citing a preferred embodiment, the invention is not limited to the illustrated embodiment. Although specific examples have been cited and described previously, it is obvious that other equivalent changes or modifications made without departing from the spirit disclosed by the present invention should be included in the scope of patent application of the present invention. In addition, all other similar and approximate changes or modifications made without departing from the spirit disclosed by the present invention are also included in the scope of patent application of the present invention. At the same time, the scope of the present invention should be construed in the broadest definition so as to encompass all modifications and similar structures.

Page 22, 200408707 Brief description of the drawings Brief description of the drawings: In order to make the technical means, inventive features, objectives and effects of the present invention easy to understand, the detailed description with the drawings and the symbols is as follows: The chromatographic spectrum of UV absorption and fluorescence emission of phycocyanin obtained by high-performance liquid chromatography (HPLC) is shown in the figure. Leaf laver life history;

The fourth picture shows the life history of the sea surface; the fifth pictures A to C show the 280nm, 565nm, and 615nm chromatograms obtained from the high-performance liquid chromatograph of the pure phycoerythrin extracted from the hair dish. Spectrum. The sixth A to C diagrams show the chromatographic spectra of 280 nm, 5 6 5 nm, and 6 1 5 nm obtained from high-performance liquid chromatography of pure phycoerythrin extracted from P. angustifolia. Illustration.

The seventh A to C diagrams show the chromatographic spectra of 2 8 0 n m, 5 6 5 n m, and 6 1 5 n m obtained from high-performance liquid chromatography with purified phycoerythrin extracted from P. longifolia.

Page 23 200408707 Simple illustration of the diagram. IHiil

Claims (1)

200408707 VI. Scope of patent application Patent scope: 1. A new application of algae, whose life history has sexual, asexual, and vegetative reproduction alternated three generations, using its spore filaments to produce high absorbance ratios Phycoerythrin. 2. The new use of the algae species as described in item 1 of the scope of the patent application, wherein the spore filaments are fruit spores (c a r ρ 〇 s ρ 〇 ere). 3. The new use of the algae species as described in item 2 of the patent application scope, wherein the algae species is Porphyra dentata, and the phycoerythrin extracted from the fruit spore filaments is subjected to high performance liquid chromatography The chromatographic spectrum of 5 6 5 nm obtained by the instrument is shown in Figure 7 of the patent specification. 4. The new application of P. latifolia according to item 3 of the patent application scope, wherein the phycophyllin of P. longifolia fruit spores to produce a high absorbance ratio of phycoerythrin comprises the following steps: The gametophyte of the fruit sporangia is matured and fruit spores are obtained; the fruit spores are cultured at 15-30 ° C, 50 0 lux to 6 0 0 0 lux, with an illumination and light-dark ratio of 10: 14 or more , Make it grow into filaments; collect the filaments; add acid test buffer, mix and centrifuge to get dark pigment Page 25 200408707 6. Scope of patent application Protein solution; and salting-out method to remove impurities in the dark pigmented protein solution and precipitate excellent protein. 5. For example, the new application of Porphyra longifolia in item 4 of the patent application scope, wherein the culture medium is SWM-III culture medium. 6. For example, the new application of Porphyra longifolia in the scope of patent application No. 5, wherein the SWM-II culture solution is a SWM-II culture solution without organic matter. 7. The new application of Porphyra latifolia according to item 4 of the patent application, wherein the method for cultivating the fruit spore filaments is a rotary float culture method. 8. For the new application of P. longifolia in the scope of the patent application, the culture environment is 20 ° C, and the illumination and light-dark ratio of 2000 lux is 12:12. 9. According to the new application of P. longifolia in the scope of patent application, the step of collecting the filaments further comprises: collecting the filaments by a mesh collection method; drying the filaments; and grinding the filaments To make it powder. 10. For the new application of Porphyra latifolia in item 9 of the scope of patent application, the net collection method uses 20 to 400 net mesh. Page 26 200408707 6. Scope of Patent Application 11. For the new application of Porphyra latifolia in item 9 of the scope of patent application, the method of drying the filaments is vacuum method. 12. For the new application of Porphyra longifolia in item 9 of the scope of patent application, the condition for drying the filaments is the warm air method. 13. As for the new application of P. longifolia in item 4 of the scope of patent application, the conditions for centrifugation are a rotation speed of 6 0 0 r p m, a time of 10 m i η, and a temperature of 4 ° C. 14. For example, the new application of Porphyra longifolia in item 4 of the scope of patent application, wherein the acid buffer solution is an acid bell solution to maintain the p 维持 value of the solution between 5-10. 15. For the new application of Porphyra latifolia in item 4 of the patent application scope, the removal of impurities by salting out method is to add 20% ammonium sulfate solution. 16. As for the new application of Porphyra longifolia in the scope of the patent application, the pigmentation protein is precipitated by salting out method by adding 60% ~ 65% ammonium sulfate solution. 200408707 6. Scope of patent application 19. If the new application of Porphyra longifolia in item 18 of the scope of patent application is applied, the gel filtration chromatography method is a gel filtration chromatography method using Se p h a d e X G 2 0. 2 0 · A kind of phycoerythrin with carp 〇sp 〇re spores of two generations of father-in-law generations with sexual reproduction, asexual reproduction and vegetative reproduction in life history to produce phycoerythrin with high absorbance The device includes: a first: a culture tank having culture medium for cultivating gametophyte containing mature fruit sporangia of Porphyra dentata and obtaining fruit spores; A second culture tank 'has an illuminance and light-dark ratio of 500: lux ~ 6, 0,0 lux of 10:14 or more and 15 ~ 3 ° C, and is used to cultivate the fruit spores so that It grows into filaments; one collects f 'to collect the filaments; two grinds f' to grind the filaments; <«# @ 'Add acid and alkali buffer solution and grind the filaments, To obtain a dark pigmented protein solution; a precipitator, the dark pigmented protein solution precipitates phycoerythrin. 2 1. Equipment such as the patent application “m”, in which the fruit spore filaments of phycoerythrin and laver are produced from the high-absorbance ratio of long leaf through high-performance liquid chromatography instructions in the seventh figure B of the high-performance liquid chromatography specification. The chromatographic spectrum of 5 6 5nm obtained from the phycoerythrin extracted in the middle is as patent 2 2. & + #Patent Range No. 20 Use of Long-Leaf Laver Filament to Produce Height Page 28 200408707 VI. Application scope Patented phycoerythrin equipment with absorbance ratio, where the culture solution is swm-m culture solution. 2 3. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments as described in item 22 of the scope of patent application, wherein the SWM-IIII culture solution is an organic-free SWM-III culture solution. 2 4. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments, such as item 20 in the scope of patent application, wherein the method for cultivating the fruit spore filaments is a rotary float culture method. 25. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments as in item 20 of the patent application scope, wherein the culture environment is 20 ° C, and the illumination and brightness ratio of 2000 lux For 12: 12. 2 6. The device for producing phycoerythrin with high absorbance ratio using long-leaf laver filaments as in item 20 of the scope of patent application, wherein the collector collects the filaments with a mesh cloth. 27. For example, a device for producing phycoerythrin with high absorbance ratio using long-leaf laver filaments, such as item 26 in the scope of patent application, wherein the net cloth is a net mesh of 20 to 400. 28. Use of long-leaf laver filaments to produce height as in the scope of patent application No. 20 200408707 6. Scope of patent application The equipment of phycoerythrin with absorbance ratio, wherein the grinder further includes a dryer. 29. The equipment for producing phycoerythrin with high absorbance ratio using long-leaf laver filaments as in item 28 of the patent application scope, wherein the dryer is a vacuum dryer. 30. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments according to item 28 of the patent application scope, wherein the dryer is a warm air dryer. 3 1. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments as in item 20 of the scope of patent application, wherein the precipitator is the salt precipitation method Shendian phycoerythrin. 32. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments according to item 31 of the scope of patent application, wherein the phycoerythrin precipitated by the salting out method is added with 60% ~ 65% ammonium sulfate Solution. Page 30, 200408707 6. Scope of patent application 33. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments, such as item 20 of the scope of patent application, further includes a purifier. The phycoerythrin was purified by chromatography. 3 4. The equipment for producing phycoerythrin with high absorbance ratio using filamentous laver filaments as described in item 33 of the scope of patent application, wherein the gel filtration chromatography method is the gel filtration and chromatography method using Sephadex G200. 35. The device for producing phycoerythrin with high absorptivity ratio using filamentous laver filaments as claimed in item 20 of the scope of patent application, which further comprises a purifier, by ultrafiltration (u 11 rafi 1 trati ο η ) Purify the phycoerythrin. Page 31