CN115448992B - Oil tea fruit chitin extraction method and application thereof - Google Patents
Oil tea fruit chitin extraction method and application thereof Download PDFInfo
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Abstract
The invention provides a method for extracting camellia oleifera fruit shell polysaccharide, which comprises the steps of fully drying camellia oleifera fruit shells, crushing and sieving by a flash extractor to obtain camellia oleifera fruit shell powder, heating and extracting the camellia oleifera fruit shell powder by distilled water, and centrifugally retaining supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, precipitating with an organic solvent, centrifuging to retain precipitate, dissolving the precipitate with distilled water, decolorizing with active carbon, centrifuging, filtering the supernatant, and lyophilizing to obtain the oil tea fruit chitin. The camellia oleifera fruit chitin prepared by the method has the advantages of high yield and content, simple preparation process, low cost and good immunocompetence.
Description
Technical Field
The invention belongs to the field of extraction and application of natural active ingredients, and particularly relates to an extraction method and application of camellia oleifera chitin.
Background
The camellia oleifera (Camellia oleifera Abel) belongs to the camellia family, the camellia genus small arbor, is a first large woody oil crop which is mainly developed in China, and has a long planting history in the middle and downstream regions of the Yangtze river. The oil tea can produce a large amount of byproducts in the process of processing oil, and the oil tea fruit shell is taken as one of main oil extraction residues, accounts for about 60% of the weight of the oil tea fruit, and mainly comprises lignin, cellulose, hemicellulose, polysaccharide, flavonoids, phenols and terpenes. At present, the development and utilization of high added value of the camellia oleifera shells are not common.
Immunization is an important barrier for the body against invasion of external pathogenic microorganisms and pathogens, and macrophages play an important role in immune response. RAW264.7 cells, which are a type of macrophage, are commonly used as model cells for research and development of immunocompetent substances. Currently, many researches on extraction of natural plant active polysaccharide and regulation of organism immunity are reported, and some products have important influence on human health and economic and social development. However, the research on the extraction and application of the oil tea fruit shell immunocompetence substance is reported recently. Therefore, the extraction, development and application of immune components of the camellia oleifera shells are of great significance for improving the added value of products.
Disclosure of Invention
The invention aims to provide an extraction method and application of camellia oleifera chitin, wherein camellia oleifera chitin is prepared by taking camellia oleifera hulls as RAW materials through the steps of crushing by a flash extractor, sieving, heating and extracting by distilled water, centrifuging, concentrating under reduced pressure, precipitating by an organic solvent, decoloring by activated carbon, freeze-drying and the like, and the obtained camellia oleifera chitin is subjected to an immune activity test, so that the obtained camellia oleifera chitin can stimulate RAW264.7 cell immune molecules to generate, enhance immune protein expression, promote TNF alpha release and mRNA expression, activate immune related signal channels and has weak influence on cell morphology and survival rate. The camellia oleifera fruit chitin prepared by the method has the advantages of high yield and content, simple process, low cost and good immunocompetence.
In order to achieve the above purpose, the invention adopts the following technical scheme: the method for extracting the camellia oleifera fruit shell polysaccharide comprises the following steps of:
(1) Fully drying oil tea shells, crushing by a flash extractor, sieving, heating and extracting by distilled water, and centrifuging to keep supernatant;
(2) Concentrating the supernatant under reduced pressure by using a rotary evaporator, precipitating by using an organic solvent, and centrifugally retaining the precipitate;
(3) Dissolving the precipitate with distilled water, decolorizing with active carbon, centrifuging to retain supernatant, suction filtering the supernatant, and lyophilizing to obtain camellia oleifera chitin.
Further preferably, the moisture content of the dried camellia oleifera shells is 15% -20%.
Further preferably, the flash extractor parameters are set to 4000-6000 rpm/min and the extraction time is 1-3 min.
Further preferably, the sieving specification is 80-100 mesh.
Further preferably, the mass-volume ratio of the oil tea fruit shell powder to the distilled water is 1:20-40, and the heating time is 8-12h.
Further preferably, the rotary evaporator parameters are set to 40-60bar,55-65℃and 50-65rpm.
Further preferably, the concentration under reduced pressure is carried out so that the concentration after concentration should reach a density of 1.1 to 1.3g/cm 3.
Further preferably, the organic solvent is selected from one of methanol, ethanol or acetone.
Further preferably, the volume ratio of the concentrated solution to the organic solvent is 1:4-6.
Further preferably, the organic solvent precipitation is carried out by keeping the temperature at 4 ℃ and standing for 12-18 hours.
Further preferably, the mass volume ratio of the sediment to the distilled water is 1:10-30.
Further preferably, the mass ratio of the activated carbon to the crude polysaccharide is 1:4-9.
Further preferably, when the activated carbon is decolorized, constant-temperature water bath heating is adopted, the temperature is 50-60 ℃ and the time is 2-4 hours;
the application of the camellia oleifera chitin is that the obtained camellia oleifera chitin is used for preparing a medicament for enhancing RAW264.7 cell immune response.
The invention has the following beneficial effects: according to the invention, camellia oleifera shells are used as RAW materials, and the steps of crushing by a flash extractor, sieving, heating and extracting by distilled water, concentrating under reduced pressure, precipitating by an organic solvent, decoloring by activated carbon, freeze drying and the like are adopted to prepare camellia oleifera shell polysaccharide, and an immunocompetence test is carried out by a RAW264.7 cell model, so that the camellia oleifera shell polysaccharide can stimulate NO and PGE2 to generate, enhance iNOS and COX2 protein expression, promote TNFa release and mRNA expression, up-regulate MAPKs signal proteins ERK, P38 and JNK phosphorylation levels, promote NF- κB nuclear translocation, has weak influence on cell morphology and survival rate, and is beneficial to being applied to preparation of medicaments for enhancing RAW264.7 cell immunoreaction. The camellia oleifera fruit chitin prepared by the method has the advantages of high yield and content, simple preparation process, low cost and good immunocompetence.
Drawings
FIG. 1 is a graph showing the data of the effect of oil tea Chitin (CPS) on cell morphology in example 1 of the present invention;
FIG. 2 is a graph showing the data of the effect of oil tea Chitin (CPS) on cell viability in example 1 of the present invention;
FIG. 3 is a graph showing the data of the effect of oil tea Chitin (CPS) on NO production in example 1 of the present invention;
FIG. 4 is a graph showing the data of the effect of oleanolic Chitin (CPS) on PGE2 production in example 1 of the present invention;
FIG. 5 is a graph showing the data of the effect of oleoresin on expression of iNOS protein in example 1 of the present invention;
FIG. 6 is a graph showing the data of the effect of oil tea Chitin (CPS) on COX2 protein expression in example 1 of the present invention;
FIG. 7 is a graph showing the data of the effect of oil tea Chitin (CPS) on TNF alpha release in example 1 of the present invention;
FIG. 8 is a graph showing the data of the effect of oil tea Chitin (CPS) on IL-6 release in example 1 of the present invention;
FIG. 9 is a graph showing data of the effect of oleoresin on IL-1β release by CPS in example 1 of the present invention;
FIG. 10 is a graph showing the data of the effect of oleoresin on expression of iNOS gene in example 1 of the present invention;
FIG. 11 is a graph showing the data of the effect of oil tea Chitin (CPS) on COX2 gene expression in example 1 of the present invention;
FIG. 12 is a graph showing the data of the effect of nectar Chitin (CPS) on TNF alpha gene expression in example 1 of the present invention;
FIG. 13 is a graph showing the data of the effect of oil tea Chitin (CPS) on IL-1. Beta. Gene expression in example 1 of the present invention;
FIG. 14 is a graph showing the data of the effect of oil tea Chitin (CPS) on IL-6 gene expression in example 1 of the present invention;
FIG. 15 is a graph showing the data of the effect of nectar Chitin (CPS) on GAPDH gene expression in example 1 of the present invention;
FIG. 16 is a graph showing the data of the effect of oleoresin (CPS) on the phosphorylation levels of MAPKs signaling pathway proteins ERK, P38 and JNK in example 1 of the present invention;
FIG. 17 is a graph of quantitative data for the effect of oleoresin (CPS) on ERK phosphorylation levels of MAPKs signaling pathway protein in example 1 of the present invention;
FIG. 18 is a graph of quantitative data for the effect of oleoresin (CPS) on the phosphorylation level of MAPKs signaling pathway protein P38 in example 1 of the present invention;
FIG. 19 is a graph of quantitative data of the effect of oleoresin (CPS) on the phosphorylation level of the MAPKs signal pathway protein JNK in example 1 of the present invention;
FIG. 20 is a data graph of the effect of oil tea Chitin (CPS) on NF- κB nuclear translocation in example 1 of the present invention.
Detailed Description
The invention will now be further illustrated with reference to specific examples, which are intended to be illustrative rather than limiting.
Example 1
Fully drying oil tea fruit shells to ensure that the water content of the oil tea fruit shells reaches about 20%, weighing 1kg of the oil tea fruit shells, putting the oil tea fruit shells into a flash extractor for crushing, sieving the crushed oil tea fruit shells with a 100-mesh sieve, adding distilled water with the volume of 20 times, heating the distilled water to 80 ℃, leaching the oil tea fruit shells for 8 hours, and centrifuging (4000 rpm,10 min) to keep supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to be 60bar, 55 ℃ and 50rpm, enabling the volume of the concentrated solution to be about 500mL, enabling the density to be 1.1g/cm 3, precipitating the concentrated solution by using 4 times of methanol, uniformly stirring, standing for 12h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 10 times volume of distilled water, adding 10% active carbon, heating to 50deg.C in water bath, maintaining for 2 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, freeze drying, detecting with phenol-sulfuric acid method to obtain oil tea fruit chitin with total sugar content of 37.1% and yield of 25.34%, and storing.
Accurately weighing 100mg of camellia oleifera fruit chitin, adding 10mL of distilled water for full dissolution, sterilizing by ultraviolet for 30min, and preserving at 4 ℃ for later use; diluting the oil tea fruit chitin mother liquor into 400, 200, 100, 50, 25 and 12.5 mug/mL with serum-free DMEM high-sugar culture solution according to a multiple ratio dilution mode, and preparing the oil tea fruit chitin mother liquor for use at present; culturing RAW264.7 cells to logarithmic growth phase, inoculating the cells into a 96-well plate overnight, wherein the density is 5x10≡4/hole, and the volume is 100 mu L/hole; treating the prepared camellia oleifera chitin and LPS (2 mug/mL) with the cells for 24h, wherein the cell is 100 mug/hole; taking a photograph through a microscope to display cell morphology, detecting the cell survival rate by an MTT method, and detecting NO production in cell supernatant by a Griess method; the results show that the camellia oleifera chitin has weak influence on cell morphology change and survival rate (refer to fig. 1 and 2), and can stimulate RAW264.7 cell NO production (refer to fig. 3).
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1x10≡6/well and a volume of 3 mL/well; treating the cells with chitin and LPS for 24 hours, detecting expression of iNOS and COX2 proteins by western blotting, detecting content of PGE2, TNF alpha, IL-6 and IL-1 beta in the cell supernatant by ELISA, and detecting mRNA levels of iNOS, COX2, TNF alpha, IL-6 and IL-1 beta by RT-PCR; the results show that camellia oleifera chitin can promote RAW264.7 cell PGE2 production (fig. 4), enhance iNOS and COX2 protein expression (fig. 5 and 6), and promote tnfα release (fig. 7 to 9) and mRNA expression (fig. 10 to 15).
RAW264.7 cells were cultured to log phase and inoculated in 60mm dishes overnight at a density of 1.5x10ζ6/well and a volume of 4 mL/dish; treating cells 1,2 and 6 hours by using oil tea fruit chitin and LPS, and detecting ERK, P38 and JNK protein expression and phosphorylation levels by western blotting; the results showed that camellia oleifera chitin can up-regulate the ERK, P38 and JNK protein phosphorylation levels of RAW264.7 cells (fig. 16 to 19).
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1x10≡5 per well and a volume of 3mL per well; treating cells for 6 hours by using camellia oleifera chitin and LPS, and analyzing NF- κB nuclear translocation by immunofluorescence technology; the results show that camellia oleifera chitin can promote NF- κb nuclear translocation of RAW264.7 cells (fig. 20).
Example 2
Fully drying oil tea fruit shells to ensure that the water content of the oil tea fruit shells reaches about 18%, weighing 1kg of the oil tea fruit shells, putting the oil tea fruit shells into a flash extractor for crushing, sieving the crushed oil tea fruit shells with a 100-mesh sieve, adding distilled water with the volume being 25 times that of the crushed oil tea fruit shells, heating the crushed oil tea fruit shells to 80 ℃, leaching the crushed oil tea fruit shells for 10 hours, and centrifuging (4000 rpm,10 min) to keep supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to be 50bar, 60 ℃ and 50rpm, enabling the volume of the concentrated solution to be about 500mL, enabling the density to be 1.2g/cm 3, precipitating the concentrated solution by using acetone with 5 times of volume, uniformly stirring, standing for 14h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 20 times of distilled water, adding 15% active carbon, heating to 55deg.C in water bath, maintaining for 3 hr, centrifuging, filtering supernatant, concentrating the filtrate under reduced pressure with rotary evaporator, freeze drying, detecting with phenol-sulfuric acid method to obtain oil tea fruit chitin with total sugar content of 38.5% and yield of 26.17%, and storing.
Accurately weighing 100mg of camellia oleifera fruit chitin, adding 10mL of distilled water for full dissolution, sterilizing by ultraviolet for 30min, and preserving at 4 ℃ for later use; diluting the oil tea fruit chitin mother liquor into 400, 200, 100, 50, 25 and 12.5 mug/mL with serum-free DMEM high-sugar culture solution according to a multiple ratio dilution mode, and preparing the oil tea fruit chitin mother liquor for use at present; culturing RAW264.7 cells to logarithmic phase, inoculating the cells into 96-well plates overnight at a density of 8x10ζ4/well and a volume of 100 μL/well; treating the prepared camellia oleifera chitin and LPS (2 mug/mL) with the cells for 24h, wherein the cell is 100 mug/hole; taking a photograph through a microscope to display cell morphology, detecting the cell survival rate by an MTT method, and detecting NO production in cell supernatant by a Griess method; the results show that the camellia oleifera chitin has weak influence on cell morphology and survival rate, and can stimulate the NO production of RAW264.7 cells.
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1.5x10ζ6/well and a volume of 3 mL/well; treating the cells with chitin and LPS for 24 hours, detecting expression of iNOS and COX2 proteins by western blotting, detecting content of PGE2, TNF alpha, IL-6 and IL-1 beta in the cell supernatant by ELISA, and detecting mRNA levels of iNOS, COX2, TNF alpha, IL-6 and IL-1 beta by RT-PCR; the results show that the camellia oleifera chitin can promote the production of PGE2 by RAW264.7 cells, enhance iNOS and COX2 protein expression, and promote tnfα release and mRNA expression.
RAW264.7 cells were cultured to log phase and inoculated in 60mm dishes overnight at a density of 1x10≡6/well and a volume of 4 mL/dish; treating cells 1,2 and 6 hours by using oil tea fruit chitin and LPS, and detecting ERK, P38 and JNK protein expression and phosphorylation levels by western blotting; the results show that the camellia oleifera chitin can up-regulate the ERK, P38 and JNK protein phosphorylation levels of RAW264.7 cells.
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1.5x10ζ5/well and a volume of 3 mL/well; treating cells for 6 hours by using camellia oleifera chitin and LPS, and analyzing NF- κB nuclear translocation by immunofluorescence technology; the result shows that the camellia oleifera chitin can promote NF- κB nuclear translocation of RAW264.7 cells.
Example 3
Fully drying oil tea fruit shells to enable the water content to reach about 15%, weighing 1kg of the oil tea fruit shells, putting into a flash extractor for crushing, sieving with a 100-mesh sieve, adding 30 times of distilled water, heating to 85 ℃, leaching for 12h, centrifuging (4000 rpm,10 min), and reserving supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to 40bar, 60 ℃ and 55rpm to enable the volume of the concentrated solution to be about 500mL, precipitating the concentrated solution with 6 times of volume of ethanol with the density of 1.3g/cm 3, uniformly stirring, standing at 4 ℃ for 15h, and centrifuging to obtain a precipitate; dissolving the precipitate with 30 times of distilled water, adding 20% active carbon, heating to 60deg.C in water bath, maintaining for 4 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, freeze drying, detecting with phenol-sulfuric acid method to obtain oil tea fruit chitin with total sugar content of 39.2% and yield of 27.83%, and storing.
Accurately weighing 100mg of camellia oleifera fruit chitin, adding 10mL of distilled water for full dissolution, sterilizing by ultraviolet for 30min, and preserving at 4 ℃ for later use; diluting the oil tea fruit chitin mother liquor into 400, 200, 100, 50, 25 and 12.5 mug/mL with serum-free DMEM high-sugar culture solution according to a multiple ratio dilution mode, and preparing the oil tea fruit chitin mother liquor for use at present; culturing RAW264.7 cells to logarithmic phase, inoculating the cells into 96-well plates overnight at a density of 1x10≡5/well and a volume of 100 μl/well; treating the prepared camellia oleifera chitin and LPS (2 mug/mL) with the cells for 24h, wherein the cell is 100 mug/hole; taking a photograph through a microscope to display cell morphology, detecting the cell survival rate by an MTT method, and detecting NO production in cell supernatant by a Griess method; the results show that the camellia oleifera chitin has weak influence on cell morphology and survival rate, and can stimulate the NO production of RAW264.7 cells.
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1x10≡6/well and a volume of 3 mL/well; treating the cells with chitin and LPS for 24 hours, detecting expression of iNOS and COX2 proteins by western blotting, detecting content of PGE2, TNF alpha, IL-6 and IL-1 beta in the cell supernatant by ELISA, and detecting mRNA levels of iNOS, COX2, TNF alpha, IL-6 and IL-1 beta by RT-PCR; the results show that the camellia oleifera chitin can promote the production of PGE2 by RAW264.7 cells, enhance iNOS and COX2 protein expression, and promote tnfα release and mRNA expression.
RAW264.7 cells were cultured to log phase and inoculated in 60mm dishes overnight at a density of 1.5x10ζ6/well and a volume of 4 mL/dish; treating cells 1,2 and 6 hours by using oil tea fruit chitin and LPS, and detecting ERK, P38 and JNK protein expression and phosphorylation levels by western blotting; the results show that the camellia oleifera chitin can up-regulate the ERK, P38 and JNK protein phosphorylation levels of RAW264.7 cells.
RAW264.7 cells were cultured to log phase and seeded overnight in 6-well plates at a density of 1x10≡5 per well and a volume of 3mL per well; treating cells for 6 hours by using camellia oleifera chitin and LPS, and analyzing NF- κB nuclear translocation by immunofluorescence technology; the result shows that the camellia oleifera chitin can promote NF- κB nuclear translocation of RAW264.7 cells.
The tea fruit chitin in the embodiment has the effects of stimulating NO and PGE2 production, enhancing iNOS and COX2 protein expression, promoting TNFa release and mRNA expression, up-regulating MAPKs signal proteins ERK, P38 and JNK phosphorylation level, promoting NF- κB nuclear translocation, and having weak influence on cell morphology and survival rate, and is beneficial to being applied to preparing medicaments for enhancing RAW264.7 cell immune response.
Comparative example
Fully drying oil tea fruit shells to ensure that the water content of the oil tea fruit shells reaches about 20%, weighing 1kg of the oil tea fruit shells, crushing, sieving with a 100-mesh sieve, adding distilled water with 20 times of volume, heating to 80 ℃, leaching for 8 hours, and centrifuging (4000 rpm,10 min) to keep supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to be 60bar, 55 ℃ and 50rpm, enabling the volume of the concentrated solution to be about 500mL, precipitating the concentrated solution by using 1 time of ethanol, uniformly stirring, standing for 12h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 10 times volume of distilled water, adding 25% active carbon, heating to 50deg.C in water bath, maintaining for 2 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, freeze drying, and detecting with phenol-sulfuric acid method to obtain oil tea fruit chitin with total sugar content of 27.9% and yield of 16.34%. According to the invention, the flash extractor is adopted for crushing, and the ratio of the concentrated solution to the organic solvent is adjusted when the organic solvent is precipitated, so that better extraction effect is obtained and the yield of the camellia oleifera chitin is greatly improved compared with the mode that the flash extractor is not adopted and the ratio of the concentrated solution to the organic solvent is 1:1.
The above disclosure is only a preferred embodiment of the present invention, and it should be understood that the scope of the invention is not limited thereto, but all or part of the procedures for implementing the above embodiments can be modified by one skilled in the art according to the scope of the appended claims.
Claims (4)
1. The application of the camellia oleifera fruit chitin is characterized in that the extraction of the camellia oleifera fruit chitin comprises the following steps:
(1) Fully drying the camellia oleifera shells, wherein the moisture content of the dried camellia oleifera shells is 15% -20%, crushing by a flash extractor, sieving, heating and leaching by distilled water, wherein the mass-volume ratio of camellia oleifera shell powder to distilled water is 1:20-40, the heating time is 8-12h, and centrifuging to retain supernatant;
(2) Concentrating the supernatant under reduced pressure by adopting a rotary evaporator to ensure that the density of the concentrated solution reaches 1.1-1.3g/cm 3, precipitating by using an organic solvent, uniformly stirring, standing at 4 ℃ for 12-15h, centrifuging to keep the precipitate, wherein the organic solvent is selected from any one of methanol, ethanol or acetone, and the volume ratio of the concentrated solution to the organic solvent is 1:4-6;
(3) Dissolving the precipitate with distilled water, decolorizing with active carbon, heating to 50-60deg.C in water bath, maintaining for 2-4 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, and freeze drying to obtain oil tea fruit chitin;
the obtained oil tea fruit chitin is used for preparing medicines for promoting PGE2 production, enhancing iNOS and COX2 protein expression, promoting mRNA expression, and up-regulating MAPKs signaling proteins ERK, P38 and JNK phosphorylation level.
2. The use of camellia oleifera chitin according to claim 1, characterized in that the camellia oleifera chitin extraction process is: fully drying oil tea fruit shells to ensure that the water content is 20%, weighing 1kg of oil tea fruit shells, putting into a flash extractor for crushing, sieving with a 100-mesh sieve, adding distilled water with 20 times of volume, heating to 80 ℃, leaching for 8 hours, centrifuging, and reserving supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to be 60bar, 55 ℃ and 50rpm, enabling the volume of the concentrated solution to be 500mL and the density to be 1.1g/cm 3, precipitating the concentrated solution by using 4 times of methanol, uniformly stirring, standing for 12h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 10 times volume of distilled water, adding active carbon, heating to 50deg.C in water bath, maintaining for 2 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, and freeze drying to obtain oil tea fruit chitin.
3. The use of camellia oleifera chitin according to claim 1, characterized in that the extraction process of camellia oleifera chitin is: fully drying oil tea fruit shells to enable the water content to be 18%, weighing 1kg of oil tea fruit shells, putting into a flash extractor for crushing, sieving with a 100-mesh sieve, adding distilled water with 25 times of volume, heating to 80 ℃, leaching for 10h, centrifuging, and reserving supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to be 50bar, 60 ℃ and 50rpm, enabling the volume of the concentrated solution to be 500mL, enabling the density to be 1.2g/cm 3, precipitating the concentrated solution by using acetone with 5 times of volume, uniformly stirring, standing for 14h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 20 times of distilled water, adding active carbon, heating to 55deg.C in water bath, maintaining for 3 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, and freeze drying to obtain oil tea fruit chitin.
4. The use of camellia oleifera chitin according to claim 1, characterized in that the extraction process of camellia oleifera chitin is: fully drying oil tea fruit shells to enable the water content to be 15%, weighing 1kg of oil tea fruit shells, putting into a flash extractor for crushing, sieving with a 100-mesh sieve, adding 30 times of distilled water, heating to 85 ℃, leaching for 12 hours, centrifuging, and reserving supernatant; concentrating the supernatant under reduced pressure by using a rotary evaporator, setting parameters to 40bar, 60 ℃ and 55rpm to make the volume of the concentrated solution be 500mL and the density be 1.3g/cm 3, precipitating the concentrated solution by using 6 times of ethanol, uniformly stirring, standing for 15h at 4 ℃, and centrifuging to obtain a precipitate; dissolving the precipitate with 30 times of distilled water, adding active carbon, heating to 60deg.C in water bath, maintaining for 4 hr, centrifuging, filtering supernatant, concentrating filtrate under reduced pressure with rotary evaporator, and freeze drying to obtain oil tea fruit chitin.
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| CN202211236364.0A CN115448992B (en) | 2022-10-10 | Oil tea fruit chitin extraction method and application thereof |
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Non-Patent Citations (5)
| Title |
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| The polysaccharide from Camellia oleifera fruit shell enhances immune responses via activating MAPKs and NF-κB signaling pathways in RAW264.7 macrophages;Chuanqi Xie等;《Food & Nutrition Research》;第66卷;第1-10页 * |
| 沈建福等.油茶果壳多糖的提取及抗氧化作用研究.《中国粮油学报》.2010,第25卷(第8期),第51-54页. * |
| 油茶果壳多糖的提取及抗氧化作用研究;沈建福等;《中国粮油学报》;第25卷(第8期);第51-54页 * |
| 油茶果壳多糖的提取工艺及免疫活性研究;林欣颖等;《生物化工》;第8卷(第5期);第24-28、59页 * |
| 油茶酸性多糖OTAPS-1的分离纯化及其免疫调节活性研究;黎青等;《中国化学会第十二届全国天然有机化学学术会议论文摘要集》;第553页 * |
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