DENDROBIUM OFFICINALE OLIGOSACCHRIDE, DENDROBIUM OFFICINALE OLIGOSACCHRIDE DERIVATIVE, AND METHOD OF MANUFACTURING THEM AND USES THEREOF

Description

CROSS REFERENCE TO RELATED APPLICATION

[0001] This patent application claims the benefit and priority of Chinese Patent Application No. 202110247117.X filed on March 05, 2021, entitled "Dendrobium officinale oligosaccharide, Dendrobium officinale oligosaccharide derivative and preparation method and use thereof".TECHNICAL FIELD

[0002] The present disclosure relates to the technical field of natural medicine chemistry, and in particular to an oligosaccharide from Dendrobium officinale, a derivative of an oligosaccharide from Dendrobium officinale and a preparation method and use thereof.BACKGROUND ART

[0003] Dendrobium officinale (Dendrobium officinale Kimura et Migo) is a perennial herb of the genus Dendrobium in the family Orchidaceae, which is listed in each edition of the Pharmacopoeia. According to the records in the Pharmacopoeia, it has the effects of such as engendering liquid and nourishing stomach, nourishing yin and clearing heat, moistening lung and boosting kidney, and brightening eyes and strengthening lumbus. Modern pharmacological research shows that Dendrobium officinale has antioxidative, hypoglycemic, immunomodulatory and anti-inflammatory activities. It is recorded in ancient literatures that the stem of Dendrobium officinale can be used for moisturizing skin and treating diabetes. Studies have shown that Dendrobium officinale mainly contains polysaccharides, stilbene, flavonoids, phenanthrene, lignin and other chemical components.

[0004] Aging is an inevitable result of human growth, and the way to delay aging has always been the ultimate proposition in the fields of skin care products and biomedicine. Aging is roughly divided into two categories: physiological aging and pathological aging. Physiological aging refers to the physiological degeneration process that occurs after the mature period, and pathological aging is the senility changes caused by various external factors (including various diseases). The two are actually difficult to distinguish. It is recorded in ancient literatures that Dendrobium officinale has the anti-aging effects, and the natural, green, safe and efficient anti-aging substances in Dendrobium officinale are thus the focus of research.

[0005] At present, many domestic and foreign researchers have conducted research on the extraction and separation of chemical components in various parts of Dendrobium officinale. Most of the researchers mainly obtain small-molecule phenol components or polysaccharides with a molecular weight of more than 100,000. Some researchers have isolated 20 compounds from Dendrobium officinale, mainly bibenzil components, and have found that the small-molecule phenol components of Dendrobium officinale have antioxidative, anti-inflammatory and anti-tumor effects through activity screening. Some researchers have extracted and separated two polysaccharides from Dendrobium officinale with a molecular weight of 7.4×10 5< Da and 5.4×10 5< Da, respectively. Some researchers have used a method of water extraction, alcohol precipitation and sulfuric acid hydrolysis to obtain polysaccharides from Dendrobium officinale, and have found that the polysaccharides of Dendrobium officinale influence the count of peripheral white blood cells in mice through activity screening, indicating an immunomodulatory effect. Some researchers have obtained polysaccharides from Dendrobium officinale by ultrasonic extraction and have found that the polysaccharides of Dendrobium officinale influence the SOD, MDA and CAT of Drosophila through activity screening, indicating an antioxidative activity. Some researchers have demonstrated that there are mainly four homogeneous polysaccharides among polysaccharides of Dendrobium officinale, with a molecular weight in a range of 6,800-1.78×10 5< Da. The previous research work mostly concentrates on the study of the pharmacological activity of the small-molecule phenol components and the polysaccharides, whereas there are few reports on the research of oligosaccharides.

[0006] CN 109295131 A discloses an oligosaccharide from Dendrobium officinale having seven glycosidic residues and a glucose residue at the non-reducing end.

[0007] CN 108635300 A discloses a polysaccharide from Dendrobium officinale having a small molecular.

[0008] Zhang et al. (Organic & Biomolecular Chemistry 15, 2017, 1222-1227) discloses a pnitrophenol derivative of a heptasaccharide having glucuronic acid at the reducing end.SUMMARY

[0009] In view of this, the present disclosure provides an oligosaccharide from Dendrobium officinale, a derivative of an oligosaccharide from Dendrobium officinale and a preparation method and use thereof. It is illustrated that the oligosaccharide from Dendrobium officinale has anti-aging and immunomodulatory effects, and is suitable for applying to skin cosmetics, health foods and medicines with anti-aging and immunomodulatory effects.

[0010] In order to achieve the above objectives, the present disclosure mainly provides the following technical solutions: The present disclosure provides an oligosaccharide from Dendrobium officinale, having a structure represented by formula (I):

[0011] The present disclosure provides a derivative of an oligosaccharide from Dendrobium officinale, where the derivative is generated by reacting the glucuronic acid residue of the oligosaccharide from Dendrobium officinale according to the above technical solution with a hydrophobic residue, and the hydrophobic residue includes any one of linear or branched C 1 -C 18 alkyl, C 1 -C 18 alkyl carboxylic acid, C 1 -C 18 alkyl amine, C 1 -C 18 aryl alkyl and C 1 -C 18 alkyl amide, or the hydrophobic residue includes any one of benzoyl, phenylacetyl and phenylpropionyl.

[0012] The present disclosure provides the oligosaccharide from Dendrobium officinale or the derivative according to the above technical solutions for use in preparation of anti-aging or immunomodulatory products.

[0013] In some embodiments, the products include medicines, foods, formulations and cosmetics.

[0014] In some embodiments, a type of the cosmetics includes a face cream, an emulsion, a toning lotion or a repair cream.

[0015] The present disclosure provides a method for extracting the oligosaccharide from Dendrobium officinale described in above technical solutions, comprising: 1) preparation of a Dendrobium officinale powder: drying a stem of Dendrobium officinale, then smashing and sieving to obtain a Dendrobium officinale powder; 2) preparation of a water extract of Dendrobium officinale: mixing the Dendrobium officinale powder with water, then subjecting to a boiling water bath for one or more times, and then merging water extracts to obtain a water extract of Dendrobium officinale; 3) preparation of a concentrated water extract of Dendrobium officinale: centrifuging the water extract of Dendrobium officinale, and then concentrating under reduced pressure to obtain a concentrated water extract of Dendrobium officinale; 4) preparation of a redissolving liquid: adding ethanol to the concentrated water extract of Dendrobium officinale, precipitating, centrifuging and then discarding a supernatant to obtain a precipitate; adding water to the obtained precipitate for redissolving to obtain a redissolving liquid; 5) preparation of an aqueous solution of the oligosaccharide from Dendrobium officinale: deproteinizing the redissolving liquid, and freeze-drying to obtain a crude polysaccharide of Dendrobium officinale; adding water to dissolve the crude polysaccharide of Dendrobium officinale, then adding cellulase for a reaction, followed by inactivating and centrifuging, and discarding a precipitate to obtain an aqueous solution of the oligosaccharide from Dendrobium officinale; 6) preparation of the oligosaccharide from Dendrobium officinale: purifying the aqueous solution of the oligosaccharide from Dendrobium officinale to obtain the oligosaccharide from Dendrobium officinale.

[0016] In some embodiments, in step 1), a temperature for the drying is 50-60°C; a moisture mass content of the stem of Dendrobium officinale after the drying is less than 6%; a mesh number of the sieving is 20-40 meshes; in step 2), the Dendrobium officinale powder and water are mixed in a mass-volume ratio of 1 : (50-80) g / mL; the boiling water bath is performed for 2-3 times; a time for each boiling water bath is 1-2 h; in step 3), the centrifugation is conducted under conditions of 3000-4000 rpm / min, 8-12 min and room temperature; the concentration under reduced pressure is conducted under conditions of a rotary evaporation method, an ethanol recycling temperature of less than 50°C, and a pressure of 0.7 Pa-0.9 Pa; a material-liquid mass-volume ratio of the concentrated water extract of Dendrobium officinale is 1 : (5-10) g / mL.

[0017] In some embodiments, in step 4), the addition of ethanol to the concentrated water extract of Dendrobium officinale and the precipitation are specifically conducted as follows: adding ethanol with a volume concentration of 95% to the concentrated water extract of Dendrobium officinale until an alcohol concentration of a mixture is 70%-80% in volume percentage, and then precipitating overnight; the centrifugation is conducted under conditions of 3500-4500 rpm, 18-22 min and room temperature; a weight-volume ratio of the precipitate and water for redissolving is 1 : (4-6) g / mL; in step 5), the deproteinization of the redissolving liquid is conducted by a Sevage method; a temperature for the freeze-drying is 45°C; the addition of water to dissolve the crude polysaccharide of Dendrobium officinale is conducted as follows: adding water to the crude polysaccharide of Dendrobium officinale for dissolving until 4.5-5.5 mg / mL; an added amount of the cellulase is 2%-4% of a weight of the crude polysaccharide of Dendrobium officinale; after adding the cellulase, the reaction is conducted under a reaction temperature of 55-65°C for a reaction time of 2 h-4 h; the inactivation is conducted by boiling for 25-35 min; the centrifugation is conducted under conditions of 3500-4500 rpm, 15-25 min and room temperature; an oligosaccharide content in the aqueous solution of oligosaccharide from Dendrobium officinale is greater than 80%; in step 6), the purification is conducted by passing through a G75 gel column; a purity of the oligosaccharide from Dendrobium officinale is 99.86%.

[0018] The present disclosure provides a method for preparing the derivative described in the above technical solutions, comprising: 1) reducing a fatty acid into a fatty alcohol; 2) glycosylating the fatty alcohol with the oligosaccharide from Dendrobium officinale according to the above technical solutions to obtain the derivative.

[0019] In some embodiments, in step 1), a reaction process of the reduction of a fatty acid into a fatty alcohol is as follows: mixing 9-11 mmol of the fatty acid, 18-22 mmol of NaBH 4 , 38-42 mmol of AlCl 3 , 25-29 ml of THF, and 2-4 ml of MeOH for a reaction, and the reaction is conducted under a temperature of 95-105°C for a time of 7.5-8.5 h with a yield of 88-92%; in step 2), a reaction process of the glycosylation is as follows: mixing the alcohol and the saccharide in a molar ratio of 1 : (8-10), and then using cetyl ammonium bromide as a catalyst for catalyzation, and the reaction is conducted under a temperature of 110-115°C for a time of 2.5-3.5 h with a yield of 78-82%.

[0020] Compared with the prior art, the oligosaccharide from Dendrobium officinale, the derivative and the preparation method and use thereof in the present disclosure have at least the following beneficial effects:

[0021] In one aspect, the present disclosure illustrates for the first time the oligosaccharides with a purity of more than or equal to 99% prepared from Dendrobium officinale, and carries out structural analysis to identify the structures of the above oligosaccharides from Dendrobium officinale.

[0022] Further, the present disclosure conducts for the first time a systematic study on the anti-aging activity of the above oligosaccharides from Dendrobium officinale, and has found that the above oligosaccharides from Dendrobium officinale can well inhibit collagenase and the like and thus have a strong anti-aging effect. Moreover, they also have a certain activity on TNF-α and IL-6, and a good immunomodulatory effect.

[0023] In another aspect, the preparation method and active application of the above oligosaccharides from Dendrobium officinale involved in the present disclosure provide a scientific basis for the application of Dendrobium officinale in related directions.

[0024] In yet another aspect, the oligosaccharide from Dendrobium officinale prepared by the present disclosure can be used as an intermediate for the development of Dendrobium officinale series products (such as application in the fields of medicine, food, daily necessities, etc.).BRIEF DESCRIPTION OF THE DRAWINGS

[0025] FIGs. 1-3 show the structure and purity identification HPLC-ELSD, NMR and MS spectra of the oligosaccharides from Dendrobium officinale of the present disclosure: FIG. 1 shows the purity identification HPLC-ELSD spectrum of the oligosaccharides from Dendrobium officinale; FIG. 2 shows the 1< H-NMR and 13< C-NMR spectra of the oligosaccharides from Dendrobium officinale, where FIG. 2A is a 1< H-NMR spectrum, and FIG. 2B is a 13< C-NMR spectrum; FIGs. 3-6 show the test results of the anti-aging activity of the oligosaccharides from Dendrobium officinale: FIG. 3 shows the anti-oxidation rate of the oligosaccharides from Dendrobium officinale based on the scavenging rate of DPPH free radicals (oligosaccharide concentrations of 4%, 8%, 11%, 15%, 20%); where the ordinate is the anti-oxidation rate % represented by the scavenging rate of DPPH free radicals, and the abscissa (from 1 to 5) corresponds to the oligosaccharide concentration of 4%, 8%, 11%, 15%, 20%, respectively; FIG. 4 shows the inhibition rate of the oligosaccharides from Dendrobium officinale on elastase activity (oligosaccharide concentrations of 4%, 8%, 11%, 15%, 20%); where the ordinate is the elastase inhibition rate %, and the abscissa (from 1 to 5) corresponds to the oligosaccharide concentration of 4%, 8%, 11%, 15%, 20%, respectively; FIG. 5 shows the inhibition rate of oligosaccharides from Dendrobium officinale on collagenase activity (oligosaccharide concentrations of 0.4%, 0.8%, 1%, 2%, 4%); where the ordinate is the collagenase inhibition rate %, and the abscissa (from 1 to 5) corresponds to the oligosaccharide concentration of 0.4%, 0.8%, 1%, 2%, 4%, respectively; FIG. 6 shows the activity of oligosaccharides from Dendrobium officinale on promoting the secretion of hyaluronic acid (oligosaccharide concentrations of 4%, 8%, 11%, 15%, 20%); where the ordinate is the secretion of hyaluronic acid (ng / mL), and the abscissa (from 1 to 5) corresponds to the oligosaccharide concentration of 4%, 8%, 11%, 15%, 20%, respectively; FIG. 7 shows the effect of oligosaccharides from Dendrobium officinale on the immunomodulatory activity of RAW264.7 cells (oligosaccharide concentrations of 3.125 µg / mL, 6.25 µg / mL, 12.5 µg / mL, 50 µg / mL, 100 µg / mL, 200 µg / mL); where A in FIG. 7 shows the effect of oligosaccharides on TNF-α secretion, the ordinate is TNF-α secretion pg / mL and the abscissa (from 1 to 5) corresponds to blank, lipopolysaccharide LPS and oligosaccharide with a final reaction concentration of 12.5 µg / mL, 6.25 µg / mL, 3.125 µg / mL, respectively; B in FIG. 7 shows the effect of oligosaccharides on IL-6 secretion, the ordinate is IL-6 secretion pg / mL, and the abscissa (from 1 to 5) corresponds to blank, lipopolysaccharide LPS and oligosaccharide with a final reaction concentration of 200 µg / mL, 100 µg / mL, 50 µg / mL, respectively. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0026] In order to further explain the technical means and effects of the present disclosure to achieve the intended purpose of the present disclosure, the specific implementations, structures, features and effects of the disclosure according to the present disclosure will be described in detail below with reference to the accompanying drawings and preferred embodiments. In the following description, different "one embodiment" or "an embodiment" do not necessarily refer to the same embodiment. In addition, specific features, structures or characteristics in one or more embodiments may be combined in any suitable form.

[0027] The term "skin care product" in the present disclosure may refer to a skin care toner, a skin care lotion, a skin care mask, a skin cream, etc.; the skin care products in the embodiments of the present disclosure can also refer to cosmetics.

[0028] The present disclosure provides an oligosaccharide from Dendrobium officinale, having a structure represented by formula (I):

[0029] The present disclosure provides a method for extracting the oligosaccharide from Dendrobium officinale described in above technical solution, comprising: 1) preparation of Dendrobium officinale powder: drying a stem of Dendrobium officinale, then smashing and sieving to obtain the Dendrobium officinale powder; In some embodiments, a temperature for the drying is 50-60°C; a moisture mass content of the stem of Dendrobium officinale after the drying is less than 6%; and a mesh number of the sieving is 20-40 meshes; 2) preparation of a water extract of Dendrobium officinale: mixing the Dendrobium officinale powder with water, then subjecting to a boiling water bath for one or more times, and merging the water extracts obtained after the boiling water bath to obtain a water extract of Dendrobium officinale; in some embodiments, the Dendrobium officinale powder and water are mixed according to a mass-volume ratio of 1: (50-80) g / mL; the boiling water bath is performed for 2-3 times; and a time for each boiling water bath is 1-2 h; 3) preparation of a concentrated water extract of Dendrobium officinale: centrifuging the water extract of Dendrobium officinale, and then concentrating under reduced pressure to obtain a concentrated water extract of Dendrobium officinale; in some embodiments, the centrifugation is conducted under 3000-4000 rpm / min, 8-12 min, and room temperature; the concentration under reduced pressure is conducted by using a rotary evaporation method, an ethanol recycling temperature of less than 50°C and a pressure of 0.7 Pa-0.9 Pa; a material-liquid mass-volume ratio of the concentrated water extract of Dendrobium officinale is 1 : (5-10) g / mL; 4) preparation of a redissolving liquid: adding ethanol to the concentrated water extract of Dendrobium officinale, precipitating, centrifuging and then discarding a supernatant to obtain a precipitate; and then adding water to the obtained precipitate for redissolving to obtain a redissolving liquid; in some embodiments, ethanol with a volume concentration of 95% is added to the concentrated water extract of Dendrobium officinale until an alcohol concentration of the mixture is 70%-80% by volume percentage, and then the overnight precipitation is carried out; the conditions of centrifugation are 3500-4500 rpm, 18-22 min, and room temperature; and water is added according to a weight-volume ratio of the precipitate and water of 1: (4-6) g / mL for redissolving; 5) preparation of an aqueous solution of the oligosaccharide from Dendrobium officinale: deproteinizing the redissolving liquid, and freeze-drying to obtain a crude polysaccharide of Dendrobium officinale; and adding water to dissolve the crude polysaccharide of Dendrobium officinale, then adding cellulase for a reaction, followed by inactivating and centrifuging, and discarding a precipitate to obtain an aqueous solution of the oligosaccharide from Dendrobium officinale; in some embodiments, the redissolving liquid is deproteinized by a Sevage method; the freeze-drying temperature is 45°C; adding water to dissolve the crude polysaccharide of Dendrobium officinale to 4.5-5.5 mg / mL; an added amount of the cellulase is 2%-4% of the weight of the crude polysaccharide of Dendrobium officinale; after the cellulase is added, the reaction is conducted under a reaction temperature of 55-65°C, and a reaction time of 2 h-4 h; inactivating by boiling for 25-35 min; the centrifugation are conducted under 3500-4500 rpm, 15-25 min, and room temperature; the oligosaccharide content in the aqueous solution of the oligosaccharide from Dendrobium officinale is greater than 80%; 6) preparation of the oligosaccharide from Dendrobium officinale: purifying the aqueous solution of the oligosaccharide from Dendrobium officinale to obtain the oligosaccharide from Dendrobium officinale; in some embodiments, the purification is conducted by using a G75 gel column; the purity of the oligosaccharide from Dendrobium officinale is 99.86%.

[0030] The present disclosure provides the oligosaccharide from Dendrobium officinale described in above technical solutions for use in the preparation of medicines and formulations for anti-aging or immunomodulatory.

[0031] The present disclosure provides the oligosaccharide from Dendrobium officinale described in above technical solutions for use in the preparation of foods and cosmetics for anti-aging or immunomodulatory. In some embodiments, the cosmetics include a face cream, an emulsion, a toning lotion or a repair cream.

[0032] The present disclosure provides a derivative of an oligosaccharide from Dendrobium officinale, where the derivative is generated by reacting the glucose residue of the oligosaccharide from Dendrobium officinale according to the above technical solutions with a hydrophobic residue, and the hydrophobic residue includes any one of linear or branched C 1 -C 18 alkyl, C 1 -C 18 alkyl carboxylic acid, C 1 -C 18 alkyl amine, C 1 -C 18 aryl alkyl and C 1 -C 18 alkyl amide, or the hydrophobic residue includes any one of benzoyl, phenylacetyl, and phenylpropionyl (Replacing hydroxy and methoxy with phenyl).

[0033] The present disclosure provides a method for preparing the oligosaccharide derivative described in the above technical solutions, comprising: 1) reducing a fatty acid into a fatty alcohol; in some embodiments, a reaction process of reducing the fatty acid into the fatty alcohol is as follows: mixing 10 mmol of fatty acid, 20 mmol of NaBH 4 , 40 mmol of AlCl 3 , 27 ml of THF, 3 ml of MeOH for a reaction, the reaction temperature is 100°C, the reaction time is 8 h, and the yield is 90%; 2) glycosylating the fatty alcohol with the oligosaccharide from Dendrobium officinale according to the above technical solutions; in some embodiments, performing the glycosylation reaction of fatty alcohols obtained by reduction and the oligosaccharides from Dendrobium officinale, the glycosylation reaction is conducted as follows: a molar ratio of alcohol to saccharide is 1: (8-10), the catalyst is cetylammonium bromide, the reaction temperature is 110-115°C, the reaction time is 3 h, and the yield is 80%.

[0034] The present disclosure provides the derivative described in above technical solutions for use in the preparation of anti-aging or immunomodulatory products.

[0035] In some embodiments, the products include medicines, foods, formulations and cosmetics; in some embodiments, the cosmetics include a face cream, an emulsion, a toning lotion or a repair cream.

[0036] The present disclosure provides a method for separating and purifying oligosaccharides from Dendrobium officinale. The polysaccharide of Dendrobium officinale has a large molecular weight (more than or equal to 100,000) and is difficult to be absorbed through skin cells, and thus mainly acts on the outside of the skin and has a moisturizing effect, while the oligosaccharide formed by enzymatic hydrolysis of polysaccharides has a small molecular weight and is easily absorbed through skin cells, which plays an anti-aging effect. According to the different molecular weights of the two, they can be used differently.

[0037] On the other hand, it is also found that the oligosaccharides from Dendrobium officinale can increase the survival rate and survival time of Hacat and RAW264.7 cells. Considering that the oligosaccharides from Dendrobium officinale may have good anti-aging activities and immunomodulatory activities, the activities on several main target proteins (elastase, collagenase, hyaluronic acid) contributing to aging have been verified. The results show that the oligosaccharides from Dendrobium officinale have a strong inhibitory effect on collagenase and can effectively alleviate the degradation and metabolism of collagen, thus maintaining the youth state of skin. They also have a certain inhibitory effect on elastase, and meanwhile a certain activity to promote the secretion of hyaluronic acid. The oligosaccharides from Dendrobium officinale also have a certain activity on TNF-α and IL-6, and have a good immunomodulatory effect.

[0038] Hereinafter, the present disclosure is further described in detail with reference to the specific examples as follows.EXAMPLE 1

[0039] In this example, substantially, Dendrobium officinale oligosaccharides were separated and purified from a stem of Dendrobium officinale, and further subjected to structural identification:

[0040] The fresh stem of Dendrobium officinale was cut and placed in a blast drying box to dry at 55°C, and the moisture content was measured. When the moisture content reached 6% or less, the stem was smashed and passed through a 20-40 mesh sieve; 50 g of a Dendrobium officinale powder was taken, 2500 mL of ultrapure water was added thereto, then the mixture was subjected to boiling water bath for 2 times with 2 h each time, and the water extracts were merged. Then a centrifugation was conducted at 3000-4000 rpm for 10 min at room temperature of 25°C to obtain a water extract of Dendrobium officinale; the water extract of Dendrobium officinale was concentrated under reduced pressure to 500 mL to obtain a concentrated water extract of Dendrobium officinale. An appropriate amount of 95% ethanol was added to the concentrated water extract of Dendrobium officinale until an alcohol concentration of the mixture was 70-80%, then the mixture was precipitated overnight, centrifuged at 4000 rpm for 20 min at room temperature of 25°C, and the supernatant was discarded to obtain a precipitate; water was added according to the ratio of the weight of the precipitate to the volume of water of 1 : 5 (m : v) (g / mL) for redissolving to obtain a redissolving liquid. The redissolving liquid was deproteinized by the Sevage method, and then freeze-dried to obtain a crude polysaccharide of Dendrobium officinale. 200 mg of the crude polysaccharide of Dendrobium officinale was taken and dissolved with water to 5 mg / mL, cellulase was added thereto in 2%-4% of the weight of the polysaccharide of Dendrobium officinale and mixed evenly, then the mixture was reacted at 60°C for 2 h-4 h, boiled for 30 min for inactivation, centrifuged at 4000 rpm for 20 min at room temperature of 25°C, and a precipitate was discarded to obtain an aqueous solution of Dendrobium officinale oligosaccharide (the content of oligosaccharides in the aqueous solution of Dendrobium officinale oligosaccharide is greater than 80%). An appropriate amount of the aqueous solution of Dendrobium officinale oligosaccharide was taken and passed through a G75 gel column to obtain 150 mg of purified Dendrobium officinale oligosaccharide (with a purity of 99.86% and a molecular weight of about 1,000).

[0041] The purity of the Dendrobium officinale oligosaccharides was identified by high performance liquid chromatography, and the results are shown in FIG. 1. The structure of Dendrobium officinale oligosaccharides was identified by hydrogen nuclear magnetic resonance analysis and C13 nuclear magnetic resonance analysis, and the results are shown in FIG. 2.EXAMPLE 2 Experiment on the DPPH free radical scavenging1. Reagents:

[0042] DPPH, water-soluble vitamin E (Trolox) (Trolox dissolved in absolute ethanol, with a concentration of 10 mM, stored at -20°C), pure water.2. Experimental method:

[0043] The drug to be tested was mixed with DPPH (a final concentration of 100 M) and reacted at 30°C for 1 h. 3 repeating wells were set, and a blank control well without drug and a Trolox positive control well were set at the same time. The OD value was measured by a microplate reader with a detection wavelength of 515 nm, and the anti-oxidation rate was calculated. 3. Experimental results:

[0044] As shown in Table 1. Table 1SampleFinal reaction concentrationAnti-oxidation rate (%)Trolox2595.668%Dendrobium officinale oligosaccharide4%11.335%Dendrobium officinale oligosaccharide8%14.346%Dendrobium officinale oligosaccharide11%16.213%Dendrobium officinale oligosaccharide15%21.155%Dendrobium officinale oligosaccharide20%25.217% 4. Conclusion:

[0045] According to the data in Table 1, FIG. 3 is made for a clearer comparison. As can be seen from FIG. 3 and Table 1, the scavenging ability of DPPH free radicals of Dendrobium officinale oligosaccharides is evaluated in the experiment. The results show that the Dendrobium officinale oligosaccharides have a certain anti-oxidation rate (> 20%) at a concentration of 15%-20%.EXAMPLE 3 Experiment on the elastase inhibition rate1. Reagents:

[0046] Sample, positive control of catechin, Tris, HCl, porcine pancreatic elastase, N-succinyl-alanine-alanine-alanine-p-nitroaniline, DMSO.2. Experimental method:

[0047] (1) Preparation of solution Preparation of a sample solution: 1 mg / mL sample solution was prepared (pure water or a buffer solution was used for preparation, and if insoluble in water or the buffer solution, the sample was firstly dissolved with DMSO, and then water or the buffer solution was added. In principle, the content of DMSO should not exceed 5%). Preparation of a positive control: 1 mg / mL catechin; Preparation of a buffer solution: 0.1 M of Tris-HCl buffer solution, PH = 8.0; Preparation of a reaction substrate solution: N-succinyl-alanine-alanine-alanine-p-nitroaniline (AAAPVN) solution (2 mM); Preparation: 28.5 µL of porcine pancreatic elastase solution (0.171 U / mL) was taken and dissolved in 10 mL of Tris-HCl buffer solution. (2) The solution of 50 µL of sample, 50 µL of porcine pancreatic elastase and 100 µL of reaction substrate was added to a 96-well plate. (3) After keeping at room temperature for 5 min, an absorbance at 420 nm was measured. 3. Experimental results:

[0048] As shown in Table 2. Table 2SampleFinal reaction concentrationElastase inhibition rate %epicatechin gallate100 µg / ml68.540%Dendrobium officinale oligosaccharide4%-2.135%Dendrobium officinale oligosaccharide8%3.886%Dendrobium officinale oligosaccharide11%8.417%Dendrobium officinale oligosaccharide15%9.723%Dendrobium officinale oligosaccharide20%15.827% 4. Conclusion:

[0049] According to the data in Table 2, FIG. 4 is made for a clearer comparison. As can be seen from FIG. 4 and Table 2, the elastase inhibition rate of Dendrobium officinale oligosaccharides is evaluated in the experiment. The results show that the Dendrobium officinale oligosaccharides have a certain elastase inhibition activity (> 10%) at a concentration of 20%.EXAMPLE 4 Experiment on collagenase inhibition rate1. Reagents:

[0050] Protease buffer solution, protease, protease substrate, inhibitor (1,10-Phenanthroline). All reagents were stored at -20°C.

[0051] 2. Experimental method: (1) Solution preparation: The collagenase reaction mixture is shown in Table 3. Table 3CompositionContent (%)Collagenase buffer solution60Collagenase substrate40 (2) The compositions were designed as follows: a. Background blank well: 100 µL / well of protease buffer solution. b. Sample well: 100 µL, prepared into different concentrations. c. Positive control well: 10 µL of protease + 90 µL of protease buffer solution. d. Inhibitor control well: 10 µL of protease + 2 µL of inhibitor + 88 µL of protease buffer solution as shown in Table 4. Table 4 GroupSample well (µl)Positive control well (µl)Inhibitor control well (µl)Background blank well (µl)Sample to be tested1-90---Protease101010-Inhibitor--2-Protease buffer solutionDiluted to 1009088100 (3) After adding reagents according to the above groups, 100 µL of collagenase reaction mixture was added to each well, incubated in the dark, and then the OD value at 345 nm was measured.

[0052] The incubation time depended on the activity of the sample to be tested, and the low activity sample could be incubated for 1-3 h. 3. Experimental results:

[0053] As shown in Table 5. Table 5SampleFinal reaction concentrationCollagenase inhibition rate (%)Positive inhibitor1%102.23%Dendrobium officinale oligosaccharide0.4%50.32%Dendrobium officinale oligosaccharide0.8%63.63%Dendrobium officinale oligosaccharide1%72.20%Dendrobium officinale oligosaccharide2%84.80%Dendrobium officinale oligosaccharide4%110.54% 4. Conclusion:

[0054] According to the data in Table 5, FIG. 5 is made for a clearer comparison. As can be seen from FIG. 5 and Table 5, the collagenase inhibition rate of Dendrobium officinale oligosaccharides is evaluated in the experiment. The results show that the Dendrobium officinale oligosaccharides have a strong collagenase inhibition activity (> 50%) at a concentration of 0.4%-4%.EXAMPLE 5 Experiment on promoting secretion of hyaluronic acid1. Reagents and cells:

[0055] Human immortalized cortical cells (Hacat cells), human hyaluronic acid Elisa kit, distilled water.2. Experimental method:

[0056] A 96-well plate was inoculated with 100 mL of cell suspension per well, and a concentration of the suspension was 4×10 5< cells / mL. After 24 h of incubation, the original culture medium was discarded, then the cells were washed with PBS for 1-2 times and the serum-free medium containing different concentrations of the drug to be tested was added thereto, and continued to incubate for 24 h. A blank control without drugs and a TGF-β positive control were set. Then the cells were lysed with ripa lysis buffer solution containing 1% PMSF, the supernatant after centrifugation was taken, and the secretion of hyaluronic acid was detected according to the method provided in the collagen ELISA kit. The OD value was measured by a microplate reader with a detection wavelength of 450 nm. The OD value was substituted into the regression equation of the standard curve to calculate the increase rate of the secretion of hyaluronic acid.3. Experimental results:

[0057] As shown in Table 6. Table 6SampleFinal reaction concentrationSecretion of hyaluronic acid (ng / mL)Blank control0.2508Dendrobium officinale oligosaccharide4%0.2294Dendrobium officinale oligosaccharide8%0.5530Dendrobium officinale oligosaccharide11%0.3691Dendrobium officinale oligosaccharide15%0.4930Dendrobium officinale oligosaccharide20%0.5392 4. Conclusion:

[0058] According to the data in Table 6, FIG. 6 is made for a clearer comparison. As can be seen from FIG. 6 and Table 6, the secretion-promoting ability of hyaluronic acid of Dendrobium officinale oligosaccharides is evaluated. The results show that the Dendrobium officinale oligosaccharides have a certain activity to promote the secretion of hyaluronic acid (> 0.3 ng / mL) at a concentration of 8%-20%.EXAMPLE 6

[0059] Evaluation of the immunomodulatory activity of Dendrobium officinale oligosaccharides1. Experimental cells and reagents:

[0060] RAW264.7 cells, TNF-α kit, IL-6 kit, distilled water.2. Experimental methods:

[0061] The logarithmic-growth RAW264.7 cells were taken and inoculated into 12-well plates after cell counting. The plate was placed in an incubator overnight, and then the cell supernatant was discarded. Different concentrations of Dendrobium officinale oligosaccharides, lipopolysaccharides and basal medium 1640 were added under aseptic operation, and the supernatant after 24 h of intervention was collected and stored at -20°C for later use. The changes in TNF-α and IL-6 secretion were detected according to the instructions of the ELISA kit. The OD value was measured by a microplate reader with a detection wavelength of 450 nm. The OD value was substituted into the regression equation of the standard curve to calculate the increase rate of TNF-α and IL-6 secretion.3. Experimental results:

[0062] As shown in Table 7. Table 7SampleFinal reaction concentration (µg / mL)TNF-α pg / mLFinal reaction concentration (µg / mL)IL-6 pg / mLBlank control64.711013.72LPS1085.131406.19Dendrobium officinale oligosaccharide12.553.862001117.33Dendrobium officinale oligosaccharide6.2535.661001376.42Dendrobium officinale oligosaccharide3.1251062.29501593.01 4. Conclusion:

[0063] According to the data in Table 7, FIG. 7 is made for a clearer comparison. As can be seen from FIG. 7 and Table 7, the effects of Dendrobium officinale oligosaccharide on the immunomodulatory activity of RAW264.7 cells is evaluated. The results show that the secretion of TNF-α by RAW264.7 macrophages is significantly increased at a concentration of 3.125 µg / mL, which is equivalent to the positive control LPS. The secretion of IL-6 by RAW264.7 macrophages can be significantly increased at a concentration of 50 µg / mL, which is equivalent to the positive control LPS.EXAMPLE 7: glycosylation of fatty acids 1. Experimental reagents:

[0064] Dendrobium officinale oligosaccharides, fatty acids, NaBH 4 , MeOH, metal salts, AlCl 3 , THF, catalysts, etc.

[0065] 2. Experimental methods and results: 1) Reducing a C 6 fatty acid into a fatty alcohol: 10 mmol of fatty acid, 20 mmol of NaBH 4 , 40 mmol of AlCl 3 , 27 mL of THF and 3 mL of MeOH were reacted at a temperature of 100°C for a time of 8 h with a yield of 90%; 2) Glycosylation: the reduced fatty alcohol and the Dendrobium officinale oligosaccharide were subjected to a glycosylation reaction with a molar ratio of the alcohol and the saccharide of 1 : (8-10); the catalyst was cetyl ammonium bromide; the reaction was conducted under a temperature of 110-115°C for a time of 3 h with a yield of 80%. 3. Results discussion:

[0066] The reaction results show that Dendrobium officinale oligosaccharides with terminal glycosides can also undergo the glycosylation reaction with fatty acids.EXAMPLE 8

[0067] This example provides an anti-aging or immunomodulatory face cream, comprising components in mass percentage: 0.4-0.6% of Dendrobium officinale stock solution rich in oligosaccharides, 7.5-8.5% of stearic acid, 1.5-2.5% of C 16 alcohol, 1.5-2.5% of self-emulsifying monoglyceride, 1.5-2.5% of hydrogenated lanolin, 11-13% of liquid paraffin, 6-8% of glycerin, 1-2% of an emulsifier, 0.1-0.3% of a preservative, 0.1-0.3% of an essence, and the balance of water. The face cream of the above formula was prepared according to a conventional method for preparing cosmetics.EXAMPLE 9

[0068] This example provides an anti-aging or immunomodulatory emulsion, comprising components in mass percentage: 0.4-0.6% of Dendrobium officinale stock solution rich in oligosaccharides, 1.3-1.5% of stearic acid, 0.08-0.12% of cetyl alcohol, 1.7-1.9% of 2-ethyl alcohol cetyl stearate, 0.1-0.3% of isopropyl myristate, 0.9-1.1% of 2-hexyl-1-decanol, 7-8% of liquid paraffin, 2.5-3.5% of glycerol, 7-9% of propylene glycol, 0.08-0.12% of triethanolamine, 0.3-0.4% of a carboxyvinyl polymer, 1.5-2.5% of Arlacel 165, 0.1-0.3% of a preservative, 0.1-0.3% of an essence, and the balance of water; The emulsion of the above formula was prepared according to a conventional method for preparing cosmetics.EXAMPLE 10

[0069] This example provides an anti-aging or immunomodulatory toner, comprising components in mass percentage: 1-10% of Dendrobium officinale stock solution rich in oligosaccharides, 0.3-0.4% of sodium polyacrylate, 3.5-4.5% of glycerol, 2-3% of 1,3-butanediol, 0.4-0.6% of vitamin B 5 , 0.4-0.6% of arbutin, 0.04-0.06% of EDTA-Na 2 , and the balance of deionized water; the method for preparing the toning lotion is as follows: the sodium polyacrylate was dissolved in water, stirred to fully swelling, then the remaining ingredients were added, and continued to stirring; then the Dendrobium officinale stock solution rich in oligosaccharides was added thereto, the mixture was stirred, and then deionized water was added to a constant volume to obtain the toner.

[0070] A preferred solution is as follows: 0.35 g of sodium polyacrylate was dissolved in 70 mL of water, stirred to fully swelling, then the remaining ingredients were added, and continued to stirring; then 10% of the Dendrobium officinale stock solution rich in oligosaccharides obtained in Example 1 was added thereto, the mixture was stirred uniformly, and then deionized water was added to 100 mL and packed.EXAMPLE 11

[0071] This example provides an anti-aging or immunomodulatory repair cream, comprising components in mass percentage: 5-7% of glycerin, 1.4-1.6% of carbomer, 1.4-1.6% of triethanolamine, 5.5-6.5% of propylene glycol, 0.15-0.25% of ethyl p-hydrobenzoate, 9-11% of Dendrobium officinale stock solution rich in oligosaccharides, 0.4-0.6% of an essential oil, and the balance of deionized water.

[0072] The method for preparing the repair cream is as follows: the carbomer was taken in an appropriate amount of deionized water, the mixture was stirred uniformly, let stand overnight to fully swelling, then glycerin was added, and the pH value was adjusted with triethanolamine to increase the viscosity of the gel matrix to obtain a carbomer gel; the Dendrobium officinale stock solution rich in oligosaccharides, propylene glycol and deionized water were mixed evenly, then the carbomer gel, essential oil and ethyl p-hydrobenzoate were added thereto, the resulting mixture was stirred until uniform and delicate, distilled water was added to a sufficient amount, and the mixture was grinded evenly to obtain a repair cream.

[0073] A preferred solution is as follows: 6 g of glycerin, 1.5 g of carbomer, 1.5 g of triethanolamine, 6 g of propylene glycol, 0.2 g of ethyl p-hydrobenzoate, 10% of Dendrobium officinale stock solution rich in oligosaccharide obtained in Example 1, and an appropriate amount of deionized water were added to a total weight of 100 g. The preparation method is as follows: the carbomer was taken in an appropriate amount of deionized water, the mixture was stirred uniformly, let stand overnight to fully swelling, then glycerin was added, and the pH value was adjusted with triethanolamine to increase the viscosity of the gel matrix to obtain a carbomer gel; a prescription amount of tanshinone extract, propylene glycol and deionized water were mixed evenly, then the carbomer gel, 0.5% of an essential oil and ethyl p-hydrobenzoate were added thereto, the resulting mixture was stirred until uniform and delicate, distilled water was added to a sufficient amount, and the mixture was grinded evenly to obtain a repair cream.

[0074] The above are only preferred embodiments of the present disclosure, and are not intended to be used to limit the present disclosure in any form.

Claims

1. An oligosaccharide from Dendrobium officinale, having a structure represented by formula (I):

2. A derivative of an oligosaccharide from Dendrobium officinale, wherein the derivative is generated by reacting the glucuronic acid residue of the oligosaccharide from Dendrobium officinale according to claim 1 with a hydrophobic residue, and the hydrophobic residue comprises any one of linear or branched C1-C18 alkyl, C1-C18 alkyl carboxylic acid, C1-C18 alkyl amine, C1-C18 aryl alkyl and Ci-Cis alkyl amide, or the hydrophobic residue comprises any one of benzoyl, phenylacetyl and phenylpropionyl.

3. The oligosaccharide from Dendrobium officinale according to claim 1 or the derivative according to claim 2 for use in preparation of anti-aging or immunomodulatory products.

4. The oligosaccharide from Dendrobium officinale or the derivative according to claim 3, wherein the products comprise medicines, foods, formulations and cosmetics.

5. The oligosaccharide from Dendrobium officinale or the derivative according to claim 4, wherein a type of the cosmetics comprises a face cream, an emulsion, a toning lotion or a repair cream.

6. A method for extracting the oligosaccharide from Dendrobium officinale according to claim 1, comprising: 1) preparation of a Dendrobium officinale powder: drying a stem of Dendrobium officinale, then smashing and sieving to obtain a Dendrobium officinale powder; 2) preparation of a water extract of Dendrobium officinale: mixing the Dendrobium officinale powder with water, subjecting to a boiling water bath for one or more times, and then merging water extracts to obtain a water extract of Dendrobium officinale; 3) preparation of a concentrated water extract of Dendrobium officinale: centrifuging the water extract of Dendrobium officinale, and then concentrating under reduced pressure to obtain a concentrated water extract of Dendrobium officinale; 4) preparation of a redissolving liquid: adding ethanol to the concentrated water extract of Dendrobium officinale, precipitating, centrifuging and then discarding a supernatant to obtain a precipitate; adding water to the obtained precipitate for redissolving to obtain a redissolving liquid; 5) preparation of an aqueous solution of the oligosaccharide from Dendrobium officinale: deproteinizing the redissolving liquid, and freeze-drying to obtain a crude polysaccharide of Dendrobium officinale; adding water to dissolve the crude polysaccharide of Dendrobium officinale, then adding cellulase for a reaction, followed by inactivating and centrifuging, and discarding a precipitate to obtain an aqueous solution of the oligosaccharide from Dendrobium officinale; 6) preparation of the oligosaccharide from Dendrobium officinale: purifying the aqueous solution of the oligosaccharide from Dendrobium officinale to obtain the oligosaccharide from Dendrobium officinale.

7. The method according to claim 6, wherein, in step 1), a temperature for the drying is 50-60°C; a moisture mass content of the stem of Dendrobium officinale after the drying is less than 6%; a mesh number of the sieving is 20-40 meshes; in step 2), the Dendrobium officinale powder and water are mixed in a mass-volume ratio of 1 : (50-80) g / mL; the boiling water bath is performed for 2-3 times; a time for each boiling water bath is 1-2 h; in step 3), the centrifugation is conducted under conditions of 3000-4000 rpm / min, 8-12 min and room temperature; the concentration under reduced pressure is conducted under conditions of a rotary evaporation method, an ethanol recycling temperature of less than 50°C and a pressure of 0.7 Pa-0.9 Pa; a material-liquid mass-volume ratio of the concentrated water extract of Dendrobium officinale is 1 : (5-10) g / mL; in step 4), the addition of ethanol to the concentrated water extract of Dendrobium officinale and the precipitation are specifically conducted as follows: adding ethanol with a volume concentration of 95% to the concentrated water extract of Dendrobium officinale until an alcohol concentration of a mixture is 70%-80% in volume percentage, and then precipitating overnight; the centrifugation is conducted under conditions of 3500-4500 rpm, 18-22 min and room temperature; a weight-volume ratio of the precipitate and water for the redissolving is 1 : (4-6) g / mL; in step 5), the deproteinization of the redissolving liquid is conducted by a Sevage method; a temperature for the freeze-drying is 45°C; the addition of water to dissolve the crude polysaccharide of Dendrobium officinale is conducted as follows: adding water to the crude polysaccharide of Dendrobium officinale for dissolving until 4.5-5.5 mg / mL; an added amount of the cellulase is 2%-4% of a weight of the crude polysaccharide of Dendrobium officinale; after adding the cellulase, the reaction is conducted under a reaction temperature of 55-65°C for a reaction time of 2 h-4 h; the inactivation is conducted by boiling for 25-35 min; the centrifugation is conducted under conditions of 3500-4500 rpm, 15-25 min and room temperature; an oligosaccharide content in the aqueous solution of the oligosaccharide from Dendrobium officinale is greater than 80%; in step 6), the purification is conducted by passing through a G75 gel column; a purity of the oligosaccharide from Dendrobium officinale is 99.86%.

8. A method for preparing the derivative according to claim 2, comprising: 1) reducing a fatty acid into a fatty alcohol; 2) glycosylating the fatty alcohol with the oligosaccharide from Dendrobium officinale according to claim 1 to obtain the derivative.

9. The method according to claim 8, wherein in step 1), a reaction process of the reduction of a fatty acid into a fatty alcohol is as follows: mixing 9-11 mmol of the fatty acid, 18-22 mmol of NaBH4, 38-42 mmol of AlCl3, 25-29 ml of THF and 2-4 ml of MeOH for a reaction, and the reaction is conducted under a temperature of 95-105°C for a time of 7.5-8.5 h with a yield of 88-92%; in step 2), a reaction process of the glycosylation is as follows: mixing the alcohol and the saccharide in a molar ratio of 1 : (8-10), and then using cetyl ammonium bromide as a catalyst for catalyzation, and the reaction is conducted under a temperature of 110-115°C for a time of 2.5-3.5 h with a yield of 78-82%.