WO2015124113A1

WO2015124113A1 - Semi-synthesis method for luteolin, galuteolin and luteolin rutinoside

Info

Publication number: WO2015124113A1
Application number: PCT/CN2015/073238
Authority: WO
Inventors: 闻永举
Original assignee: 闻永举
Priority date: 2014-02-23
Filing date: 2015-02-21
Publication date: 2015-08-27
Also published as: CN103833714A; CN103833714B

Abstract

Provided is a semi-synthesis method for luteolin, galuteolin and luteolin rutinoside. In the method, two steps of dehydrogenation and demethylation are combined into one step, and reaction conditions of dehydrogenation and demethylation are mild and easy to control; few reagents are used so that the method is green and environmentally friendly; and the demethylation yield is high, and industrial production is easy to realize.

Description

Method for semi-synthesis of luteolin, luteolin and luteolin rutinoside

Technical field

The present invention relates to a method for semi-synthesizing luteolin, luteolin, luteolin rutin, hesperetin-7-O-glucoside semi-synthetic luteolin, and hesperetin semi-synthetic luteolin by hesperidin Its field belongs to chemistry and medicine.

Background technique

Hesperidin and luteolin resources comparison: Hesperidin is widely found in Rutaceae plants, such as orange peel, citrus is also the world's largest fruit, China has abundant citrus resources, and the use of citrus in China is limited. In the use of citrus fruit, the peel has not received enough attention, and citrus peel as a by-product of processing accounts for 30% to 50%. Only a small portion was recovered for use as tangerine peel and extracted hesperidin, but most of it was discarded, causing great waste and environmental pollution. The 2010 edition of the Chinese Pharmacopoeia stipulates that the pericarpin should not be less than 5.0%, the hesperidin content of the tangerine peel should not be less than 3.5%, the hesperidin is rich in source, the extraction process is simple, and the price is low. 90% hesperidin per kilogram is about 180 yuan. Hesperidin can be used as a raw material to semi-synthesize dioxin, geranin, luteolin, luteolin-7-O-glucoside, and methyl hesperidin. Among them, luteolin and its glycosides are 3,4,5,7-tetrahydroxyflavonoids and their glycoside derivatives, which are mainly found in plants such as chrysanthemum, honeysuckle, travertine, goat's milk and sclerotium. High anti-inflammatory, anti-oxidant, anti-tumor and other biological activities. However, luteolin is low in the natural world and expensive, and 98% luteolin is 2,000 yuan per kilogram.

Hesperidin semi-synthetic luteolin existing method: luteolin can be prepared by hesperidin semi-synthesis, there are usually three methods reported in the literature or patents: (1) hesperidin hydrolyzed hesperetin, hesperetin Dehydrogenation produces geranyl lignin and geranyl demethylation to prepare luteolin. The reaction formula is shown in (I) ([1] Li Yushan. A new technology for the preparation of luteolin from hesperidin [P].2013, CN, 201310004892.8); (2) dehydrogenation of hesperidin to produce odisamine, diosmin hydrolyzed to produce geranyl lignin, and geranyl lignin demethylated to prepare luteolin, the total yield of 3 steps is 35%, See (II) ([2] Criminal Power, Sun Zhizhong, Han Wenhui, Han Xiaoling. Semi-synthesis of luteolin and glucoside[J]. Chinese Journal of Pharmaceutical Industry, 1994, 25(11): 484-487); (3) Hesperidin hydrolyzed hesperetin, demethylation of hesperetin to eriophthol, dehydrogenation of ergophenol to luteolin, the total yield of three steps is 45.9%, and the reaction is seen in (III) ([3 Sun Zhizhong, Hao Wenhui, Duan Shuhong, et al. Semi-synthesis. Chinese Journal of Modern Applied Pharmacy, 1999, 16(1): 30-31).

Hesperidin semi-synthetic luteolin deficiency: The above three methods of hesperidin semi-synthetic luteolin are three steps, each step uses more organic reagents, some reagents are not recycled, on the one hand, the cost is increased. On the other hand, it pollutes the environment; each step requires purification, drying, etc., on the one hand, the production cycle is long, on the other hand, the sample has a certain loss during the treatment process, which reduces the yield of the product, and the separation is troublesome. For example, the demethylation of geranyl lignin is described as follows: it is slightly soluble in glacial acetic acid, demethylated with glacial acetic acid-HBr, and glacial acetic acid-HBr is also difficult to recover. Therefore, the demethylation method is usually pyridine-aluminum trichloride, but since pyridine and aluminum trichloride form a salt, it is difficult to form a homogeneous solvent, so the amount thereof is large, since pyridine can form a salt with aluminum trichloride and phenol, and is recovered. The rate is limited and the environment is polluted.

Semi-synthesis method and defect of luteolin: luteolin is luteolin-7-O-glucoside, which is less in plants, such as the 2010 edition of the Chinese Pharmacopoeia honeysuckle regulation containing luteolin (C ₂₁ H ₂₀ O ₁₁ ) Not less than 0.05%, the chrysanthemum contains no less than 0.080%, and the content of chlorogenic acid in the honeysuckle is higher than that of honeysuckle, but does not contain luteolin. Because of the trace amount of luteolin, the price is 1/ of honeysuckle. 2-1/3, but its application range is far less than honeysuckle, the reason is that the mountain silver flower does not contain luteolin, which makes people more respect for honeysuckle. Luteolin is produced by the reaction of luteolin with 2,3,4,6-tetra-O-acetyl-aD-bromopyranose. The luteolin was synthesized from hesperidin, and the luteolin was prepared from hesperidin by 35%. The luteolin was prepared by luteolin. The total yield of the two steps was 16.8%. Based on hesperidin, the total yield of the five steps was 5.88% ([2] Criminal Power, Sun Zhizhong, Han Wenhui, Han Xiaoling. Semi-synthesis of luteolin and glucoside [J]. Chinese Journal of Pharmaceutical Industry, 1994, 25(11): 484-487. It is obvious that the existing hesperidin semi-synthetic luteolin has many steps, the yield is too low, and 2,3,4,6-tetra-O-acetyl-aD-bromopyrrol The price of glucone is relatively expensive, resulting in higher industrialization costs.

Demethylation and Defects of Glycoside Compounds: In the total synthesis of puerarin, demethylation uses expensive trimethyliodosilane and the more expensive reagent acetonitrile as solvent ([4] Zhang Peicheng. Flavonoid Chemistry [M]. Chemical Industry Press, 2009, 357), demethylation, resulting in high product costs, extracted from the natural plant Pueraria, does not have a synthetic advantage. Puerarin

As a carbon glycoside, hydrolysis does not occur during the demethylation process, and its stability is much higher than that of oxyglucoside. However, oxyglucosides are easily hydrolyzed by acid, easily dehydrated by heat, and their stability is far worse than that of phenol methyl ether. Demethylation of oxomethyl ether is carried out by the existing demethylation method, and the obtained product is first de-sugarized and then desorbed. Methyl group, the last de-saccharification and demethylation to obtain aglycon, because the sugar ring may even dehydrate at high temperature, and react with the flavonoid nucleus, the product is more complicated, and it is difficult to obtain the glycoside which is demethylated first and retains the sugar. Therefore, it has not been reported that oxoside preferentially dephenolate methyl and retains sugar ring. Therefore, hesperidin, dipyridin, and hesperetin-7-O-glucoside selectively remove methyl groups from the molecule, which is entirely a new challenge. If this method can be achieved, hesperidin, The steps of dimethoate, hesperetin-7-O-glucoside, hesperetin semi-synthetic luteolin, luteolin, luteolin rutin are greatly reduced, the cost is greatly reduced, and industrial application value is obtained.

Oxygen free radical damage and flavonoid protection. The cell membrane has its loose chemical structure, is extremely elastic and flexible, and its electrons are easily lost. Therefore, once the cell membrane is attacked by free radicals, its electrons are taken away, its function is lost, various inflammations are caused, and it is further deteriorated, resulting in various diseases such as cardiovascular diseases and tumors. Some endogenous enzymes, such as superoxide dismutase (SOD), catalase, glutathione peroxidase, and vitamin C, vitamin E, carotene and other compounds, have the ability to scavenge free radicals . In addition to the above substances, in order to reduce the risk of free radicals, it is also necessary to find and discover exogenous free radical scavengers. These substances are used in preference to free radical binding to block the attack of free radicals on the human body and protect the human body from harm. . A large number of studies have proved that flavonoids have many physiological activities and pharmacological effects such as anti-oxidation, anti-cancer, anti-AIDS, anti-bacterial, anti-allergic, anti-inflammatory, etc., and have no toxic and side effects, which are of great significance for the prevention and treatment of human tumors, aging and cardiovascular diseases. . The flavonoid molecule contains multiple phenolic hydroxyl groups, which are easily metabolized by the intestinal flora or the liver enzyme system, and the metabolites also have corresponding phenolic hydroxyl groups. The free radical scavenging effect of phenolic hydroxyl groups contained in flavonoids and metabolites may be the basis of many of their effects.

O-diphenol has significant antioxidant and pharmacological effects: A and B benzene rings are linked by three carbons to form flavonoids. When ring A has 5-OH and 7-OH dihydroxy groups, 7-OH often forms glycosidic bonds. -OH forms an intramolecular hydrogen bond with 4C=O, so the A ring has weak oxidation resistance. When the B ring contains ortho-diphenolic hydroxyl groups, it has strong antioxidant activity and relatively good pharmacological effects, such as rutin, luteolin and glycosides, and eriocitrin. For example, other compounds with ortho-diphenolic hydroxyl groups in traditional Chinese medicines also have good pharmacological activities, such as protocatechuic aldehyde, Danshensu, chlorogenic acid and the like. The above description of ortho-diphenol has an important effect on the action of flavonoids and other compounds, which may be due to its strong antioxidant activity. Antioxidant activity of flavonoids, H ₂ O ₂ clearance rate is commonly used for antioxidant activity, and erioconsin-7-rutinoside and luteolin-7-rutinoside have strong anti-H ₂ O ₂ activity. Hesperidin has weak anti-H ₂ O ₂ activity, and naringin, diosmin, and quercetin have almost no H ₂ O ₂ activity ([5]Zbigniew Sroka*, Izabela Fecka, and Wojciech Cisowski. Antiradical and Anti-H ₂ O ₂ Properties of Polyphenolic Compounds from an Aqueous Peppermint Extract. Z. Naturforsch, 2005, 60 (826): 832.).

Antibiotic abuse hazard, calling for Chinese medicine substitutes: Since Fleming discovered penicillin in 1928, antibiotics have saved countless lives. At present, antibiotics are increasingly used in medical and aquaculture, and are even in abuse in many areas. However, antibiotics can have a wide-ranging effect on the body's immune system, such as headache, nausea, vomiting, constipation, etc., while reducing inflammation and sterilization. Therefore, it is necessary to find safe, low-toxic side effects and effective alternative medicine. Many experts in the medical field are looking for Chinese medicine to replace Western medicine antibiotics. Honeysuckle is a valuable traditional Chinese medicine with the functions of clearing away heat and detoxifying and evacuating wind and heat. Its extracts are widely used in medicines such as Shuanghuanglian preparation, Yinqiao Jiedu Tablet and Honeysuckle Flower Dew. At the same time, the demand for beverages and daily chemical industry is also exploding. The products that have been developed include: honeysuckle drinks (Jiduobao, Wang Laoji), honeysuckle beer, honeysuckle toothpaste and so on. The mountain silver flower also has the effect of clearing away heat and detoxifying and evacuating wind and heat. In the 2000 edition and the previous pharmacopoeia, it was the same medicine as honeysuckle. In the 2005 edition of the pharmacopoeia, it was divided into two kinds of drugs because of the trace of luteolin. However, the content of chlorogenic acid is higher than that of honeysuckle, and the yield per mu is 2-3 times that of honeysuckle. The price is 1/2-1/3 of honeysuckle, but its application range is far less than that of honeysuckle. The reason is that the mountain silver flower does not contain limonium. Glycoside, which makes people more respect for honeysuckle. If the extract of arborvitae is added to luteolin, the chlorogenic acid and luteolin are the same as the active ingredients of honeysuckle, and the effect of antibacterial and other effects is not much different, then the silver stalk can be used instead of luteolin instead of honeysuckle. Since the yield of the mountain silver flower is much higher than that of the honeysuckle, it can save a lot of cultivated land.

Carbohydrates and polyphenolic hydroxyl groups have solubilization effect on flavonoids: flavonoids contain two benzene rings in the molecule. Due to the planarity of the benzene ring, the molecules and molecules are easily overlapped, making them insoluble in water, such as orange. Derinoside, naringin, rutin, baicalin, etc. When the A ring has a polyhydroxy compound and the B ring has no hydroxyl group, the molecules are more likely to undergo self-phase rearrangement to form crystals, and their solubility in water is small. For example, baicalin contains only o-diphenolic hydroxyl group in ring A, B ring is not, insoluble in water, and insoluble in solvents such as methanol, ethanol, acetone, etc., and breviscapine has one ortho-diphenolic hydroxyl group in addition to ring A, and one ring in B ring. Hydroxyl, soluble in methanol, ethanol, heat water. Wild laccase is insoluble in water, while luteolin-7-O-neo-hesperidin is soluble in water. The difference between the two is that the former has one less 3'-OH than the latter on the B ring, indicating an increase in phenolic hydroxyl groups. Increase polarity and help increase its water solubility. Luteolin is very soluble in water, luteolin-7-O-glucoside is slightly soluble in water, luteolin-7-O-rutinoside, soluble in water, soluble in hot water, indicating increased glycosylation It is beneficial to improve water solubility.

In summary, the use of abundant sources, simple preparation process, low price of hesperidin and its derivatives, such as hesperetin-7-O-glucoside, hesperetin semi-synthetic luteolin rutin, luteolin, Luteolin, the latter three have excellent antibacterial, anti-tumor, anti-oxidation effects, low toxic and side effects, used in the fields of medicine, food, cosmetics, for the benefit of mankind, has been urgently important.

Summary of the invention

The object of the present invention is to provide semi-synthetic luteolin, luteolin, luteolin rutinoside from hesperidin, hesperetin, hesperetin-7-O-glucoside, which has fewer steps, lower cost and high yield. , environmentally friendly industrial preparation methods.

Solution: Combine the two-step reaction of hesperetin, hesperidin, hesperetin-7-O-glucoside dehydrogenation and demethylation into a one-step reaction.

Technical features of the invention:

Pyridines, trivalent aluminum ion compounds, low-boiling alcohols, hesperetin, and iodine are mixed and dehydrogenated at room temperature to 110 ° C to form a homogenous liquid, and the dehydrogenation of the starting product is completely followed by PC/TLC/HPLC. Then, it is sealed at 80 °C-180 °C for 2-80 h, and the demethylation product is checked by PC/TLC/HPLC. (When the decarboxylation is incomplete in a closed distillation, the distillate can be poured into the reactor and dissolved by heating and stirring. , and then performing closed distillation to increase the yield of the demethylated product), the distillation product is heated and dissolved by adding alcohol, and phosphoric acid is added to precipitate luteolin;

Pyridines, trivalent aluminum ion compounds, low boiling alcohols, hesperetin-7-O-glucoside, iodine mixed, dehydrogenation at room temperature -110 ° C to form a homogeneous solution, PC / TLC / HPLC tracking starting products Dehydrogenation is complete. Then, it is sealed and sealed at 80 ° C - 180 ° C for 2-80 h. The demethylation product is traced by PC/TLC/HPLC. The distilled product is dissolved by heating with alcohol, and phosphoric acid is added to precipitate luteolin and geranyl-7-O-glucoside. , luteolin and geranyl-7-O-glucoside can be obtained by chromatography;

Pyridines, trivalent aluminum ion compounds, low boiling alcohols, hesperidin, and iodine are mixed and dehydrogenated at room temperature -110 ° C to form a homogenous liquid, and the dehydrogenation of the starting product is completely followed by PC/TLC/HPLC. Then, it is sealed and sealed at 80 ° C - 180 ° C for 2-80 h. The demethylation product is traced by PC/TLC/HPLC. The distilled product is dissolved by heating with alcohol, phosphoric acid is added, and the precipitate is precipitated (Dioxing). Acid-7-rutinoside acid solution: passed Macroporous adsorption resin adsorption, water elution and impurity removal, ethanol elution, recovery of ethanol, can obtain luteolin-7-rutinoside; luteolin-7-rutinoside acid water can be obtained by acid hydrolysis, luteolin And a mixture of luteolin, which can be obtained by silica gel column chromatography to obtain luteolin and luteolin monomer; the acid water of luteolin-7-rutino is completely hydrolyzed in alcohol to obtain luteolin.

The principle of the invention:

Dehydrogenation and demethylation of hesperetin or its glycoside derivatives to prepare luteolin or its glycosides, the mechanism is the same, 4-C=O, 5-OH in the molecule is complexed with Al ³⁺ , and is easily soluble in pyridine In the alcohol liquid, dehydrogenation by heating, the dehydrogenation product is also complexed with Al ³⁺ , still exists in the form of a solution, and the methanol and pyridine are exemplified by the closed distillation of the alcohol liquid and the excess pyridine. The process produces HI and HI and pyridine salts, in which I ions are highly polarizable and easy to attack C+ ions in OCH ₃ ; on the other hand, Al ³⁺ is complexed with ortho-dioxy to facilitate OCO _{3 de-} arming Base, two aspects of synergy, demethylation is easier and more selective. The two reactions of dehydrodemethylation can be carried out directly in the same vessel without separation.

The pyridines referred to in the present invention mainly refer to pyridine, 2-methylpyridine, 3-methylpyridine, 4-methylpyridine, 4-methylpyridine, 5-methylpyridine, 6-methylpyridine, 2 , 6-lutidine, 3,5-lutidine, 2,4,6-trimethylpyridine.

The trivalent aluminum ion compound in the present invention mainly refers to aluminum trichloride, aluminum tribromide, aluminum triiodide, aluminum methoxide, aluminum ethoxide, aluminum propoxide, etc., and preferably anhydrous aluminum trichloride, wherein Aluminum halide: orange peel The molar ratio of glycoside (or hesperetin or hesperetin-7-O-glucoside) is from 3:1 to 1:3, preferably 1:1.

The low-boiling alcohol in the present invention means methanol, ethanol, propanol or isopropanol, of which methanol is preferred, and the amount of methanol used is not more than the volume of the pyridine.

Hesperidin (CAS: 520-26-3), hesperetin (CAS: 520-33-2), hesperetin-7-O-glucoside (CAS:31712-49-9) in the present invention , the structure is as (A)) as the dehydrogenation and de-starting reactant, wherein the hesperidin (or hesperetin or hesperetin-7-O-glucoside): pyridine is in a molar ratio of 1:5-1 : 10, preferably 1:6-1:7.

In the present invention, the iodine, the amount thereof and the hesperetin (or hesperidin or hesperetin-7-O-glucoside) molar ratio are preferably 1:1.

In the closed distillation according to the present invention, the distillation temperature is from 80 ° C to 180 ° C, preferably from 90 ° C to 150 ° C. The closed state means that outside air cannot enter the reactor during the distillation, but the air in the reactor can be discharged, and the alcohol And pyridine vapors can be condensed and discharged.

The luteolin (CAS: 491-70-3), luteolin (CAS: 5373-11-5), and luteolin-7-O-rutinoside (CAS: 20633-84-5, The structure is as in (B)).

In the present invention, the dehydrogenation and deprotection are combined into one step, wherein since it is a solution type, the yield of dehydrogenation to produce geranin and the derivative is 100%, and the dislocation is directly separated without separation, and the geranyl-7-O - Demethylation of rutinoside (Diomin) to prepare luteolin-7-O-rutinoside, yield 60%-75%, luteolin-7-O- The rutinoside has high water solubility, and there is a certain loss in the enrichment and purification process, and the yield will decrease; the geranyl-7-O-glucoside demethylation yield is between 50% and 65%, and its water solubility It is less soluble, can be precipitated and precipitated, and has high yield; lignan is demethylated by double distillation to prepare luteolin. The yield is above 95%. Luteolin is insoluble in water and is easy to be purified.

Since hesperetin and its glycoside derivatives are complexed with aluminum, the dehydro-demethylated products are easily soluble in the alcohol solution of pyridine, so the amount of pyridine is particularly small. The method and the existing patents and reports of hesperidin Semi-synthetic luteolin has significant improvements. Dehydro-demethylation of hesperidin or hesperetin-7-O-glucoside to prepare luteolin glycoside is a breakthrough invention. Semi-synthetic route of luteolin, luteolin, luteolin and rutin, with mild reaction conditions and easy operation; low reagent dosage and low cost; few steps, high dehydrogenation and demethylation yield, easy industrial production . Among them, the production of luteolin-7-O-rutinoside is preferred to dehydro-demethylation in one step of hesperidin; the production of luteolin preferentially dehydrogenates in one step of hesperetin-7-O-glucoside Demethylation; luteolin is recommended for dehydrogenation in a one-step process, followed by several times of closed distillation demethylation.

In the present invention, Dioxin has been directly demethylated, but in the alcohol and pyridine, the dissolution rate with AlCl ₃ is very slow, and it is difficult to form a uniform solution, and the demethylation effect is not good; likewise, hesperidin and In the alcohol and pyridine, the demethylation yield of the complex with AlCl ₃ is very low, and it is mainly subjected to a long-term distillation and is mainly a de-sugar product. The luteolin-7-O-rutinoside is highly polar, and it is difficult to be adsorbed by the macroporous adsorption resin D101 type, AB-8, polyamide or the like in the acid aqueous solution of pyridine. After repeated screening, T-02, T-01, HPD-D, DM130, HPD400, and HPD500 have certain adsorption capacity for luteolin rutinoside. Hesperidin is prepared by hesperidin. If the method of patent No. 201310692053.X is used, the preparation of hesperidin is extremely low. Hesperidin-7-O-glucoside is prepared by hesperidin. If the method is patent No. 201310692053.X or 201410051283.2, hesperetin-7-O-glucoside is inexpensive to prepare.

When the closed distillation is carried out, if the distillation temperature is too high, the glycoside is dehydrated. At the same time, the pyridine or alcohol is too fast, the solution becomes solid faster, the demethylation ability will prolong with time, the demethylation ability gradually decreases, and the effect is not good. When the alcohol or pyridine is distilled at a low temperature and then at a high temperature, expansion phenomenon occurs (the bottom solid gradually expands, and the volume expands the original volume several times or even several tens of times, possibly overflowing the bottle). The closed distillation time is too long, and if there is sugar in the molecule, the de-sugar product increases.

The present invention will be further described below in conjunction with the embodiments.

The invention discloses a method for one-step dehydrogenation and demethylation of hesperetin, hesperetin-7-O-glucoside and hesperidin, and those skilled in the art can learn from the contents of this paper to screen the types of pyridines and alcohols. , optimizing the demethylation temperature and the amount of alcohol in the closed distillation. In particular, it is necessary to point out that the process of dehydro- and demethylation of hesperetin and its glycosides to prepare luteolin and its glycosides is accomplished by a one-step method of complexation with aluminum ions, which will be regarded as the present invention, and the relevant personnel obviously cannot The methods and principles described herein are appropriately modified or altered and combined to achieve and apply the techniques of the present invention without departing from the spirit, scope, and scope of the present invention; since the alcohol, pyridine, and trivalent aluminum ion compounds involved are more For the preparation of luteolin and its glycosides, can not be enumerated. However, in order to further understand the present invention, luteolin-7-O-rutinoside is prepared by dehydro-demethylation of hesperidin, and luteolin and luteolin, hesperetin-7-O-glucoside are prepared by hydrolysis. Dehydro-demethylation to prepare luteolin, hesperidin dehydro-demethylation to prepare luteolin, alcohol as methanol, pyridine, pyridine, trivalent aluminum ion compound with anhydrous AlCl ₃ as an example, Wherein the reagent AR indicates analytical purity.

Example 1 Preparation of luteolin-7-O-rutinoside by hesperidin

Quickly weigh anhydrous aluminum trichloride (AR) 3.5g in a 250ml iodine flask, quickly add 15ml of pyridine (AR), shake well, add 8ml of methanol (AR), 2min, add 92% hesperidin 16.5g Stir for 2min, add iodine (AR) 6.8g, add 7ml of methanol (AR), stir for 2min, end the iodine bottle with a condenser or air condensing tube, and seal at 80 °C (prevent the absorption of moisture in the air) every 1h After stirring for 1 time, a homogeneous solution was formed after 2 hours, and the closed reaction was continued for 6 hours (dehydrogenation was completed). In an air bath, sealed at 114 ° C for 15 h (demethylation reaction, the solution changed from solution to solid, soft when hot, very hard when cooled), placed at 80 ° C, added 10 ml of ethanol and 10 ml of glycerol, and After incubating at 80 °C for 2 h, the solid was converted into a solution, and 1.0 g of the insurance powder was added. After stirring for 5 min, it was poured into 200 ml of 5% phosphoric acid water solution, rapidly stirred, and sealed in a water bath at 50 ° C for 1 h, filtered (insoluble matter for the ground) Omin, about 4.8g after drying, get acid water, pass 350g macroporous adsorption resin, wash with water to remove pyridine, HI, phosphoric acid, glycerol, ethanol, AlCl ₃ , elute with ethanol, recover ethanol to get rhododendron The oxacin-7-O-rutinoside was 10.8 g, and the purity was determined by HPLC to be 90.8% (with 98% luteolin as an internal standard). ¹³ C NMR (DMSO-d6, 100 MHz): d 182.08 (s, C-4), 164.74 (s, C-2), 163.01 (s, C-7), 161.37 (s, C-5), 157.08 ( s, C-9), 150.12 (s, C-4'), 145.92 (s, C-3'), 121.47 (s, C-1'), 119.40 (d, C-6'), 116.32 (d , C-5'), 113.73 (d, C-2'), 105.53 (s, C-10), 103.28 (d, C-3), 100.65 (d, Gly-1), 99.99 (d, C- 6), 94.95 (d, C-8), 76.42 (t, Gly-5), 75.67, (s, Gly-3), 73.25 (d, Gly-2), 69.70 (d, Gly-4), 66.15 (t, Gly-6).99.66 (Rha-1), 70.28 (Rha-2), 70.88 (Rha-3), 72.20 (Rha-4), 68.50 (Rha-5) 17.99.50 (Rha-6) ; TOF MS ES+m/z 617.1464 [M+Na] ⁺ , m/z 449.1066 [M-146+H] ⁺ , m/z 287.0550 [M-146-162+H] ⁺ . TOF MS ES-m/z 593.1527[MH] ^- , m/z285.0414[M-146-162-H] ^- , (calc.for.C ₂₇ H ₃₀ O ₁₅ ) (90.8% of the sample for structural identification) Luteolin-7-O-rutinoside was separated on a silica gel column).

Example 2 Preparation of hesperidin to produce luteolin-7-O-rutinoside by acid hydrolysis to obtain luteolin and luteolin

Quickly weigh anhydrous aluminum trichloride (AR) 3.5g in a 250ml iodine flask, quickly add 15ml of pyridine (AR), shake well, add 8ml of methanol (AR), 2min, add 92% hesperidin 16.5g Stir for 5min, add iodine (AR) 6.8g, add 7ml of methanol (AR), stir for 3min, end the iodine bottle with a condenser or air condensing tube, and seal at 80 °C (to prevent absorption of moisture in the air) every 1h After stirring for 1 time, a homogeneous solution was formed after 2 hours, and the closed reaction was continued for 6 hours (dehydrogenation was completed). In an air bath, sealed at 112 ° C for 17 h (demethylation reaction, the solution changed from solution to solid, soft when hot, very hard when cooled), placed at 80 ° C, added 20 ml of ethanol, and kept at 80 ° C for 3 h, The solid is converted into a solution, and 1.0 g of the powder is added. After stirring for 5 minutes, it is poured into 100 ml of 5% phosphoric acid water solution, rapidly stirred, and sealed in a water bath at 50 ° C for 1 h, filtered (insoluble matter is deomin, dried) After about 4.7g), the acid water solution was added, 20ml of hydrochloric acid was added, and the mixture was hydrolyzed at 80 ° C for 3 hours to precipitate a precipitate. The mixture was filtered to obtain a mixture of luteolin and luteolin, which was separated by silica gel column chromatography to obtain luteolin 3.0. g, luteolin 3.2g.

Example 3 Preparation of hesperidin Luteolin-7-O-rutinoside was hydrolyzed by acid alcohol solution, and luteolin was quickly weighed into anhydrous aluminum chloride (AR) 3.5 g in 250 ml iodine flask, and quickly added. 15ml of pyridine (AR), shake well, add 8ml of methanol (AR), after 2min, add 16.5g of 92% hesperidin, stir for 5min, add iodine (AR) 6.8g, add methanol (AR) 7ml, stir for 3min, iodine The measuring bottle is terminated with a condensing tube or an air condensing tube, and the reaction is sealed at 80 ° C (to prevent absorption of moisture in the air), and stirred once every 1 hour. After 2 hours, a uniform solution is formed, and the sealing reaction is continued for 6 hours (complete dehydrogenation). In an air bath, sealed at 116 ° C for 15 h (demethylation reaction, the solution changed from solution to solid, soft when hot, very hard when cooled), placed at 80 ° C, added 20 ml of ethanol, and kept at 80 ° C for 3 h, The solid is converted into a solution, and 1.0 g of the powder is added. After stirring for 5 minutes, it is poured into 100 ml of 5% phosphoric acid water solution, rapidly stirred, and sealed in a water bath at 50 ° C for 1 h, filtered (insoluble matter is deomin, dried) After about 5.0 g), an aqueous acid solution was obtained, 100 ml of ethanol was added, 30 ml of hydrochloric acid was added, and the mixture was hydrolyzed at 80 ° C for 5 hours to precipitate a precipitate, which was filtered to obtain 5.6 g of luteolin. UV λ max nm in MeOH: 346, 258; + NaOMe 407, 267; + AlCl ₃ 422, 320 sh, 265; + AlCl ₃ / HCl 390, 375, 273.

Example 4 Preparation of luteolin by hesperetin-7-O-glucoside

Quickly weigh anhydrous aluminum trichloride (AR) 3.5g in a 250ml iodine volumetric flask, quickly add 15ml of pyridine (AR), shake well, add methanol (AR) 8ml, 2min, add 92% hesperidin 12g, Stir for 5min, add iodine (AR) 6.8g, add 7ml of methanol (AR), stir for 3min, end the iodine bottle with a condenser or air condensing tube, seal at 80 °C (prevent the absorption of moisture in the air), stir every 1h One time, a homogeneous solution was formed after 2 hours, and the closed reaction was continued for 6 hours (complete dehydrogenation). In an air bath, sealed at 113 ° C for 15 h (demethylation reaction, the solution changed from solution to solid, soft when hot, very hard when cooled), placed at 80 ° C, added 20 ml of ethanol, and kept at 80 ° C for 3 h, The solid was converted into a solution, and 1.0 g of the powder was added. After stirring for 5 minutes, it was poured into 100 ml of 5% phosphoric acid aqueous solution, rapidly stirred, and sealed in a water bath at 50 ° C for 1 h, and filtered to obtain luteolin and geranin-7. The mixture of -O-glucoside was separated by silica gel column chromatography to obtain 5.7 g of luteolin, and the purity was 94% by HPLC. ¹ H NMR (DMSO-d6, 400 MHz): δ 13.01 (1H, s, 5-OH), 10.05 (1H, s, 4'-OH), 9.45 (1H, s, 3'-OH), 7.45 (1H) , dd, J=8.4/2.2, H-6'), 7.44 (1H, d, J=2.2, H-2'), 6.80 (1H, d, J=8.4, H-5'), 6.77 (1H , dd, J = 1.8, H-8), 6.77 (1H, s, H-3), 6.46 (1H, dd, J = 1.8, H-6), 5.11 (1H, d, J = 7.3, H- 1′′), 4.06 (1H, d, J=9.5, H-6′′), 3.88 (1H, dd, J=3.5/1.6 Hz, H-2′′′), 3.69, (1H, dd, J=9.5 /3.3 Hz, H-3"'), 3.18 - 3.68 (7H, m, H-2", 3", 4", 5", 6" a, 4", 5"'), 1.10 (3H, d, J = 6.2 Hz, H-6"'); ¹³ CNMR (DMSO-d6, 100 MHz): d 181.97 (s, C-4), 164.50 (s, C-2), 162.97 (s, C- 7), 161.18 (s, C-5), 156.99 (s, C-9), 149.97 (s, C-4'), 145.83 (s, C-3'), 121.42 (s, C-1') , 119.23 (d, C-6'), 116.01 (d, C-5'), 113.60 (d, C-2'), 105.37 (s, C-10), 103.31 (d, C-3), 99.85 (d, Gly-1), 99.57 (d, C-6), 94.74 (d, C-8), 77.18 (t, Gly-5), 76.42, (s, Gly-3), 73.14 (d, Gly -2), 69.53 (d, Gly-4), 60.63 (t, Gly-6); TOF MS ES+m/z 471.0909 [M+Na] ⁺ , m/z 449.1145 [M+H] ⁺ , m/z 287.0571 [M-162+H] ⁺ . TOF MSES-m/z 447.0922 [MH] ^- , m/z 285.0401 [M-162-H] ^- , (calc. for. C ₂₁ H ₂₀ O ₁₁ ).

Example 5 Preparation of luteolin by hesperetin

Quickly weigh anhydrous aluminum trichloride (AR) 3.5g in a 250ml iodine volumetric flask, quickly add 15ml of pyridine (AR), shake well, add methanol (AR) 8ml, 2min, add 98% hesperetin 8.5g Stir for 5min, add iodine (AR) 6.8g, add 7ml of methanol (AR), stir for 3min, end the iodine bottle with a condenser or air condensing tube, and seal at 80 °C (to prevent absorption of moisture in the air) every 1h After stirring for 1 time, a homogeneous solution was formed after 2 hours, and the closed reaction was continued for 6 hours (dehydrogenation was completed). In an air bath, sealed at 113 ° C for 15 h (demethylation reaction, the solution changed from solution to solid, soft when hot, very hard when cooled), placed at 80 ° C. The distillate was poured into an iodine measuring flask and incubated at 80 ° C for 3 h. The solid was converted into a solution, and then sealed in an air bath at 116 ° C for 15 h (second demethylation), placed at 80 ° C, and added with 100 ml of ethanol. Dissolve, quickly add 20 ml of phosphoric acid, stir rapidly, and filter at 70 ° C for 30 min. The filter cake was washed with water and dried to obtain 7.5 g of luteolin. The purity was 98.1% by HPLC. ¹ H-NMR (DMSO d6, 400 MHz): δ 7.41 (1H, bd, J = 8.4 Hz, H 6 '), 7.39 (1H, bs, H2'), 6.89 (1H, d, J = 8.4 Hz, H 5 '), 6.61 (1H, s, H ₃ ), 6.44 (1H, d, J = 2 Hz, H 8), 6.19 (1H, d, J = 2 Hz, H 6); ¹³ C-NMR (DMSO) D6,100 MHz): δ182.1 (C-4), 164.5 (C-7), 164.4 (C-2), 161.9 (C-5), 157.7 (C-9), 150.1 (C-4') , 146.2 (C-3'), 122.0 (C-1'), 119.4 (C-6'), 116.5 (C-5'), 113.8 (C-2'), 104.2 (C-10), 103.3 ( C-3), 99.3 (C-6) 94.3 (C-8); EIMS (40 eV.): m/z% 286 (100), 152 (10) 137 (15); UV λ max nm in MeOH: 346, 258; NaOMe 407, 267; + AlCl ₃ 422, 320 sh, 265; + AlCl ₃ / HCl 390, 375, 273.

Claims

A method for semi-synthesis of luteolin, luteolin, and luteolin rutinoside, specifically characterized by three pathways A, B, and C: A pathway: pyridine, trivalent aluminum ion compound, low boiling alcohol, hesperetin, Iodine mixing, dehydrogenation by heating at 20-110 °C to form a homogenous liquid, PC/TLC/HPLC to trace the dehydrogenation of the starting product completely; then sealed at 80 ° C -180 ° C for 2-80 h, PC / TLC / HPLC follow-up inspection Demethylation product (when the demethylation in a closed distillation is incomplete, the distillate can be poured into the reactor, dissolved by heating, and then subjected to closed distillation to increase the yield of the demethylated product), and the distillation product is obtained. Add alcohol to dissolve, add phosphoric acid, and precipitate luteolin;

Route B: pyridine, trivalent aluminum ion compound, low boiling point alcohol, hesperetin-7-O-glucoside, iodine mixed, dehydrogenation by heating at 20-110 ° C to form a homogeneous solution, PC / TLC / HPLC The starting product is completely dehydrogenated; then it is sealed and sealed at 80 ° C - 180 ° C for 2-80 h. The demethylation product is traced by PC/TLC/HPLC. The distilled product is heated and dissolved by alcohol, and phosphoric acid is added to precipitate luteolin and geranium. -7-7-O-glucoside, which can be obtained by chromatography to obtain luteolin and geranyl-7-O-glucoside;

Route C: pyridine, trivalent aluminum ion compound, low boiling point alcohol, hesperidin, iodine mixed, dehydrogenation by heating at 20-110 ° C to form a homogenous liquid, PC/TLC/HPLC to trace the dehydrogenation of the starting product completely Then, it is sealed and sealed at 80 ° C - 180 ° C for 2 - 80 h. The demethylation product is traced by PC/TLC/HPLC. The distilled product is heated and dissolved by alcohol, phosphoric acid is added, and the precipitate is precipitated (Dioxing). Oxalate-7-rutinoside acid solution: adsorption by macroporous adsorption resin, water elution and impurity removal, ethanol elution, recovery of ethanol, luteolin-7-rutinoside; luteolin-7-rutin The acid water solution can be obtained by acid hydrolysis, and a mixture of luteolin and luteolin can be obtained, and the luteolin and luteolin monomers can be obtained by silica gel column chromatography; the acid water of luteolin-7-rutinoside is obtained. Fully hydrolyzed in the alcohol to obtain luteolin.
According to claim 1, it is characterized in that the pyridines in the present invention mainly refer to pyridine, 2-methylpyridine, 3-methylpyridine, 4-methylpyridine, 4-methylpyridine, 5- Methylpyridine, 6-methylpyridine, 2,6-lutidine, 3,5-lutidine, 2,4,6-trimethylpyridine, of which pyridine is preferred.
According to claim 1, it is characterized in that the trivalent aluminum ion compound in the present invention mainly refers to aluminum trichloride, aluminum tribromide, aluminum triiodide, aluminum methoxide, aluminum ethoxide, aluminum propoxide, and the like. Preferred is anhydrous aluminum trichloride, wherein the molar ratio of aluminum halide: hesperidin (or hesperetin or hesperetin-7-O-glucoside) is from 3:1 to 1:3, with a priority of 1: 1.
According to claim 1, it is characterized in that the low boiling point alcohol in the present invention means methanol, ethanol, Propanol, isopropanol, the amount of methanol is preferred, and the amount of methanol used is not more than the volume of the pyridine.
According to claim 1, it is characterized by the hesperidin (CAS: 520-26-3), hesperetin (CAS: 520-33-2), and hesperetin-7-O according to the present invention. - Glucosin (CAS: 31712-49-9, structure (A)) as a dehydrogenation and de-starting reactant, wherein hesperidin (or hesperetin or hesperetin-7-O-glucoside): The molar ratio of the pyridines is from 1:5 to 1:10, preferably from 1:6 to 1:7.
It is characterized in that, in the present invention, the iodine, the amount thereof and the hesperetin (or hesperidin or hesperetin-7-O-glucoside) molar ratio are preferably 1:1.
According to claim 1, it is characterized in that the closed distillation in the present invention means that the distillation temperature is in the range of 80 ° C to 180 ° C, preferably 90 ° C to 150 ° C. The sealing means that outside air cannot enter the reactor during the distillation process. Medium, but the air in the reactor can be discharged, and the alcohol and pyridine vapor can be condensed and discharged.
According to claim 1, it is characterized by: luteolin (CAS: 491-70-3), luteolin (CAS: 5373-11-5), luteolin-7-O- as described in the present invention. The rutinoside (CAS: 20633-84-5, structure (B)) was prepared by one-step dehydrogenation and demethylation, respectively.