CN112353842A - Rabdosia rubescens stem water extract powder, preparation thereof, free radical scavenging effect and application - Google Patents

Rabdosia rubescens stem water extract powder, preparation thereof, free radical scavenging effect and application Download PDF

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CN112353842A
CN112353842A CN202011231955.XA CN202011231955A CN112353842A CN 112353842 A CN112353842 A CN 112353842A CN 202011231955 A CN202011231955 A CN 202011231955A CN 112353842 A CN112353842 A CN 112353842A
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hydroxy
carboxy
cinnamoyl
quinic acid
rabdosia rubescens
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彭师奇
赵明
黄飞
卢斌
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Beijing Hengrun Taisheng Pharmaceutical Technology Co ltd
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Beijing Hengrun Taisheng Pharmaceutical Technology Co ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/53—Lamiaceae or Labiatae (Mint family), e.g. thyme, rosemary or lavender
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P39/00—General protective or antinoxious agents
    • A61P39/06—Free radical scavengers or antioxidants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/30—Extraction of the material
    • A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
    • A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation, decoction
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
    • A61K2236/50—Methods involving additional extraction steps
    • A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying

Abstract

The invention discloses rabdosia rubescens stem aqueous extract powder, a preparation method thereof and application thereof in removing free radicals. The rabdosia rubescens stem water extract powder prepared by the method contains 17 quinic acid compounds and 11 quercetin compounds. Experiments prove that the rabdosia rubescens stem water extract powder has excellent free radical scavenging effect. Therefore, the invention provides an effective technical means for resisting oxidative stress.

Description

Rabdosia rubescens stem water extract powder, preparation thereof, free radical scavenging effect and application
Technical Field
The invention relates to a method for preparing rabdosia rubescens stem water extract powder, and relates to application of the rabdosia rubescens stem water extract powder prepared by the method in preparation of an anti-oxidative stress medicament. The invention belongs to the field of biological medicine.
Background
Oxidative stress is the tilt of the balance between oxidation and antioxidation in the body, resulting in inflammatory infiltration of neutrophils, increased secretion of proteases and generation of largeStress phenomenon of oxidation intermediate. In oxidative stress, free radicals accumulate in the body and have negative health effects. Free radical accumulation in the body is generally considered to be one of the important factors leading to deterioration of cell function, aging, and even death. Free radicals include reactive oxygen species free Radicals (ROS) and reactive nitrogen species free Radicals (RNS). The active oxygen radical being a superoxide anion (. O)2 -) Hydroxyl radical (. OH) and hydrogen peroxide (H)2O2) Is a general term for (1). The active nitrogen radical is nitrogen monoxide radical (NO.), nitrogen dioxide radical (NO.)2) And peroxynitrite radical (. ONO)2 -) Is a general term for (1). Free radical accumulation in the body refers to the accumulation of ROS and RNS. Free radicals are exceptionally reactive molecules that can react with all cellular components. Under normal conditions, organs in physiological response produce free radicals. Too high a concentration of free radicals causes a loss of balance in cell function and morphology. Primary radicals generated in the cell further expand into secondary radicals when the equilibrium moves in the direction of oxidation. Cellular proteins are ideal targets for secondary free radical attack. Secondary free radicals target cellular proteins and trigger biological damage to cells.
Numerous studies have demonstrated that free radicals are an important pathological indication in the pathogenesis of many human diseases. Free radicals randomly oxidize proteins or polypeptides to side chain carbonyl derivatives to inactivate them. The degree of protein carbonylation was used to quantify the degree of functional senescence in human dermal fibroblasts, lens and brain tissue. While there is no direct evidence to associate free radicals with the etiology of degenerative diseases of the central nervous system (e.g., motor neuron disease, cerebellar ataxia and parkinson's disease), indirect evidence leads to the hypothesis that free radicals contribute to the pathogenesis of these diseases.
Oxidative stress is involved in the pathological process of acute and chronic liver diseases by inducing hepatocyte dysfunction. Although free radicals are derived from the metabolism of normal cells, the metabolism of cells in liver diseases overproduces free radicals. The overproduced free radicals cross the antioxidant defenses and cause liver damage. Among liver diseases, the incidence of nonalcoholic fatty liver diseases characterized by developing oxidative stress and breaking redox balance is the highest. The pathology of nonalcoholic fatty liver disease belongs to a multifactorial pathology, including alterations in lipid metabolism, mitochondrial dysfunction, inflammation and oxidative stress.
Cardiovascular diseases including ischemic heart disease and cerebrovascular disease are the top among the morbidity and mortality of diseases on a global scale. Atherosclerosis is the major preferential causative agent of most cardiovascular diseases. It is well understood that oxidative stress and inflammation are the two main mechanisms responsible for atherosclerosis. Under oxidative stress conditions, higher unsaturated fatty acid-containing phospholipids and cholesterol esters in cell membranes and ester proteins are readily oxidized to complex mixed oxidation products in a free radical-induced peroxidation process of esters. Most evidence suggests that these oxidized lipids are active in the inflammatory response of atherosclerosis by interacting with immune cells (e.g., macrophages) and endothelial cells.
Sometimes tumor onset is broken down into three steps, initiation, acceleration and spread. In the priming step, the carcinogens modulate the genetic identity of the cells and allow the cells to divide rapidly. At the acceleration step, the genetically damaged cells fail to lock into the genetic lesion and contribute to unregulated cell growth. In the epidemic step, the cell system growing without control establishes a blood supply network through angiogenesis and causes tumor and carcinogenesis to grow. There is evidence that oxidative stress, chronic inflammation and tumors are closely associated. Free radical induced oxidative stress is prevalent in many types of tumor cells associated with altered redox signaling pathways. Unlike normal cells, oxidative stress induces a loss of balance in the redox of various tumor cells. In this sense, redox imbalance may be associated with an oncogenic stimulus. DNA mutation is a critical step in the carcinogenic process, while an increased level of oxidative DNA damage is observed in a variety of tumors. It can be seen that such lesions are strongly implicated in the etiology of tumors. Increased production of reactive oxygen radicals and altered redox status in tumor cells has been observed early, while tumor cells of advanced tumors often show multiple genetic variations and strong oxidative stress.
Primary radicals generated in cells and tissues under oxidative stress further expand to become secondary radicals. Proteins in cells and tissues are ideal reactants for secondary free radicals. Secondary free radicals target these proteins and initiate biological damage to cells and tissues. Oxidative stress-induced biological damage to cells and tissues has led to neonatal morbidity, particularly in premature infants for which antioxidant defense systems have not yet been established. Mitochondria are not only the subject of oxidative damage, but also tend to be involved in oxidative self-damage in the onset of neonatal lung disease. Similarly, both intrauterine growth restriction and giant fetuses are associated with mitochondrial dysfunction due to oxidative stress. In addition, the ability of the lamb to produce pro-inflammatory cytokines is significantly enhanced by the pure oxygen-dominated reoxygenation on hypoxic ischemic newborn lamb models.
In summary, oxidative stress, which is characterized by the accumulation of free radicals, is the cause of a number of major common diseases. The invention is safe and effective free radical scavenging medicine and can provide effective technical means for scavenging free radicals.
Rabdosia rubescens is whole plant of Rabdosia rubescens (Hemsl.) Hara of Rabdosia of Labiatae. Rabdosia rubescens is a perennial herb, widely distributed in the Yangtze river basin of the yellow river of China, and mainly produced in the areas of the south Henan Jiyuan Taihang mountain and the Wang House mountain. Because of the functions of clearing away heat and toxic material, clearing and nourishing throat, and relieving swelling and pain, the rabdosia rubescens whole herb is boiled in water for drinking all the time in folk. In the past 30 years, the research on the chemical components and pharmacological action of rabdosia rubescens mainly focuses on the fat-soluble parts of leaves and stems, and the representative components are rubescensin A and rubescensin B. To date, there has been no study concerning the chemical composition and therapeutic effect of aqueous extracts of Rabdosia Rubescens (Hemsl.) Hara stems. The Rabdosia Rubescens (Hemsl.) Hara has high stem content compared with leaves. Effective application of Rabdosia rubescens stems is of great significance.
The inventor has long conducted research on chemical components and therapeutic effects of the rabdosia rubescens stem aqueous extract powder. The inventor carries out mass spectrometry on chemical components of rabdosia rubescens stem water extract powder, and finds 28 unreported components which mainly comprise cinnamoyl quinic acid and multi-substituted quercetin. Although the biological activity of single components of an aqueous extract powder of rabdosia rubescens stems has been reported, the biological activity of the combination of the whole components is unknown. Through repeated studies, the inventors found that these 28 components can effectively scavenge free radicals. Thus, the rabdosia rubescens stem aqueous extract powder of the present invention can prevent oxidative stress. Based on these findings, the inventors have proposed the present invention.
Disclosure of Invention
The technical problem to be solved by the invention is to provide a method for preparing rabdosia rubescens stem aqueous extract powder and the rabdosia rubescens stem aqueous extract powder prepared by the method. Experiments prove that the rabdosia rubescens stem water extract powder prepared by the invention has the function of removing free radicals. In order to achieve the purpose, the invention adopts the following five technical means.
The first technical means is to provide a rabdosia rubescens stem water extract powder of the invention, which contains 17 kinds of quinic acid compounds and 11 kinds of quercetin compounds, wherein the 17 kinds of quinic acid compounds are 3-O- [ E-2,4, 6-tri (3-carboxyl-2-acryloyloxy) -5- (3-hydroxyl-2-acryloyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (6-hydroxyl-1, 3, 5-hexatrienylcarbonyloxy) -4- (4-carboxyl-1, 3-butadienylcarbonyloxy) -5-methoxy-6- (3-hydroxyl-1-acryloylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-bis (4-carboxy-1-butenylcarbonyloxy) -3-hydroxy-5-methoxy-6- (6-carboxy-1, 3-hexadienylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-hydroxy-1, 3-butadienylcarbonyloxy) -4- (3-hydroxyacryloyloxy) -5-hydroxy-6- (3-hydroxy-1-propenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (5-hydroxy-1-pentenylcarbonyloxy) -3-methoxy-4- (3-carboxy-1-propenylcarbonyloxy) -5- (3-hydroxypropanoyloxy) -6- (3-hydroxy-propenylcarbonyloxy) - (5-carboxy-1-pentenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 5-dimethoxy-3-hydroxy-4- (4-carboxybutylcarbonyloxy) -6- (5-carboxypentylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutylcarbonyloxy) -3-hydroxy-4- (3-carboxypropylcarbonyloxy) -5-formyloxy-6- (4-carboxy-1-butenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2-formyloxy-3, 4-bis (5-carboxy-1-pentenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (7- Carboxy-1-heptenecarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2-carboxyacetoxy-3, 6-bis (5-carboxy-1-pentenylcarbonyloxy) -4- (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-heptenecarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-bis (4-carboxy-1-butenylcarbonyloxy) -3-hydroxy-5- (5-hydroxypentylcarbonyloxy) -6- (5-hydroxy-1, 3-pentadienylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutylcarbonyloxy) -3-hydroxy-4- (4-carboxybutylcarbonyloxy) -1-butenylcarbonyloxy) -5, 6-bis (5-hydroxy-1-pentenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-bis (4-carboxybutylcarbonyloxy) -3-hydroxy-5- (3-hydroxypropanoyloxy) -6-carboxyacetyloxy cinnamoyl ] quinic acid, 3-O- [ E-2-formyloxy-3- (5-carboxy-1-pentenylcarbonyloxy) -4- (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (5-hydroxypentylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 5-dihydroxymethoxy-4- (3-carboxypentanoyloxy) -6- (5-carboxypentanoyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (5-hydroxypentanoyloxy) -3-hydroxy-4- (4-carboxybutanecarbonyloxy) -5, 6-dicarboxyacetyloxy cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutanecarbonyloxy) -4, 6-bis (4-carboxy-1-butenoyloxy) -5- (3-carboxypropanoyloxy) cinnamoyl ] quinic acid and 3-O- [ E-2- (3-carboxypropanoyloxy) -3- (5-carboxy-1-pentenoyloxy) -4, 6-bis (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) cinnamoyl ] quinic acid.
The 11 kinds of quercetin compounds are 3-O-glucoside-3 '- (6-hydroxyhexylcarbonyloxy) -4' -methoxy-5 '- (5-hydroxypentanoyloxy) quercetin, 3-O-glucoside-2' -hydroxy-3 '-hydroxymethoxy-4' -hydroxyacetoxy-5 ', 6' -dicarboxyacetyloxy quercetin, 3-O-glucoside-2 ', 6' -dihydroxymethoxy-3 '- (3-hydroxypropionyloxy) -4' -hydroxy-5 '- (3-carboxypropanoyloxy) quercetin, 3-O-glucoside-2', 6 '-dimethoxy-3' - (5-hydroxy-1, 3-pentadienecarbonyloxy) -4 '- (3-hydroxy-1-propenylcarbonyloxy) quercetin, 3-O-glucoside-3' - (4-carboxy-1-butenylcarbonyloxy) -4 '- (4-carboxybutylcarbonyloxy) quercetin, 3-O-glucoside-2' -hydroxy-4 '- (3-carboxypropenyloxy) -5' - (3-carboxypropylcarbonyloxy) quercetin, 3-O-glucoside-2 '-methoxy-4' - (3-carboxy-1-propenylcarbonyloxy) -5 '- (5-hydroxy-1-pentenylcarbonyloxy) -6' - (7-hydroxy-1, 3-heptadienylcarbonyloxy) quercetin, 3-O-glucoside-2 ' -methoxy-4 ', 5 ' -dihydroxymethoxy-6 ' - (7-hydroxyheptacarbonyloxy) quercetin, 3-O-glucoside-2 ', 3 ' -diformyloxy-4 ' - (4-carboxy-1-butenylcarbonyloxy) -5 ' - (3-carboxypropylcarbonyloxy) -6 ' - (4-hydroxy-1-butenylcarbonyloxy) quercetin, 3-O-glucoside-2 ' -hydroxy-3 ', 5 ' -dihydroxymethoxy-4 ' -carboxyacetoxy-6 ' - (3-carboxypropyloxy) quercetin and 3-O-glucoside-2 ', 3 ', 4' -Tricarbonyloxy-5 '- (4-hydroxy-1, 3-butadienecarbonyloxy) -6' -carboxyacetoxyquercetin.
The second technical means is to provide a method for preparing the rabdosia rubescens stem water extract powder. Cleaning Rabdosia rubescens stem with tap water, cutting, heating in distilled water at 50-100 deg.C for 0.5-4 hr, stirring at 200-400rpm, cooling to room temperature, filtering, washing filter cake with distilled water for 3 times, and concentrating the combined filtrate under reduced pressure to obtain Rabdosia rubescens stem water extract powder.
The third technical means is to provide a mass spectrum total ion current spectrum of the rabdosia rubescens stem water extract powder.
The fourth technical means is to provide the chemical structures of 28 components corresponding to the peaks in the mass spectrum total ion flow spectrum of the rabdosia rubescens stem aqueous extract powder.
The fifth technical means is to evaluate the free radical scavenging activity of the rabdosia rubescens stem aqueous extract powder.
Drawings
FIG. 1 UPLC-Mass Spectroscopy Total ion flux Profile of Rabdosia Rubescens (Hemsl.) Hara stem aqueous extract powder.
FIG. 2 shows 3-O- [ E-2,4, 6-tris (3-carboxy-2-acryloyloxy) -5- (3-hydroxy-2-acryloyloxy) cinnamoyl ] quinic acid and its cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 3 shows 3-O- [ E-2- (6-hydroxy-1, 3, 5-hexatrienylcarbonyloxy) -4- (4-carboxy-1, 3-butadienylcarbonyloxy) -5-methoxy-6- (3-hydroxy-1-propenylcarbonyloxy) cinnamoyl ] quinic acid and its cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 4 shows 3-O- [ E-2, 4-bis (4-carboxy-1-butenylcarbonyloxy) -3-hydroxy-5-methoxy-6- (6-carboxy-1, 3-hexadienylcarbonyloxy) cinnamoyl ] quinic acid in Rabdosia rubescens stem aqueous extract powder and its cleavage product.
FIG. 5 shows 3-O- [ E-2- (4-hydroxy-1, 3-butadienecarbonyloxy) -4- (3-hydroxyacryloyloxy) -5-hydroxy-6- (3-hydroxy-1-propenecarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 6 shows 3-O- [ E-2- (5-hydroxy-1-pentenylcarbonyloxy) -3-methoxy-4- (3-carboxy-1-propenylcarbonyloxy) -5- (3-hydroxypropionyloxy) -6- (5-carboxy-1-pentenylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 7 3-O- [ E-2, 5-dimethoxy-3-hydroxy-4- (4-carboxybutylcarbonyloxy) -6- (5-carboxypentyloxy) cinnamoyl ] quinic acid and cleavage products from an aqueous extract powder of Rabdosia rubescens stems.
FIG. 8 shows 3-O- [ E-2- (4-carboxybutoxycarbonyloxy) -3-hydroxy-4- (3-carboxypropylcarbonyloxy) -5-formyloxy-6- (4-carboxy-1-butenylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 9 shows 3-O- [ E-2-formyloxy-3, 4-bis (5-carboxy-1-pentenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (7-carboxy-1-heptenecarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 10 shows 3-O- [ E-2-carboxyacetoxy-3, 6-bis (5-carboxy-1-pentenoyloxy) -4- (4-carboxy-1-butenoyloxy) -5- (8-carboxy-1-heptenylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stems.
FIG. 11 shows 3-O- [ E-2, 4-bis (4-carboxy-1-butenylcarbonyloxy) -3-hydroxy-5- (5-hydroxypentanecarbonyloxy) -6- (5-hydroxy-1, 3-pentadienylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 12 shows 3-O- [ E-2- (4-carboxybutylcarbonyloxy) -3-hydroxy-4- (4-carboxy-1-butenylcarbonyloxy) -5, 6-bis (5-hydroxy-1-pentenylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 13 shows 3-O- [ E-2, 4-bis (4-carboxybutylcarbonyloxy) -3-hydroxy-5- (3-hydroxypropionyloxy) -6-carboxyacetoxycinnamoyl ] quinic acid and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 14 shows 3-O- [ E-2-formyloxy-3- (5-carboxy-1-pentenylcarbonyloxy) -4- (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (5-hydroxypentylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara.
FIG. 15 shows 3-O- [ E-2, 5-dihydroxymethoxy-4- (3-carboxypropionyloxy) -6- (5-carboxypentanoyloxy) cinnamoyl ] quinic acid and its cleavage products from an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stems.
FIG. 16 shows 3-O- [ E-2- (5-hydroxypentanecarbonyloxy) -3-hydroxy-4- (4-carboxybutanecarbonyloxy) -5, 6-dicarboxyacetylcinnamoyl ] quinic acid and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 17 shows 3-O- [ E-2- (4-carboxybutoxycarbonyloxy) -4, 6-bis (4-carboxy-1-butenylcarbonyloxy) -5- (3-carboxypropylcarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stems.
FIG. 18 shows 3-O- [ E-2- (3-carboxypropionyloxy) -3- (5-carboxy-1-pentenylcarbonyloxy) -4, 6-bis (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) cinnamoyl ] quinic acid and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 19 shows 3-O-glucoside-3 ' - (6-hydroxycarbonyloxy) -4 ' -methoxy-5 ' - (5-hydroxypentancarbonyloxy) quercetin and cleavage products in an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stem.
FIG. 20 3-O-glucoside-2 ' -hydroxy-3 ' -hydroxymethoxy-4 ' -hydroxyacetoxy-5 ', 6 ' -dicarboxyacetoxy quercetin and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 21 shows 3-O-glucoside-2 ', 6 ' -dihydroxymethoxy-3 ' - (3-hydroxypropionyloxy) -4 ' -hydroxy-5 ' - (3-carboxypropanoyloxy) quercetin and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 22 shows 3-O-glucoside-2 ', 6' -dimethoxy-3 '- (5-hydroxy-1, 3-pentadiene carbonyloxy) -4' - (3-hydroxy-1-propenylcarbonyloxy) quercetin and its cleavage products in an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stem.
FIG. 23 shows 3-O-glucoside-3 '- (4-carboxy-1-butenylcarbonyloxy) -4' - (4-carboxybutylcarbonyloxy) quercetin and cleavage products in an aqueous extract powder of Rabdosia rubescens stems.
FIG. 24 shows 3-O-glucoside-2 ' -hydroxy-4 ' - (3-carboxy acryloyloxy) -5 ' - (3-carboxy propylcarbonyloxy) quercetin and its cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 25 shows 3-O-glucoside-2 '-methoxy-4' - (3-carboxy-1-propenylcarbonyloxy) -5 '- (5-hydroxy-1-pentenylcarbonyloxy) -6' - (7-hydroxy-1, 3-heptadienylcarbonyloxy) quercetin and cleavage products in an aqueous extract powder of Rabdosia rubescens stems.
FIG. 26 shows 3-O-glucoside-2 '-methoxy-4', 5 '-dihydroxymethoxy-6' - (7-hydroxyheptacarbonyloxy) quercetin and its cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 27 shows 3-O-glucoside-2 ', 3 ' -dicarboxyloxy-4 ' - (4-carboxy-1-butenylcarbonyloxy) -5 ' - (3-carboxypropylcarbonyloxy) -6 ' - (4-hydroxy-1-butenylcarbonyloxy) quercetin and cleavage products in an aqueous extract powder of Rabdosia rubescens stem.
FIG. 28 3-O-glucoside-2 ' -hydroxy-3 ', 5 ' -dihydroxymethoxy-4 ' -carboxyacetoxy-6 ' - (3-carboxypropionyloxy) quercetin and cleavage products in Rabdosia rubescens stem aqueous extract powder.
FIG. 29 shows 3-O-glucoside-2 ', 3 ', 4 ' -trimethyloyloxy-5 ' - (4-hydroxy-1, 3-butadienecarbonyloxy) -6 ' -carboxyacetoxyquercetin and cleavage products in an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stems.
Detailed Description
To further illustrate the invention, a series of examples are given below. These examples are purely illustrative and are intended to be a detailed description of the invention only and should not be taken as limiting the invention.
Example 1 preparation of Rabdosia Rubescens Stem aqueous extract powder
Washing 300g of dry Rabdosia Rubescens (Hemsl.) Hara stem with tap water, and cutting into pieces. The cut stem was heated in 700-1200mL distilled water at 50-100 ℃ for 0.5-4 hours while stirring (200-400 rpm). Cooling to room temperature, filtering, and concentrating the filtrate under reduced pressure to obtain 72g Rabdosia Rubescens (Hemsl.) Hara stem water extract powder.
Example 2 determination of chromatography and Mass Spectrometry ion mobility Spectroscopy of Rabdosia Rubescens Stem aqueous extract powder
2-1 preparation of sample solution (10mg/mL)
26.7mg of Rabdosia Rubescens (Hemsl.) Hara stem water extract powder was weighed and dissolved in 2.67mL of ultrapure water. The resulting solution was sonicated for 1 minute, followed by centrifugation at 13000rpm for 10 minutes. The supernatant was taken, filtered through a 0.22 μm filter and placed in a sample vial for chromatographic and mass spectrometric determination.
2-2 chromatographic conditions
Chromatographic column Waters, Acquity
Figure BDA0002765505320000061
A HSS T3 column (2.1 × 100mm i.d.,1.7 μm); the sample injection volume is 2 mu L; 190-400nm of PDA detector; mobile phase water (0.1% formic acid), acetonitrile; the mobile phase was used and the column was washed with the gradient of table 1.
TABLE 1 mobile phase gradiometer
Figure BDA0002765505320000071
2-3 measuring chromatogram
UPLC chromatogram of the aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stem was determined and recorded according to the above chromatographic conditions (see FIG. 1 of the specification).
2-4. conditions for measuring ion flow spectrum and mass spectrum
Electrospray ionization modes are Positive (PI) and Negative (NI) modes. The ion mode parameters comprise that the capillary tube voltage is 1000V, the desolvation air flow rate is 800L/h, the temperature is 450 ℃, the source temperature is 120 ℃, the taper hole air flow rate is 50L/h, the spraying air pressure is 6bar, the fragmentation voltage is 20-45V, the sampling cone voltage is 6V, the acquisition mode is an MSE continuum resolution mode, the data acquisition range of the ratio (m/z) of the mass number of the charged particles to the charge number is 100-1500, the Trap fragmentation voltage of the low-energy channel is 6V, the gradient voltage of the Trap fragmentation voltage of the high-energy channel is 20-60V, LE (leucine enkephalin) is selected as a mass lock to acquire m/z, and the range is 100-1500.
2-5, recording ion flow spectrum and mass spectrum
Ion flow spectra of the rabdosia rubescens stem aqueous extract powder were determined and recorded according to the above conditions (see figure 1 of the specification).
Example 3 specifying the Structure of 28 Components in an aqueous extract powder of Rabdosia Rubescens Stem
The UPLC chromatogram of example 2 was coupled to a mass spectrum and the UPLC-mass spectrum of the rabdosia rubescens stem aqueous extract powder was determined. The mass spectrometry conditions are two modes of electrospray ionization, positive and negative ions. The ion mode parameters comprise that the capillary tube voltage is 1000V, the desolvation air flow rate is 800L/h, the temperature is 450 ℃, the source temperature is 120 ℃, the taper hole air flow rate is 50L/h, the spraying air pressure is 6bar, the fragmentation voltage is 20-45V, the sampling cone voltage is 6V, the acquisition mode is an MSE continuum resolution mode, the data acquisition range of the ratio (m/z) of the mass number of the charged particles to the charge number is 100-1500, the Trap fragmentation voltage of the low-energy channel is 6V, the gradient voltage of the Trap fragmentation voltage of the high-energy channel is 20-60V, LE (leucine enkephalin) is selected as a mass lock to acquire m/z, and the range is 100-1500. 28 independent peaks separated out within 30 minutes. The structural assignment of these peaks (in order of peaks from left to right in the total ion current spectrum) according to the mass spectrometry fragmentation law is given in table 2.
Table 2 shows the retention time of the peak-corresponding component, the mass number of the negative ion, the structure and the name of the negative ion in the total ion current spectrum
Figure BDA0002765505320000072
Figure BDA0002765505320000081
Figure BDA0002765505320000091
Figure BDA0002765505320000101
Figure BDA0002765505320000111
Figure BDA0002765505320000121
Figure BDA0002765505320000131
Example 4 evaluation of DPPH radical scavenging Activity of Rabdosia Rubescens Stem aqueous extract powder
DPPH is a stable nitrogenous radical that can be rapidly scavenged by protic radical scavengers. The activity of the proton radical scavenger DPPH in food and biological systems can be measured on a paramagnetic resonance apparatus. Antioxidant scavenging activity from natural sources can also be measured by DPPH on a paramagnetic resonance spectrometer. Concentration-dependent removal of DPPH from Rabdosia Rubescens (Hemsl.) Hara stem aqueous extract powder based on the third peak in paramagnetic resonance spectrumThe compound is shown in the specification. The results are shown in Table 3. The concentration of the aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara that scavenges 50% DPPH free radicals is 0.65mg/mL interpolated from the percentage DPPH scavenged shown in Table 3. That is, the Rabdosia Rubescens stem water extract powder removes the IC of DPPH free radical50It was 0.62 mg/mL. It can be seen that the aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stem is an excellent therapeutic agent for DPPH-induced oxidative stress.
TABLE 3 Rabdosia rubescens stem water extract powder for eliminating DPPH activity
Figure BDA0002765505320000132
Example 5 evaluation of hydroxyl radical scavenging Activity of Rabdosia Rubescens Stem aqueous extract powder
Fe2+/H2O2The hydroxyl radical formed by the system can be captured by DMPO and formed into DMPO-OH adduct for determination on paramagnetic resonance. The activity of the scavenger to scavenge hydroxyl radicals can be calculated from the height of the third peak of the paramagnetic resonance spectrum. The activity is expressed in relative amounts of DMPO-OH adduct. Therefore, the activity of the rabdosia rubescens stem water extract powder for removing hydroxyl free radicals is determined. The invention discovers that the concentration of the rabdosia rubescens stem water extract powder is increased, and the intensity of the third peak of the paramagnetic resonance spectrum is reduced. The results are shown in Table 4. The concentration of aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara that scavenges 50% of hydroxyl radicals as interpolated by the percentage of hydroxyl radical scavenges in Table 4 was 11.43 mg/mL. That is, the powder of Rabdosia rubescens stem water extract eliminates the IC of hydroxyl radical50It was 11.43 mg/mL. It can be seen that the aqueous extract powder of rabdosia rubescens stem is an excellent therapeutic agent for oxidative stress induced by hydroxyl radical.
TABLE 4 Rabdosia rubescens stem aqueous extract powder hydroxyl radical scavenging Activity
Figure BDA0002765505320000141

Claims (5)

1. An aqueous extract powder of rabdosia rubescens stems, wherein the aqueous extract powder of rabdosia rubescens stems contains 17 kinds of quinic acid compounds; wherein the 17 quinic acid compounds are 3-O- [ E-2,4, 6-tri (3-carboxyl-2-acryloyloxy) -5- (3-hydroxyl-2-acryloyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (6-hydroxyl-1, 3, 5-hexatrienylcarbonyloxy) -4- (4-carboxyl-1, 3-butadiene carbonyloxy) -5-methoxy-6- (3-hydroxyl-1-propylene carbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-di (4-carboxyl-1-butylene carbonyloxy) -3-hydroxyl-5-methoxy-6- (6-carboxyl-1, 3-hexadienecarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-hydroxy-1, 3-butadienylcarbonyloxy) -4- (3-hydroxyacryloyloxy) -5-hydroxy-6- (3-hydroxy-1-propenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (5-hydroxy-1-pentenylcarbonyloxy) -3-methoxy-4- (3-carboxy-1-propenylcarbonyloxy) -5- (3-hydroxypropionyloxy) -6- (5-carboxy-1-pentenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 5-dimethoxy-3-hydroxy-4- (4-carboxybutoxycarbonyloxy) -6- (5-carboxypentylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutoxycarbonyloxy) -3-hydroxy-4- (3-carboxypropylcarbonyloxy) -5-formyloxy-6- (4-carboxy-1-butenylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2-formyloxy-3, 4-bis (5-carboxy-1-pentenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (7-carboxy-1-heptenecarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2-carboxyacetoxy-3, 6-bis (5-carboxy-1-pentenoyloxy) -4- (4-carboxy-1-butenoyloxy) -5- (8-carboxy-1-heptenecarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-bis (4-carboxy-1-butenoyloxy) -3-hydroxy-5- (5-hydroxypentylcarbonyloxy) -6- (5-hydroxy-1, 3-pentadienoyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutylcarbonyloxy) -3-hydroxy-4- (4-carboxy-1-butenoyloxy) -5, 6-bis (5-hydroxy-1-pentenoyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 4-bis (4-carboxybutylcarbonyloxy) -3-hydroxy-5- (3-hydroxypropanoyloxy) -6-carboxyacetyloxy cinnamoyl ] quinic acid, 3-O- [ E-2-formyloxy-3- (5-carboxy-1-pentenoyloxy) -4- (4-carboxy-1-butenoyloxy) -5- (8-carboxy-1-octenecarbonyloxy) -6- (5-hydroxypentylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2, 5-dimethyloxy-4- (3-carboxypropyloxy) -6- (5-carboxycarbonyloxy) Pentylcarbonyloxy) cinnamoyl ] quinic acid, 3-O- [ E-2- (5-hydroxypentylcarbonyloxy) -3-hydroxy-4- (4-carboxybutoxycarbonyloxy) -5, 6-dicarboxyacarbonyloxy cinnamoyl ] quinic acid, 3-O- [ E-2- (4-carboxybutoxycarbonyloxy) -4, 6-bis (4-carboxy-1-butenecarbonyloxy) -5- (3-carboxypropylcarbonyloxy) cinnamoyl ] quinic acid and 3-O- [ E-2- (3-carboxypropyloxy) -3- (5-carboxy-1-pentenylcarbonyloxy) -4, 6-bis (4-carboxy-1-butenylcarbonyloxy) -5- (8-carboxy-1-octenecarbonyloxy) cinnamoyl ] quinic acid .
2. The rabdosia rubescens stem aqueous extract powder of claim 1, further comprising 11 quercetin-based compounds; wherein the 11 kinds of quercetin compounds are 3-O-glucoside-3 '- (6-hydroxyhexylcarbonyloxy) -4' -methoxy-5 '- (5-hydroxypentanoyloxy) quercetin, 3-O-glucoside-2' -hydroxy-3 '-hydroxymethoxy-4' -hydroxyacetoxy-5 ', 6' -dicarboxyacetyloxy quercetin, 3-O-glucoside-2 ', 6' -dihydroxymethoxy-3 '- (3-hydroxypropionyloxy) -4' -hydroxy-5 '- (3-carboxypropanoyloxy) quercetin, 3-O-glucoside-2', 6 '-dimethoxy-3' - (5-hydroxy-1, 3-pentadienecarbonyloxy) -4 '- (3-hydroxy-1-propenylcarbonyloxy) quercetin, 3-O-glucoside-3' - (4-carboxy-1-butenylcarbonyloxy) -4 '- (4-carboxybutylcarbonyloxy) quercetin, 3-O-glucoside-2' -hydroxy-4 '- (3-carboxypropenyloxy) -5' - (3-carboxypropylcarbonyloxy) quercetin, 3-O-glucoside-2 '-methoxy-4' - (3-carboxy-1-propenylcarbonyloxy) -5 '- (5-hydroxy-1-pentenylcarbonyloxy) -6' - (7-hydroxy-1, 3-heptadienylcarbonyloxy) quercetin, 3-O-glucoside-2 ' -methoxy-4 ', 5 ' -dihydroxymethoxy-6 ' - (7-hydroxyheptacarbonyloxy) quercetin, 3-O-glucoside-2 ', 3 ' -diformyloxy-4 ' - (4-carboxy-1-butenylcarbonyloxy) -5 ' - (3-carboxypropylcarbonyloxy) -6 ' - (4-hydroxy-1-butenylcarbonyloxy) quercetin, 3-O-glucoside-2 ' -hydroxy-3 ', 5 ' -dihydroxymethoxy-4 ' -carboxyacetoxy-6 ' - (3-carboxypropyloxy) quercetin and 3-O-glucoside-2 ', 3 ', 4' -Tricarbonyloxy-5 '- (4-hydroxy-1, 3-butadienecarbonyloxy) -6' -carboxyacetoxyquercetin.
3. A method of preparing an aqueous extract powder of Rabdosia Rubescens (Hemsl.) Hara stem of claim 1 or 2, comprising the steps of:
cleaning Rabdosia Rubescens (Hemsl.) Hara stem with tap water, cutting, soaking in distilled water under heating while stirring slowly, filtering, and concentrating the filtrate under reduced pressure to obtain powder which is Rabdosia Rubescens (Hemsl.) Hara stem water extract powder.
4. The method as claimed in claim 3, wherein the rabdosia stem is washed with tap water, cut up, heated preferably in distilled water at 50 ℃ -100 ℃ for 0.5-4 hours and stirred at 200-.
5. Use of the rabdosia rubescens stem aqueous extract powder of claim 1 or 2 for preparing a medicament against oxidative stress.
CN202011231955.XA 2020-11-06 2020-11-06 Rabdosia rubescens stem water extract powder, preparation thereof, free radical scavenging effect and application Withdrawn CN112353842A (en)

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