CN109223904B - Kidney tonifying and blood circulation promoting formula and preparation method thereof - Google Patents
Kidney tonifying and blood circulation promoting formula and preparation method thereof Download PDFInfo
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Abstract
The invention relates to the field of traditional Chinese medicines, and in particular relates to a resveratrol-containing traditional Chinese medicine preparation and a preparation method thereof. The common Chinese medicinal preparation is decocted by water, can not effectively extract effective components in medicinal materials, has direct relation between the content of the resveratrol in the compound preparation and the curative effect for a plurality of prescriptions with obvious curative effects for treating the nephropathy and delaying the aging, and has lower solubility of the resveratrol in water. Therefore, the invention provides a formula for tonifying kidney and activating blood circulation, which comprises the following traditional Chinese medicines in parts by weight: 5-15 parts of wolfberry fruit, 4-8 parts of giant knotweed, 5-15 parts of mulberry, 5-15 parts of angelica and 2-6 parts of pseudo-ginseng; and the preparation method comprises the following steps: pretreating the medicinal materials, extracting with alcohol, and making into Chinese medicinal extract. The kidney tonifying and blood activating prescription has the effects of tonifying spleen and kidney, activating blood circulation to dissipate turbid pathogen, resisting aging and the like; the process of ethanol extraction can extract resveratrol to the maximum extent, improve the effective components of the traditional Chinese medicine prescription and optimize the treatment effect.
Description
Technical Field
The invention relates to the field of traditional Chinese medicines, and in particular relates to a resveratrol-containing traditional Chinese medicine preparation and a preparation method thereof.
Background
Resveratrol (Resveratro, Res for short) is a non-flavonoid polyphenol compound with a chemical name of (E) 3, 5, 4-trihydroxy stilbene and a molecular formula of C14H1203The resveratrol has a relative molecular mass of 228.25g/mol, is a white needle crystal, exists in a plurality of plants, and is a phytoalexin generated when the plants are stressed by organisms or non-organisms (extraction and analysis detection technology research progress of resveratrol in giant knotweed rhizome in Song Hainan, Qianxingjun, Liu Wei [ [ J ] Henwei ]]Guangzhou chemical industry, 2014, 42 (21): 18-19.). The high-dose resveratrol has effects of resisting inflammation, scavenging free radicals, inhibiting bacteria and preventing cancer. In China, the Chinese medicinal material giant knotweed containing resveratrol has been used for treating and preventing hyperlipidemia and arteriosclerosis. Meanwhile, the resveratrol can play a role in protecting the cardiovascular from multiple aspects.
Experiments show that Res has good effect of delaying senescence, and can improve the oxidation resistance of organisms, improve the functions of testicular endocrine cells, promote the recovery of spermatogenic function and delay the decline of gonadal function. And can improve sperm quality (such as increasing sperm density, motility, and decreasing teratospermia rate) in senescent male mice, and improve the seminiferous epithelial structure and function of the major reproductive organs (testis) to improve reproductive function of the senescent body. In addition, a large amount of experimental evidences show that the resveratrol has the functions of resisting aging, tumors, inflammation and immunoregulation, and can improve the immunologic function of old mice and people.
The general Chinese medicine preparation is a water decoction preparation, i.e. Chinese medicine decoction pieces are put into boiling water for decoction, and a patient takes the water extraction product of the decoction pieces. However, for many prescriptions with obvious curative effect for treating nephropathy and delaying senility, the content of resveratrol in the compound prescription is directly related to the curative effect. However, resveratrol has low solubility in water, and thus decocting cannot sufficiently extract resveratrol from the traditional Chinese medicinal materials. Therefore, the traditional Chinese medicine theory is combined with a modern natural product research method, the preparation method for improving the resveratrol content in the prescription for treating the nephropathy and delaying the aging is developed, the preparation method has good production and application prospects, and has important significance for developing and inheriting traditional Chinese medicine culture.
Disclosure of Invention
The general Chinese medicine preparation is a water decoction, i.e. Chinese medicine decoction pieces are put into boiling water for decoction, and a patient takes the extract of the decoction pieces. However, for many prescriptions with obvious curative effect for treating nephropathy and delaying senility, the content of resveratrol in the compound prescription is directly related to the curative effect. However, the solubility of resveratrol in water is low, so that the common water decoction cannot sufficiently extract the resveratrol in the traditional Chinese medicinal materials. Therefore, the invention provides a formula for tonifying kidney and activating blood circulation, which comprises the following traditional Chinese medicines in parts by weight: 5-15 parts of wolfberry fruit, 4-8 parts of giant knotweed rhizome, 5-15 parts of mulberry, 5-15 parts of angelica and 2-6 parts of pseudo-ginseng.
The formula for tonifying kidney and activating blood circulation comprises the following traditional Chinese medicines in parts by weight: 10 parts of wolfberry fruit, 6 parts of giant knotweed rhizome, 10 parts of mulberry, 10 parts of Chinese angelica and 4 parts of pseudo-ginseng.
A formula for tonifying kidney and activating blood circulation is prepared by the following steps:
(1) weighing the wolfberry fruit, the giant knotweed rhizome, the mulberry fruit, the Chinese angelica and the pseudo-ginseng according to the weight parts, and pretreating the medicinal materials;
(2) adding 30-60% ethanol 4-10 times of the amount of the above pretreated medicinal materials, soaking for 1-21 days, filtering the medicinal liquid, centrifuging, and collecting supernatant;
(3) placing the supernatant obtained in the last step in an evaporating dish, rotary evaporating to dryness under the pressure of 30-50Pka at 30-70 deg.C and rotation speed of 60-90 r/min, taking out, and cooling in a drying dish for 10-60 min to obtain Chinese medicinal extract.
The pretreatment method comprises the steps of preparing decoction pieces or crushing.
The invention has the beneficial effects that: the clinical curative effect of the traditional Chinese medicine preparation disclosed by the invention comprises the following steps: invigorating spleen and kidney, promoting blood circulation, eliminating turbid pathogen, resisting aging, and treating nephropathy, premature ovarian failure caused by Tripterygium wilfordii preparation, and lumbago. The traditional Chinese medicine preparation prepared by the preparation method disclosed by the invention has the advantages that the content of resveratrol is obviously improved, the solubility of resveratrol in water is low, the resveratrol in the resveratrol can be extracted to the maximum extent by using an ethanol extraction process, the effective components of the traditional Chinese medicine prescription are improved, and the treatment effect is optimized. The preparation method is simple, and the finally prepared extract is convenient for patients to take.
Drawings
FIG. 1 HPLC profile of control solution.
FIG. 2 HPLC chromatogram of the sample solution.
FIG. 3 HPLC chromatogram of negative sample.
Detailed Description
The formula for tonifying kidney and activating blood circulation comprises the following traditional Chinese medicines in parts by weight: 5-15 parts of wolfberry fruit, 4-8 parts of giant knotweed rhizome, 5-15 parts of mulberry, 5-15 parts of angelica and 2-6 parts of pseudo-ginseng.
The preferred embodiment is as follows: 10 parts of wolfberry fruit, 6 parts of giant knotweed rhizome, 10 parts of mulberry, 10 parts of Chinese angelica and 4 parts of pseudo-ginseng.
The preparation method comprises the following steps:
(1) weighing the wolfberry fruit, the giant knotweed rhizome, the mulberry fruit, the Chinese angelica and the pseudo-ginseng according to the weight parts, and pretreating the medicinal materials.
(2) Adding 30-60% ethanol 4-10 times of the amount of the above pretreated medicinal materials, soaking for 1-21 days, filtering the medicinal liquid, centrifuging, and collecting supernatant.
(3) Placing the supernatant obtained in the last step in an evaporation dish, and performing rotary evaporation to dryness under the pressure of 30-50Pka at the temperature of 30-70 ℃ and the rotation speed of 60-90 r/min; taking out, and cooling in a drying container for 10-60 min to obtain Chinese medicinal extract.
The pretreatment method comprises the steps of preparing decoction pieces or crushing.
Analytical conditions and methods:
HPLC conditions: a pre-column (phenomenex-KJ 0-4282); a chromatographic column: phenomenex-C18 (250X 4.6 mm, 5 μm); mobile phase: acetonitrile (A) -0.1% aqueous phosphoric acid (B) (28: 72), elution conditions: 0.01-8.50 min, 28% A; 8.51-14.00 min, 28% -40% A; 14.01-20.00 min, 90% A; 20.10-28.00 min, 28% A; flow rate: 0.01-5.00 min, 1.0 ml/mim; 5.01-5.50 min, 1.0-0.8 ml/min; 5.50-9.00 min, 0.8 ml/min; 9.01-9.50 min, 0.8-1.0 ml/min; 9.51-28.00 min, 1.0 ml/min; column temperature: 30 ℃; detection wavelength: 310 nm; sample introduction amount: 5 mul.
2. Preparing a standard curve of resveratrol in the step of pretreating medicinal materials: precisely sucking 1 mul, 2 mul, 3 mul, 5 mul and 10 mul of resveratrol reference substance solution (61.628 mug/mL), dissolving in methanol solution, and fixing the volume to 10 mL: detecting according to the HPLC detection method, recording the integral value of the chromatographic peak area, and taking the resveratrol sample amount (mug) as the abscissa(s) ((g))X) Taking the integrated value of the measured peak area as the ordinate (c:)Y) And drawing a standard curve. The regression equation isY = (7.81425×106)X，（r=0.9999, detection range: 0.061628-0.61628 mug/ml).
3. Preparing a standard curve of resveratrol in the extract: accurately weighing appropriate amount of resveratrol control, dissolving with methanol to obtain control stock solution with mass concentration of 0.234 mg/ml, diluting at equal ratio to obtain series of control solutions (resveratrol concentrations of 0.234, 0.117, 0.0585, 0.02925, 0.01463, 0.0073125, and 0.0036563 mg/ml). Precisely measuring 5 μ l of each of the above series of reference solutions, detecting by HPLC to obtain peak area of each sample, and drawing standard curve. The regression equation isY=3.92×107 X55939.2 (r =0.9999, detection range is 36.56-234.00 mug/ml).
4. Preparation of control (a) solution: resveratrol control, concentration 0.234 mg/ml.
5. Preparation of sample solution (B):
taking 10g of medlar, 10g of mulberry, 6g of giant knotweed, 10g of angelica and 4g of pseudo-ginseng, grinding into powder, adding 320mL of 55% ethanol solution, soaking for 14 days, taking supernatant of the ethanol extract, centrifuging at 45000 r/min for 10 min, taking supernatant, fixing the volume to 250 mL, precisely absorbing 10mL, performing rotary evaporation to dryness (pressure: 40 Pka; temperature: 50 ℃, rotation speed: 75 r/min), adding acetonitrile-0.1% phosphoric acid (28: 72) solution into residues, fixing the volume to 10mL, filtering through a 0.45 mu m microfiltration membrane, and taking subsequent filtrate to obtain the Chinese medicinal composition.
6. Negative sample (C) solution preparation:
taking 10g of medlar, 10g of mulberry, 10g of angelica and 4g of pseudo-ginseng, grinding into powder, and soaking in 55% ethanol for 14 days. Taking the ethanol extract, centrifuging at 45000 r/min for 10 min, taking supernatant, fixing the volume to 250 mL, precisely absorbing 10mL, performing rotary evaporation to dryness (pressure: 40 Pka; temperature: 50 ℃, rotation speed: 75 r/min), adding acetonitrile-0.1% phosphoric acid (28: 72) solution into residues to fix the volume to 10mL, filtering through a 0.45 mu m microporous membrane, and taking subsequent filtrate to obtain the product.
7. The method for calculating the dry paste rate comprises the following steps:
precisely sucking 25.0 mL of sample solution, placing in an evaporation dish with constant weight, evaporating in a water bath to dryness, drying at 105 ℃ for 3 h, taking out, placing in a dryer for cooling for 30 min, precisely weighing the mass, and obtaining the dry extract rate according to a formula (the extract rate is dry extract mass multiplied by 10/medicinal material mass multiplied by 100%).
Experimental part:
1. influence of the medicinal material pretreatment method on resveratrol content.
The first pretreatment method comprises the following steps: taking 2 parts of decoction pieces in the prescription amount, respectively adding yellow wine in an amount which is 5 times of the prescription amount, and soaking for 1 month. Filtering the medicinal liquid, centrifuging, collecting supernatant, diluting with yellow wine to a constant volume of 200ml, and shaking to obtain decoction pieces-1 and decoction pieces-2.
The second pretreatment method comprises taking 2 parts of medicinal powder according to the prescription amount, wrapping the medicinal powder with gauze, adding 5 times of yellow wine according to the prescription amount, and soaking for 1 month. Filtering the medicinal liquid, centrifuging, collecting supernatant, diluting with yellow wine to a constant volume of 200ml, and shaking to obtain powder-1 and powder-2.
Table 1 total resveratrol content for different pretreatment methods.
The above results show that the resveratrol content obtained by pulverizing the medicinal materials is more desirable, and therefore, in the following examples, the medicinal materials were subjected to pulverization pretreatment.
2. Influence of different preparation processes on resveratrol content.
The control solution (A), the sample solution (B) and the negative sample (C) were detected by the HPLC detection method described above, and the HPLC profiles obtained are shown in FIGS. 1 to 1:
resveratrol is mainly present in the traditional Chinese medicine giant knotweed rhizome, so that the peak 1(t =12.5min) in fig. 2 can be determined as the resveratrol peak by combining the HPLC (high performance liquid chromatography) spectrums of a resveratrol control product and a negative control product solution (lacking the traditional Chinese medicine giant knotweed rhizome).
The factors affecting the extraction amount of resveratrol were used as variables, wherein ethanol concentration/% (a), soaking days/day (B), particle size (C), feed-to-liquid ratio (D), as shown in the following table:
table 1 variable factors affecting resveratrol content.
According to L9( 34 ) Orthogonal test, detecting the extract obtained under the orthogonal test condition of various variable combinations by the HPLC analysis method to obtain the content of resveratrol, and taking the resveratrol and dry extract rate as evaluation indexes. The results are shown in the following table:
TABLE 2L9(34) Orthogonal experimental design and results.
As can be seen from table 2, the extract has the highest resveratrol content and the preferable dry extract rate under the conditions of a1 (ethanol concentration 55%), B3 (soaking days 21 days), C3 (medium meal) and D3 (feed-liquid ratio 8).
The results of the cross-over test were analyzed using SPSS 19.0 and the results of the anova are shown in table 3. According to visual analysis, the main action of all factors on the alcohol extraction process of the traditional Chinese medicine preparation disclosed by the invention is that the ethanol concentration is more than D, the material-liquid ratio is more than C, the granularity is more than B, and the soaking days are more than B; analysis of variance shows that the ethanol concentration (A) and the feed-liquid ratio (D) have a significant influence on the test result (P is less than 0.05). The data fitting effect of the B soaking days and the C granularity is not obvious.
Table 3 analysis of variance of orthogonal factors.
As can be seen from table 3, the number of days of soaking and the particle size were not significant for the content of resveratrol; in addition, the influence of the soaking days B on the result is small, and the time cost difference between 14 days of soaking and 21 days of soaking is large in combination with actual production, so that the optimal condition is selected to be 14 days of soaking.
Wolfberry fruit: has effects in replenishing vital essence, improving eyesight, and tranquilizing mind.
Giant knotweed rhizome: has effects of promoting diuresis, eliminating jaundice, clearing heat, removing toxic substance, removing blood stasis, relieving pain, relieving cough, and eliminating phlegm.
And (3) mulberry fruit: is sweet and cold in nature and has the effects of tonifying liver and kidney, promoting the production of body fluid and moistening dryness, blackening hair and improving eyesight and the like.
Chinese angelica: has effects in nourishing blood, regulating menstruation, relieving pain, moistening dryness, smoothing intestine, resisting cancer, resisting aging, and enhancing immunity.
Pseudo-ginseng: disperse blood stasis, stop bleeding, resolve swelling and alleviate pain.
The clinical implementation case is as follows:
case one:
the traditional Chinese medicine preparation disclosed by the invention can be used for preventing and treating premature ovarian failure caused by the tripterygium wilfordii preparation.
In the study, 40 female patients with chronic nephritis were selected, and 20 patients of the treatment group and the control group were randomly divided. The treatment group is treated by the tripterygium glycosides tablet 20mg three times a day and the traditional Chinese medicine preparation disclosed by the invention, the control group is treated by the tripterygium glycosides tablet 20mg three times a day, after 3 months, two groups of curative effects are evaluated according to a clinical symptom score table, biochemical indexes and menstruation conditions of a patient, if the patient has a shortened menstruation period, a reduced menstruation amount and even amenorrhea after 3 months, the tripterygium glycosides tablet is given to a patient for stopping taking, and the menstruation recovery condition of the patient is tracked by regular follow-up visits. The results show that the preparation can effectively relieve the urine protein of patients with chronic nephritis, improve the renal function, relieve clinical symptoms and delay the progress of nephropathy. In particular to the antagonism of the hypofunction of female ovary and the ovulation obstacle caused by the tripterygium wilfordii preparation. After the tripterygium wilfordii preparation is stopped, comparing the treatment group with the control group, finding that the menstruation recovery time of the patient taking the traditional Chinese medicine preparation disclosed by the invention is faster than that of other patients, and meanwhile, in the application process, the proportion of menstrual disorder is reduced, and the proportion of abnormal serum hormone level is lower than that of the control group.
Tripterygium glycosides tablets are widely used for treating chronic kidney diseases and various immune diseases, have fewer side effects than common hormones and immunosuppressants, particularly can avoid side effects such as eyelid edema, central obesity, blood sugar rise and the like caused by using hormones, and have good effects on various kidney diseases such as diabetic nephropathy, asymptomatic proteinuria, IgA nephropathy and the like. The deficiency in the middle-jiao is that about 30% of patients can have reproductive hypofunction after taking tripterygium glycosides tablet, which is manifested by scanty menstruation, amenorrhea, hyposexuality, dysfunctional uterine bleeding and sex hormone level reduction, and type-B ultrasonic manifested by premature ovarian failure such as ovary or uterus atrophy, female uterus subnormal, etc. Although some patients may experience slow improvement of reproductive dysfunction after discontinuation of the tripterygium wilfordii formulation, some patients may experience irreversible ovarian damage. In clinical application aiming at the problems, the traditional Chinese medicine for tonifying the kidney and invigorating the blood circulation is found to have the function of obviously improving the conditions. We observe that after taking tripterygium glycosides tablets, the time for the patient to reach the normal level of the hypomenorrhea, amenorrhea and sex hormone level is obviously shortened compared with the control group, and the premature ovarian failure is obviously relieved. The Chinese medicinal preparation disclosed by the invention has the effects of tonifying kidney, regulating menstruation, strengthening spleen and discharging turbidity.
Case two:
clinical observation of patients with kidney deficiency:
clinical observation shows that 60 patients with kidney deficiency in traditional Chinese medicine diagnosis are treated by 30 patients with the traditional Chinese medicine preparation disclosed by the invention, and 30 patients with the control group are treated by pills of six ingredients with rehmannia. The treatment course is 6 weeks, and the patient symptom score is used as an evaluation criterion.
Note: the kidney deficiency syndrome differentiation standard refers to the national conference of traditional Chinese and western medicine combined deficiency syndrome research and prevention and treatment of senile diseases in 1982: soreness of the waist and back, soreness of the waist and knees or pain of the heel, tinnitus, deafness, loss of teeth, dribbling or incontinence of urine, impotence, premature ejaculation, and menoxenia. Three items are available to diagnose kidney deficiency.
The research finds that the traditional Chinese medicine preparation disclosed by the invention has the advantages of obviously improving the symptoms of patients with kidney deficiency and having good anti-aging effect compared with the pill of six ingredients with rehmannia, and the formula has the effects of tonifying kidney, securing essence, regulating kidney essence, balancing kidney yin and kidney yang, storing vital energy of a human body and strengthening five internal organs. It has effects of scavenging free radicals, regulating gene signal, enhancing immunity, repairing neuroendocrine disorder, delaying aging, relieving aging symptom, and prolonging life of elderly patients.
Case three:
for treating lumbago:
for patients with lumbago caused by kidney deficiency accompanied by cold-dampness, qi stagnation and blood stasis, the lumbago can be relieved obviously by differentiation of symptoms and signs. The medicated wine is one of various dosage forms of traditional Chinese medicine, and the Huangdi's classic has the treatise on herb liquor with amazalin. "Jiehonglin" is also known as medicated wine. The wine is the long of hundreds of drugs, and the wine can easily enter the histiocyte of the traditional Chinese medicine to dissolve various organic substances out so as to play the original therapeutic effect of the crude drugs. The medicinal liquor is used for treating the lumbago caused by the cold, can be orally taken and externally applied, and is convenient and effective.
The resveratrol in the traditional Chinese medicine preparation disclosed by the invention has an estrogen-like effect, can regulate and control ovarian cell apoptosis, repair ovarian insufficiency, improve the ovarian maturation rate, relieve diabetic renal fibrosis, relieve renal tubular epithelial cell apoptosis induced by high sugar and improve renal function. Meanwhile, the medicine can repair the oxidative damage of endothelial cells and inhibit the oxidative stress reaction of kidney tissues. It also has effects in improving memory, regulating mitochondrial pathway, and protecting cranial nerve cells. Has reliable curative effect in treating chronic kidney diseases and anti-aging treatment.
Claims (2)
1. The preparation method of the formula for tonifying kidney and activating blood is characterized in that the formula for tonifying kidney and activating blood comprises the following traditional Chinese medicines in parts by weight: 10 parts of wolfberry fruit, 6 parts of giant knotweed, 10 parts of mulberry, 10 parts of Chinese angelica and 4 parts of pseudo-ginseng; the preparation method comprises the following steps:
(1) weighing the wolfberry fruit, the giant knotweed rhizome, the mulberry fruit, the Chinese angelica and the pseudo-ginseng according to the weight parts, and pretreating the medicinal materials;
(2) adding 30-60% ethanol 4-10 times of the amount of the above pretreated medicinal materials, soaking for 1-21 days, filtering the medicinal liquid, centrifuging, and collecting supernatant;
(3) placing the supernatant obtained in the last step in an evaporation dish, and performing rotary evaporation to dryness under the pressure of 30-50kPa, the temperature of 30-70 ℃ and the rotating speed of 60-90 r/min; taking out, and cooling in a drying container for 10-60 min to obtain Chinese medicinal extract.
2. The preparation method of the kidney tonifying and blood circulation promoting formula as claimed in claim 1, wherein the pretreatment method comprises decoction piece preparation or pulverization.
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| CN112098563B (en) * | 2020-10-14 | 2022-07-15 | 中国中医科学院望京医院(中国中医科学院骨伤科研究所) | Detection method for chemical components of kidney-tonifying and blood-activating formula |
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