CN108498498B - Application of salvianolic acid component in preparation of pharmaceutical preparation for improving protein-bound uremic toxin dialysis rate and filtration rate - Google Patents
Application of salvianolic acid component in preparation of pharmaceutical preparation for improving protein-bound uremic toxin dialysis rate and filtration rate Download PDFInfo
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Abstract
本发明公开了丹酚酸类成分在制备提高蛋白结合型尿毒素透析率及滤过率药物制剂中的应用。丹酚酸类成分包括丹酚酸B、原儿茶醛、迷迭香酸等丹酚酸类单体成分，以及含丹参的药物制剂、含丹参和红花的药物制剂、不同配伍比例的丹参和红花药对等。本发明通过大量实验筛选，结果表明丹红注射液以及丹酚酸类等成分均能显著提高蛋白结合型尿毒素的透析效率，显著提高蛋白结合型尿毒素的滤过效率，可开发为肾病透析和血液滤过等体外肾脏替代治疗策略的辅助药物制剂，具有很好的开发应用前景。
The invention discloses the application of salvianolic acid components in preparing a pharmaceutical preparation for improving the dialysis rate and filtration rate of protein-bound urea toxin. Salvianolic acids include salvianolic acid B, protocatechuic aldehyde, rosmarinic acid and other salvianolic acid monomers, as well as medicinal preparations containing salvia miltiorrhiza, medicinal preparations containing salvia miltiorrhiza and safflower, and salvia miltiorrhiza in different proportions Equivalent to safflower. The present invention has screened through a large number of experiments, and the results show that Danhong injection, salvianolic acids and other components can significantly improve the dialysis efficiency of protein-bound urea toxins, significantly improve the filtration efficiency of protein-bound urea toxins, and can be developed into dialysis for kidney diseases. It is an adjuvant drug preparation for extracorporeal renal replacement therapy strategies such as hemofiltration and has good development and application prospects.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to application of salvianolic acid components in preparation of a preparation for improving the dialysis rate and filtration rate of protein-bound uremic toxin.
Background
Renal function of end-stage renal disease patients presents serious disorders, and metabolic wastes in the body cannot be effectively removed, so that more than 90 kinds of uremic toxins are accumulated in the bodies of the patients. Accumulated uremic toxins can induce complications such as bone disease, encephalopathy, cardiovascular disease, anemia, soft tissue calcification, and immune response abnormality. Reducing the level of uremic toxins in a patient can significantly improve the therapeutic efficacy of patients with renal disease. The uremic toxins mainly comprise small-molecule uremic toxins (molecular weight <500Da), protein-bound small-molecule uremic toxins (molecular weight <500Da) and medium-molecule uremic toxins (molecular weight >500 Da). The existing extracorporeal kidney replacement treatment strategies commonly used in clinic, such as dialysis (including hemodialysis and peritoneal dialysis) and hemofiltration, can effectively eliminate small-molecule uremic toxins and also can eliminate medium-molecule uremic toxins, but cannot eliminate protein-bound small-molecule uremic toxins. Because either the artificial synthetic semipermeable membrane used in hemodialysis and hemofiltration or the patient's own peritoneum used in peritoneal dialysis can only allow free uremic toxins to pass through in blood circulation, the free uremic toxins are present in a small proportion, and the dialysis efficiency and filtration efficiency are low.
Reducing the level of protein-bound uremic toxins can significantly improve clinical dialysis and filtration effects, and eliminating protein-bound uremic toxins is the biggest challenge in current dialysis and filtration technologies.
Disclosure of Invention
The purpose of the invention is as follows: the invention aims to overcome the defects of the prior art, and provides a salvianolic acid component capable of improving the dialysis efficiency and the filtration efficiency of protein-bound uremic toxin and a traditional Chinese medicine or a traditional Chinese medicine compound containing the salvianolic acid component through a large amount of experimental screening.
The technical scheme is as follows: in order to solve the technical problems, the technical scheme adopted by the invention is as follows:
application of salvianolic acid in preparing preparation for improving dialysis rate and filtration rate of protein-bound uremic toxin is provided.
Preferably, the salvianolic acid component is applied to preparation of preparations for improving the dialysis rate and filtration rate of protein-bound uremic toxin, and the salvianolic acid component comprises a medicinal preparation containing salvia miltiorrhiza, a medicinal preparation containing salvia miltiorrhiza and safflower, and a pair of salvia miltiorrhiza and safflower in different compatibility ratios.
Preferably, the salvianolic acid component is applied to preparation of preparations for improving the dialysis rate and filtration rate of protein-bound uremic toxin, and the salvianolic acid component comprises a danhong injection and a danshen dripping pill.
Preferably, the salvianolic acid components include lithospermic acid, salvianolic acid A, danshensu, caffeic acid, salvianolic acid B, salvianolic acid C, salvianolic acid D, salvianolic acid E, protocatechuic acid, protocatechuic aldehyde, rosmarinic acid and other salvianolic acid components. Particularly preferred are salvianolic acid B, protocatechualdehyde, and rosmarinic acid.
Preferably, the salvianolic acid component is used for preparing a preparation for improving the dialysis rate and filtration rate of protein-bound uremic toxin, wherein the protein-bound uremic toxin comprises protein-bound small-molecule uremic toxins such as indoxyl sulfate, p-cresol sulfate, 3-carboxy-4-methyl-5-propyl-2-furanpropionic acid, 2-methoxyresorcinol, 3-deoxyglucosone, glyoxal, methylglyoxal, p-formic acid, phenol, lysine, fructolysine, hippuric acid, p-hydroxyhippuric acid, kynurenine, kynurenic acid, homocysteine, pentostatin, putrescine, spermine, spermidine and the like. Indoxyl sulfate and p-cresol sulfate are particularly preferred.
Application of salvianolic acid in preparing preparation for improving protein-bound uremic toxin dialysis rate and filtration rate in hemodialysis, peritoneal dialysis, and hemofiltration treatment processes is provided.
Has the advantages that: compared with the prior art, the invention has the following advantages:
through a large number of experimental screens, experimental results show that components such as danhong injection or salvianolic acid and the like can obviously improve the dialysis efficiency and the filtration efficiency of protein-bound uremic toxin, and can obviously improve the clinical commonly used dialysis methods such as hemodialysis, peritoneal dialysis and the like and the extracorporeal kidney replacement treatment effects such as hemofiltration and the like. Can be developed into an auxiliary medicinal preparation for kidney disease dialysis.
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FIG. 1 is a bar graph of the efficiency of DANHONG injection in improving dialysis efficiency of protein-bound indoxyl sulfate and p-cresol sulfate.
FIG. 2 is a bar graph of the effect of salvianolic acid on increasing dialysis efficiency of protein-bound indoxyl sulfate and p-cresol sulfate.
FIG. 3 is a bar graph of Danhong injection for improving the filtration efficiency of protein-bound indoxyl sulfate and p-cresol sulfate.
FIG. 4 is a bar graph of salvianolic acid component increasing the filtration efficiency of protein-bound indoxyl sulfate and p-cresol sulfate.
Detailed Description
The foregoing aspects of the present invention are described in further detail below by way of examples, but it should not be construed that the scope of the above-described subject matter of the present invention is limited to the following examples, and that all the technologies implemented based on the above-described aspects of the present invention are within the scope of the present invention.
Example 1 Danhong injection significantly improves dialysis efficiency of protein-bound uremic toxins
1. Experimental methods
1.1 sample preparation method
Preparation of standard sample: indoxyl Sulfate (IS) and p-cresol sulfate (PCS) standard substances are precisely weighed and dissolved in 200 mu l of ultrapure water to prepare standard substance solutions with the concentrations of IS and PCS being 50 mu g/ml.
Preparation of control sample: taking 165 ul of rat plasma sample, adding 10 ul of IS aqueous solution and 5 ul of PCS aqueous solution, whirling for 30s, fully mixing, standing for 30min at room temperature, adding 20 ul of ultrapure water, whirling for 30s, and fully mixing. The total volume of the dialysate was 200. mu.l, with IS and PCS at a concentration of 50. mu.g/ml.
Preparing a danhong injection sample: taking 165 mu l of rat plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, vortexing for 30s, fully mixing, standing for 30min at room temperature, adding 20 mu l of each of the Danhong injection (Danhong injection stock solution, Danhong injection diluted 2 times, Danhong injection diluted 4 times) with different concentrations, vortexing for 30s, and fully mixing. The Danhong injection can be prepared from 750g of Salvia miltiorrhiza and 250g of safflower produced by Jinan steplength pharmacy Co.
1.2 in vitro microdialysis method
The microdialysis system (equipped with a dialysis membrane with a molecular cut-off of 15,000 Da; CMA, Stockholm, Sweden) was flushed through the line with PBS buffer for 15min, and after the control samples had equilibrated for 100min, the dialysate was collected every 20min for a total of 4 times. The method for collecting the dialysate of the Danhong injection sample is the same as the control sample. The collected dialysate was quantitatively analyzed for IS and PCS content using HPLC-PDA method.
1.3 data analysis method
Differences between groups were analyzed using a Nonparametric Mann-Whitney Test with P <0.05 for significant differences and P <0.01 for very significant differences. IS/PCS dialysis efficiency (%) -. Instrument response of IS/PCS in dialysate/Instrument response of IS/PCS in standard sample × 100%.
The IS/PCS dialysis efficiency improvement ratio (%) - (dialysis efficiency of danhong injection sample-dialysis efficiency of control sample)/dialysis efficiency of control sample × 100%.
2. Results of the experiment
The Danhong injection can obviously improve the dialysis efficiency of the IS, and the higher the concentration of the Danhong injection, the stronger the promotion effect on the IS dialysis (as A in figure 1, P IS less than 0.01). The low-concentration Danhong injection (namely the Danhong injection IS diluted by 40 times) can improve the dialysis efficiency of the IS by 77.52%, and the medium-concentration Danhong injection (namely the Danhong injection IS diluted by 20 times) and the high-concentration Danhong injection (namely the Danhong injection IS diluted by 10 times) can respectively improve the dialysis efficiency of the IS by 93.79% and 99.13%. A very good technical effect is achieved.
The danhong injection showed the same significant concentration-dependent promoting effect on PCS dialysis (B in fig. 1, P < 0.01). The low-concentration Danhong injection (namely the Danhong injection is diluted by 40 times) can improve the dialysis efficiency of PCS by 78.30 percent, and the medium-concentration (namely the Danhong injection is diluted by 20 times) and the high-concentration Danhong injection (namely the Danhong injection is diluted by 10 times) can respectively improve the dialysis efficiency of PCS by 86.33 percent and 142.00 percent. A very good technical effect is achieved.
Example 2 Salvianolic acid component significantly improves dialysis efficiency of protein-bound uremic toxins
1. Experimental methods
1.1 sample preparation method
Preparation of control sample: taking 180 mu l of rat plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, vortexing for 30s, fully mixing, standing for 30min at room temperature, and adding 5 mu l of ultrapure water. The total volume of the dialysate was 200. mu.l, with IS and PCS at a concentration of 50. mu.g/ml.
Preparing a salvianolic acid sample: taking 180 mu l of rat plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, swirling for 30s, mixing well, standing for 30min at room temperature, adding 5 mu l each of high-concentration (50 mu g/ml), medium-concentration (10 mu g/ml) and low-concentration (5 mu g/ml) alkannic acid, salvianolic acid A, tanshinol, caffeic acid, salvianolic acid B, salvianolic acid C, salvianolic acid D, salvianolic acid E, protocatechuic acid, protocatechuic aldehyde and rosmarinic acid aqueous solutions (caffeic acid, protocatechuic aldehyde and rosmarinic acid are insoluble in pure water and dissolved by 20% acetonitrile aqueous solution), swirling for 30s, mixing well. The total volume of the dialysate was 200. mu.l.
1.2 in vitro microdialysis method as in example 1 item 1.2; the data analysis method was the same as that of item 1.3 in example 1.
2. Results of the experiment
Phenolic acid component: lithospermic acid, salvianolic acid A, tanshinol, caffeic acid, salvianolic acid B, salvianolic acid C, salvianolic acid D, salvianolic acid E, protocatechuic acid, protocatechuic aldehyde and rosmarinic acid can obviously improve the dialysis efficiency of IS and PCS, wherein the salvianolic acid B and protocatechuic aldehyde have the strongest promotion effect on IS dialysis, and the high-concentration salvianolic acid B and protocatechuic aldehyde (50 mu g/ml) can respectively improve the dialysis efficiency of IS by 70.90% and 66.50% (A in figure 2, P IS less than 0.01). The most strong promoting effect on PCS dialysis is protocatechualdehyde and rosmarinic acid, and the high concentration of protocatechualdehyde and rosmarinic acid (50 mu g/ml) can respectively improve the dialysis efficiency of PCS by 77.88% and 62.87% (figure 2B, P < 0.01). A very good technical effect is achieved.
Example 3 Danhong injection significantly improves the filtration efficiency of protein-bound uremic toxins
1. Experimental methods
1.1 sample preparation method
Preparation of standard sample: IS and PCS standard substances are precisely weighed and dissolved by 200 mul of ultrapure water to prepare standard substance solutions with the IS and PCS concentration of 50 mug/ml.
Preparation of control sample: taking 165 mu l of human mixed plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, whirling for 30s, fully mixing, standing for 30min at room temperature, adding 20 mu l of ultrapure water, whirling for 30s, and fully mixing. The total volume of the dialysate was 200. mu.l, with IS and PCS at a concentration of 50. mu.g/ml.
Preparing a danhong injection sample: taking 165 mu l of human mixed plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, vortexing for 30s, fully mixing, standing for 30min at room temperature, adding 20 mu l of each of the Danhong injection solutions with different concentrations (Danhong injection stock solution, Danhong injection solution diluted 2 times, Danhong injection solution diluted 4 times), vortexing for 30s, and fully mixing.
1.2 extracorporeal hemofiltration method
Hemofiltration Devices (Amicon Ultra-0.5Centrifugal Filter Devices; Millipore, Darmstadt, Germany) were soaked with Milli-Q water for 30min and centrifuged (1,4000 Xg, 10s) to spin dry the water. 200 μ l of a control sample or a Danhong injection sample (n ═ 3) was added, and after centrifugation (1,4000 Xg, 10min), the filtrate was collected and quantitatively analyzed for IS and PCS content by HPLC-PDA method.
1.3 data analysis method
Differences between groups were analyzed using a Nonparametric Mann-Whitney Test with P <0.05 for significant differences and P <0.01 for very significant differences. IS/PCS filtration efficiency (%) -instrument response to IS/PCS in filtrate/instrument response to IS/PCS in standard sample x 100%. The IS/PCS filtration efficiency improvement ratio (%) - (filtration efficiency of Danhong injection sample-filtration efficiency of control sample)/filtration efficiency of control sample × 100%.
2. Results of the experiment
The Danhong injection can obviously improve the filtration efficiency of the IS, and the higher the concentration of the Danhong injection, the stronger the promotion effect on the filtration of the IS (A in figure 3, P IS less than 0.01). The low-concentration Danhong injection (namely the Danhong injection IS diluted by 40 times) can improve the filtration efficiency of the IS by 80.92 percent, and the medium-concentration (namely the Danhong injection IS diluted by 20 times) and the high-concentration Danhong injection (namely the Danhong injection IS diluted by 10 times) can respectively improve the filtration efficiency of the IS by 92.10 percent and 124.92 percent.
The danhong injection showed the same significant concentration-dependent promotion effect on PCS filtration (B in fig. 3). The low-concentration Danhong injection (namely the Danhong injection IS diluted by 40 times) can improve the filtration efficiency of PCS by 25.05 percent, and the medium-concentration (namely the Danhong injection IS diluted by 20 times) and the high-concentration Danhong injection (namely the Danhong injection IS diluted by 10 times) can respectively improve the dialysis efficiency of IS by 28.23 percent and 104.14 percent (P < 0.01).
Example 4 Salvianolic acid component significantly improves the filtration efficiency of protein-bound uremic toxins
1. Experimental methods
1.1 sample preparation method
Preparation of control sample: taking 180 mu l of human mixed plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, vortexing for 30s, fully mixing, standing for 30min at room temperature, and adding 5 mu l of ultrapure water. The total volume of the dialysate was 200. mu.l, with IS and PCS at a concentration of 50. mu.g/ml.
Preparing a salvianolic acid sample: taking 180 mu l of human mixed plasma sample, adding 10 mu l of IS aqueous solution and 5 mu l of PCS aqueous solution, swirling for 30s, fully mixing, standing for 30min at room temperature, respectively adding high-concentration (50 mu g/ml), medium-concentration (10 mu g/ml) and low-concentration (5 mu g/ml) alkannic acid, salvianolic acid A, tanshinol, caffeic acid, salvianolic acid B, salvianolic acid C, salvianolic acid D, salvianolic acid E, protocatechuic acid, protocatechuic aldehyde and rosmarinic acid aqueous solutions, respectively, wherein each 20 mu l of caffeic acid, protocatechuic aldehyde and rosmarinic acid aqueous solutions are insoluble in pure water and are dissolved by 20% acetonitrile aqueous solution, swirling for 30s, fully mixing. The total volume of the dialysate was 200. mu.l, wherein the high, medium and low concentrations of each salvianolic acid component were 50, 10 and 5. mu.g/ml, respectively.
1.2 extracorporeal hemofiltration method as in item 1.2 of example 3 and data analysis method as in item 1.3 of example 3.
2. Results of the experiment
Both protocatechualdehyde and rosmarinic acid significantly increased the filtration efficiency of IS and were concentration dependent (a in fig. 4). Low concentration protocatechuic aldehyde (5 mug/ml) has no significant effect on IS filtration, and medium concentration (10 mug/ml) and high concentration protocatechuic aldehyde (50 mug/ml) can respectively improve the filtration efficiency of IS by 3.15% and 18.38% (P < 0.01). While the rosmarinic acid can not only show the effect of promoting IS filtration under high concentration, but the high concentration rosmarinic acid (50 mu g/ml) can improve the filtration efficiency of IS by 12.14 percent (P < 0.01).
And protocatechuic aldehyde and rosmarinic acid can obviously improve the filtration efficiency of PCS under the high concentration level, and the high concentration protocatechuic aldehyde and rosmarinic acid (50 mu g/ml) can respectively improve the filtration efficiency of PCS by 27.03 percent (P <0.01) and 16.44 percent (P < 0.05).
The experimental results show that the components such as the danhong injection, the phenolic acids and the like can obviously improve the clinical dialysis effect and the filtering effect of the protein-bound uremic toxin, improve the clearing efficiency of the protein-bound uremic toxin, reduce the uremic toxin level in the body of a nephrotic patient, improve the life quality of the nephrotic patient and have important application prospects.
Claims (1)
1. The application of the danhong injection in preparing the medicine preparation for improving the dialyzing rate and the filtering rate of indoxyl sulfate and paracresol sulfate protein-conjugated uremic toxin in the treatment processes of hemodialysis, peritoneal dialysis and hemofiltration.
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Non-Patent Citations (3)
| Title |
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| 丹红注射液7 种药效物质基础的人血浆蛋白结合率;张倩，等;《中国药理学通报》;20170531;第33卷(第5期);712-718 * |
| 丹红注射液治疗慢性肾衰竭维持性血液透析患者血瘀证65 例临床分析;周成，等;《中国实验方剂学杂志》;20150831;第21 卷(第16 期);167-170 * |
| 尿毒症毒素研究进展;陈斌等;《吉林医学》;20130831;第34 卷(第22 期);4513-4515 * |
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