Total Flavones of Abelmoschus manihot Remodels Gut Microbiota and Inhibits Microinflammation in Chronic Renal Failure Progression by Targeting Autophagy-Mediated Macrophage Polarization.

BACKGROUND: Recently, progression of chronic renal failure (CRF) has been closely associated with gut microbiota dysbiosis and intestinal metabolite-derived microinflammation. In China, total flavones of Abelmoschus manihot (TFA), a component of Abelmoschus manihot, has been widely used to delay CRF progression in clinics for the past two decades. However, the overall therapeutic mechanisms remain obscure. In this study, we designed experiments to investigate the renoprotective effects of TFA in CRF progression and its underlying mechanisms involved in gut microbiota and microinflammation, compared with febuxostat (FEB), a potent non-purine selective inhibitor of xanthine oxidase. METHODS: In vivo, the CRF rat models were induced by uninephrectomy, potassium oxonate, and proinflammatory diet, and received either TFA suspension, FEB, or vehicle after modeling for 28 days. In vitro, the RAW 264.7 cells were exposed to lipopolysaccharide (LPS) with or without TFA or FEB. Changes in parameters related to renal injury, gut microbiota dysbiosis, gut-derived metabolites, and microinflammation were analyzed in vivo. Changes in macrophage polarization and autophagy and its related signaling were analyzed both in vivo and in vitro. RESULTS: For the modified CRF model rats, the administration of TFA and FEB improved renal injury, including renal dysfunction and renal tubulointerstitial lesions; remodeled gut microbiota dysbiosis, including decreased Bacteroidales and Lactobacillales and increased Erysipelotrichales; regulated gut-derived metabolites, including d-amino acid oxidase, serine racemase, d-serine, and l-serine; inhibited microinflammation, including interleukin 1ß (IL1ß), tumor necrosis factor-α, and nuclear factor-κB; and modulated macrophage polarization, including markers of M1/M2 macrophages. More importantly, TFA and FEB reversed the expression of beclin1 (BECN1) and phosphorylation of p62 protein and light chain 3 (LC3) conversion in the kidneys by activating the adenosine monophosphate-activated protein kinase-sirtuin 1 (AMPK-SIRT1) signaling. Further, TFA and FEB have similar effects on macrophage polarization and autophagy and its related signaling in vitro. CONCLUSION: In this study, we demonstrated that TFA, similar to FEB, exerts its renoprotective effects partially by therapeutically remodeling gut microbiota dysbiosis and inhibiting intestinal metabolite-derived microinflammation. This is achieved by adjusting autophagy-mediated macrophage polarization through AMPK-SIRT1 signaling. These findings provide more accurate information on the role of TFA in delaying CRF progression.

Qualitative and quantitative analysis of saponins in the flower bud of Panax ginseng (Ginseng Flos) by UFLC-Triple TOF-MS/MS and UFLC-QTRAP-MS/MS.

INTRODUCTION: Ginseng Flos (GF), the flower bud of Panax ginseng, is a worthy functional food with medicinal potential. A few studies have focused on the comprehensive and systematic analysis of its major bioactive constituents. OBJECTIVE: The aims are to develop the methods of ultra-fast liquid chromatography coupled with triple quadrupole-time of flight tandem mass spectrometry (UFLC-Triple TOF-MS/MS) and ultra-fast liquid chromatography coupled with triple quadrupole-linear ion trap tandem mass spectrometry (UFLC-QTRAP-MS/MS) for the qualitative and quantitative analysis of the saponins in GF. METHODOLOGY: UFLC-Triple TOF-MS/MS and UFLC-QTRAP-MS/MS were established for the qualitative and quantitative analysis of the saponins in GF, separately. RESULTS: Fifty-one saponins were identified in GF using UFLC-Triple TOF-MS/MS method; among them, 21 saponins were characterized by comparing with standards. Furthermore, 12 ginsenosides (ginsenoside Re, Rg1 , Rf, 20(S)-Rg2 , 20(R)-Rg2 , Rb1 , Rc, Ro, Rb2 , F1 , Rd, and F2 ) were synchronously determined by UFLC-QTRAP-MS/MS method after the extraction with 70% methanol. This UFLC-QTRAP-MS/MS method showed good linearity (r >0.9991), the interday and intraday precision, repeatability and stability were all satisfied, the average recoveries of standard addition for the compounds were between 94.01% and 105.16%, and the relative standard deviations were less than 5%. CONCLUSION: The results are available for the comprehensive quality control and assessment of GF and its relative products.

Quality Evaluation of Wild and Cultivated Schisandrae Chinensis Fructus Based on Simultaneous Determination of Multiple Bioactive Constituents Combined with Multivariate Statistical Analysis.

Schisandrae Chinensis Fructus, also called wuweizi in China, was a widely used folk medicine in China, Korea, and Russia. Due to the limited natural resources and huge demand of wuweizi, people tend to cultivate wuweizi to protect this species. However, the quality of wild and cultivated herbs of the same species may change. Little attention has been paid to comparing wild and cultivated wuweizi based on simultaneous determination of its active components, such as lignans and organic acids. An analytical method based on UFLC-QTRAP-MS/MS was used for the simultaneous determination of 15 components, including 11 lignans (schisandrin, gomisin D, gomisin J, schisandrol B, angeloylgomisin H, schizantherin B, schisanhenol, deoxyschizandrin, γ-schisandrin, schizandrin C, and schisantherin) and 4 organic acids (quinic acid, d(-)-tartaric acid, l-(-)-malic acid, and protocatechuic acid) in wuweizi under different ecological environments. Principal components analysis (PCA), partial least squares discrimination analysis (PLS-DA), independent sample t-test, and gray relational analysis (GRA) have been applied to classify and evaluate samples from different ecological environments according to the content of 15 components. The results showed that the differential compounds (i.e., quinic acid, l-(-)-malic acid, protocatechuic acid, schisandrol B) were significantly related to the classification of wild and cultivated wuweizi. GRA results demonstrated that the quality of cultivated wuweizi was not as good as wild wuweizi. The protocol not just provided a new method for the comprehensive evaluation and quality control of wild and cultivated wuweizi, but paved the way to differentiate them at the chemistry level.

Huangkui capsule alleviates renal tubular epithelial-mesenchymal transition in diabetic nephropathy via inhibiting NLRP3 inflammasome activation and TLR4/NF-κB signaling.

BACKGROUND: Huangkui capsule (HKC), an anti-inflammatory Chinese modern patent medicine, has been now widely applied to the clinical therapy of diabetic nephropathy (DN). However, the overall therapeutic mechanisms in vivo are still unclear. Renal tubular epithelial-to-mesenchymal transition (EMT) is one of the major pathogenesis of renal interstitial fibrosis in DN. Recently, the physiological roles of NLRP3 inflammasome activation and toll-like receptor 4 (TLR4)/nuclear factor (NF)-κB signaling are closely linked to EMT. But, it remains elusive whether HKC regulates renal tubular EMT in vivo through targeting NLRP3 inflammasome activation and TLR4/NF-κB signaling in the kidneys. PURPOSE: This study thereby aimed to clarify the therapeutic effects of HKC on renal tubular EMT in DN and its underlying mechanisms in vivo, compared to rapamycin (RAP). METHODS: Thirty-two rats were randomly divided into 4 groups: the Sham group, the Vehicle group, the HKC group and the RAP group. The early DN rat models were induced by unilateral nephrectomy combined with intraperitoneal injection of streptozotocin, and administered with HKC suspension or RAP suspension or vehicle after modeling for 4 weeks. Changes in the incipient renal lesions-related parameters in urine and blood were analyzed, respectively. Renal interstitial tissues were isolated for histomorphometry, immunohistochemistry and Western blotting at sacrifice. RESULTS: For the early DN rat models, HKC at the suitable dose of 2 g/kg/day ameliorated the general condition and biochemical parameters partially including kidney weight (KW), urinary albumin (UAlb), serum creatinine (Scr) and serum albumin (Alb), attenuated renal tubular EMT significantly and inhibited the activation of NLRP3 inflammasome in the kidneys obviously, which was superior to RAP generally. In addition to these, HKC also suppressed TLR4/NF-κB signaling in the kidneys of the DN model rats accurately, which was different from RAP specifically. CONCLUSION: The results of this study further indicated that HKC, different from RAP, can alleviate renal tubular EMT in the DN model rats, likely by inhibiting NLRP3 inflammasome activation and TLR4/NF-κB signaling in the kidneys. Our findings thus provide the more accurate information in vivo about a clinical value of HKC, a traditional anti-inflammatory phytomedicine, in the treatment of the early DN patients.

Hyperoside attenuates renal aging and injury induced by D-galactose via inhibiting AMPK-ULK1 signaling-mediated autophagy.

The kidney is a typical organ undergoing age and injury. Hyperoside is reported to be useful for preventing aging induced by D-galactose (D-gal). However, therapeutic mechanisms remain unclear. We thereby aimed to verify whether hyperoside, compared to vitamin E (VE), could alleviate renal aging and injury by regulating autophagic activity and its related signaling pathways. In vivo, rats were administered with either hyperoside or VE after renal aging modeling induced by D-gal. Changes in renal aging and injury markers, autophagic activity and AMPK-ULK1 signaling pathway in the kidneys were analysed. In vitro, the NRK-52E cells exposed to D-gal were used to investigate regulative actions of hyperoside and VE on cell viability, renal tubular cellular aging markers, autophagic activity and its related signaling pathways by histomorphometry, immunohistochemistry, immunofluorescence, lentiviral transfection and Western blot. Aging and injury in the kidneys and renal tubular cells induced by D-gal were ameliorated by hyperoside and VE. Hyperoside and VE inhibited autophagic activity through mTOR-independent and AMPK-ULK1 signaling pathways. Hyperoside, as a component of phytomedicine similar to VE, attenuated renal aging and injury induced by D-gal via inhibiting AMPK-ULK1-mediated autophagy. This study provides the first evidence that hyperoside contributes to the prevention of age-associated renal injury.

[Comparative analysis of multiple index constituents in Ophiopogonis Radix and Liriopes Radix].

An analytical method based on UFLC-QTRAP-MS/MS was established for simultaneous determination of thirty-three components including steroidal saponins, homoisoflavonoids, amino acids and nucleosides in Ophiopogonis Radix. Thirty-three target components of commercial medicinal materials of Maidong were comparative analysis. Synergi™ Hydro-RP 100 column (2.0 mm × 100 mm, 2.5 µm) was used with 0.1% formic acid solution-0.1% formic acid acetonitrile for gradient elution at a flow rate of 0.4 mL·min⁻¹. In addition, multiple reaction monitoring (MRM) mode was employed. The data were comprehensively processed and analyzed with hierarchical clustering analysis(HCA), principal component analysis(PCA) and partial least squares discriminant analysis(PLS-DA) methods. All components showed good linearity(r>0.999 0) within the tested ranges. The average recoveries were between 96.23%-102.0%, and the relative standard deviation(RSD) were less than 5%. The results showed that there were significant differences in components between Ophiopogonis Radix and Liriopes Radix, with seven components obviously different. This method was useful for providing basis for the comprehensive evaluation and intrinsic quality control of Ophiopogonis Radix and Liriopes Radix , and may provide a new method reference for the identification of Ophiopogonis Radix and Liriopes Radix.

[Simultaneous determination of lignans and organic acids in Schisandrae Chinensis Fructus by UFLC-Q-TRAP-MS/MS].

An analytical method based on UFLC-QTRAP-MS/MS was developed for simultaneous determination of fifteen components including eleven lignans (schizantherin B, schisandrol B, schizandrin C, γ-schisandrin, deoxyschizandrin, schisantherin, schisandrin, schisanhenol, gomisin D, gomisin J, and angeloylgomisin H) and organic acids (S)-malic acid, D(-)-tartaric acid, protocatechuic acid, and quinic acid) in Schisandrae Chinensis Fructus. Samples from different product specifications were evaluated and analyzed. The chromatographic separation was performed on a Synergi™ Hydro-RP 100Å column (2.0 mm×100 mm, 2.5 µm) at 40 °C with a gradient elution by employing 0.1% aqueous formic acid (A)-acetonitrile (B) as the mobile phase, and the flow rate was 0.4 mL·min⁻¹, using an electrospray ionization (ESI) source and multiple reaction monitoring (MRM) mode. Fifteen components were evaluated synthetically by TOPSIS and gray related degree. The results showed that fifteen components had good linearity (r>0.999 90), and the limits of detection were all satisfactory. The average recoveries of standard addition for the compounds were between 95.42 % and 98.86 %, and the relative standard deviations were less than 5%. The greatest difference of ri in grey related degree was 58.1%, whilst the greatest difference of Ci value in TOPSIS method was 94.8%. The results of these two methods showed that the holistic quality of No. 14 sample was the best. The developed method was accurate and reliable, which was suitable for the simultaneous determination of multiple functional substances and able to provide a new basis for the comprehensive assessment and overall control of the quality of Schisandrae Chinensis Fructus.

Inhibition of Akt/mTOR/p70S6K Signaling Activity With Huangkui Capsule Alleviates the Early Glomerular Pathological Changes in Diabetic Nephropathy.

Huangkui capsule (HKC), a Chinese modern patent medicine extracted from Abelmoschus manihot (L.) medic, has been widely applied to clinical therapy in the early diabetic nephropathy (DN) patients. However, it remains elusive whether HKC can ameliorate the inchoate glomerular injuries in hyperglycemia. Recently the activation of phosphatidylinositol-3-kinase (PI3K)/serine-threonine kinase (Akt)/mammalian target of rapamycin (mTOR) signaling and its downstream regulator, 70-kDa ribosomal protein S6 kinase (p70S6K), play important roles in the early glomerular pathological changes of DN including glomerular hypertrophy, glomerular basement membrane (GBM) thickening and mild mesangial expansion. This study thereby aimed to clarify therapeutic effects of HKC during the initial phase of DN and its underlying mechanisms. Fifteen rats were randomly divided into 3 groups: the normal group, the model group and the HKC group. The early DN model rats were induced by unilateral nephrectomy combined with intraperitoneal injection of streptozotocin, and administered with either HKC suspension or vehicle after modeling and for a period of 4 weeks. Changes in the incipient glomerular lesions-related parameters in urine and blood were analyzed. Kidneys were isolated for histomorphometry, immunohistochemistry, immunofluorescence and Western blotting (WB) at sacrifice. In vitro, murine mesangial cells (MCs) were used to investigate inhibitory actions of hyperoside (HYP), a bioactive component of HKC, on cellular hypertrophy-associated signaling pathway by WB, compared with rapamycin (RAP). For the early DN model rats, HKC ameliorated micro-urinary albumin, body weight and serum albumin, but had no significant effects on renal function and liver enzymes; HKC improved renal shape, kidney weight and kidney hypertrophy index; HKC attenuated glomerular hypertrophy, GBM thickening and mild mesangial expansion; HKC inhibited the phosphorylation of Akt, mTOR and p70S6K, and the protein over-expression of transforming growth factor-ß1 in kidneys. In vitro, the phosphorylation of PI3K, Akt, mTOR and p70S6K in MCs induced by high-glucose was abrogated by treatment of HYP or RAP. On the whole, this study further demonstrated HKC safely and efficiently alleviates the early glomerular pathological changes of DN, likely by inhibiting Akt/mTOR/p70S6K signaling activity in vivo and in vitro, and provided the first evidence that HKC directly contributes to the prevention of the early DN.

Renal protective effect and action mechanism of Huangkui capsule and its main five flavonoids.

ETHNOPHARMACOLOGICAL RELEVANCE: The flower of Abelmoschus manihot (Linn.) Medicus (A. manihot), as a traditional Chinese Herbal medicine, was used widely in China with efficacy of inducing diuresis for treating strangurtia, and subdhing swelling and detoxicating. It has been reported that Huangkui capsule, prepared by the extract of the flower of A. manihot, can reduce the content of urinary protein, serum creatinine and serum urea nitrogen in nephropathy rats and processes renoprotective activity, while the action mechanism need to illuminate deeply. AIMS OF THE STUDY: In this study, we investigated the protection effect of Huangkui capsule on tubulointerstitial fibrosis in chronic renal failure (CRF) rats and its mechanism against high glucose-induced epithelial to mesenchymal transition (EMT) in renal tubular epithelial cells (HK-2) of its bioactive components. MATERIALS AND METHODS: The animals were divided into normal group, CRF model group and Huangkui capsule-treated group. Hematoxylin eosin (HE) staining and Masson staining were applied to observe pathological changes in renal tissue of different groups. Biochemical indicators including serum urea nitrogen (BUN), urine protein (UP) and serum creatinine (Scr) were measured according to the manufacturer's instructions of kits. HK-2 cell damaged model was established to access the protection effect and action mechanism of five main flavonoids from Huangkui capsule. The experimental cells were divided into eight groups: control group, model group, positive drug group and five main flavonoids treated groups. The dichlorodihydrofluorescein diacetate (DCFH-DA) assay was used to determine the reactive oxygen species (ROS) in different groups. Western blot was applied to analyze the expression of pathogenesis-related proteins in different groups. RESULTS: The results stated that Huangkui capsule significantly inhibited the elevation of Scr, BUN, UP, the expression of α-smooth muscle actin (α-SMA), phosphorylation-extracellular signal-regulated kinase (p-ERK1/2), NADPH Oxidase 1, NADPH Oxidase 2 and NADPH Oxidase 4 in adenine-induced CRF rats. The main bioactive components of quercetin (QT), hyperoside (HY), isoquercitrin (IQT), gossypetin-8-O-ß-D-glucuronide (GG) and quercetin-3'-O-glucoside (QG) at the dosage of 100µM, like NADPH oxidase inhibitor diphenyleneiodonium, exhibited a significant effect on inhibiting the expression of α-SMA, p-ERK1/2, NADPH Oxidase 1, NADPH Oxidase 2 and NADPH Oxidase 4 in high glucose-induced HK-2 cells, especially GG. CONCLUSIONS: These results demonstrated that Huangkui capsule and the flavonoids components prevent tubulointerstitial fibrosis in CRF rat involvement in the action mechanism of inhibiting NADPH oxidase/ROS/ERK pathway.

[Antidepressant activity of flavonoid ethanol extract of Abelmoschus manihot corolla with BDNF up-regulation in the hippocampus].

Abelmoschus manihot (L.) Medic., a folk herbal medicine in China, is a flowering plant belonging to Abelmoschus L. genus and Malvaceae family, which has been reported with an antidepressant activity. The study was designed to isolate flavonoids from Abelmoschus manihot corolla and explore the action mechanism of antidepressant activities. The flavonoids were isolated and purified by D101 macroporous resin column, polyamide column and Sephadex LH-20 sequentially and identified as myricetin-3-O-ß-D-glucoside (1), gossypetin-8-O-ß-D-glucuronide (2, G-8-G), gossypetin-3'-O-ß-D-glucoside (3), quercetin-3'-glucoside (4, Q-3-G), isoquercitrin (5, IQT), hyperoside (6, HY), myricetin (7), quercetin (8, QT). Compounds 2, 4, 5, 6 and 8 (15, 30 and 60 mg·kg−1) were orally administered to mice and the reaction was observed in tail suspension test (TST) and forced swimming test (FST). Western blot analysis was used in determination of the protein expressions of brain-derived neurotrophic factor (BDNF), tyrosine receptor kinase B (TrkB) and phosphorylation eukaryotic elongation factor 2 (p-eEF2). The results revealed that only Q-3-G and G-8-G (15, 30, 60 mg ·kg−1) significantly reduced the immobility time in FST and TST. Furthermore, Q-3-G and G-8-G remarkably increased the expression of BDNF and TrkB, and decreased the expression of p-eEF2. These results suggest that Q-3-G and G-8-G had an obvious antidepressant activity via up-regulation of BDNF expression. The new observation will provide a new direction in the development of antidepressant in the treatment of major depressive disorder (MDD).

[Analysis and utilization value discussion of multiple chemical composition in different tissues of Abelmoschus manihot].

This research is to analyze the resourceful chemical composition in different tissues (root, stem, leaf and flower) of Abelmoschus manihot and evaluate their utilizing value. The flavonoids, soluble polysaccharides, cellulose, nucleosides and amino acids in the different tissues of A. manihot were determined by HPLC coupled with UV-Vis spectrophotpmetry, and UPLC-TQ/MS. The flowers are rich in the resourceful chemical compositions of flavonoids which mainly consist of hyperoside, isoquercitrin, cotton-8-O-glucuronide, myricetin, quercetin-3'-O-glucoside, rutin and quercetin. The total content of these flavonoids is 25.450 mg•g-1 in the flowers, while they are trace in the other tissues.Different tissues of A. manihot are rich in soluble polysaccharides and celluloses and the stems have the highest content(19.76%) of soluble polysaccharides, while the roots have the highest content (29.88%) of cellulose. Total of 21 amino acids and 9 nucleosides were detected in this plant, and the flowers have the highest content of amino acids(4.737 mg•g⁻¹), while the leaves have the highest content of nucleosides (1.474 mg•g⁻¹). A. manihot is rich in the resourceful chemical compositions, and its constituents and contents are various in different tissues of this plant.The results provided a scientific basis for the utilization and industrial development of A. manihot plants.

Huangkui capsule attenuates renal fibrosis in diabetic nephropathy rats through regulating oxidative stress and p38MAPK/Akt pathways, compared to α-lipoic acid.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional Chinese medicine (TCM), Abelmoschus manihot (L.) medic (AM) is a natural medicinal plant used for the treatment of inflammatory diseases. Recently, Huangkui capsule (HKC), a Chinese patent medicine extracted from AM, has been widely applied to the clinical therapy of renal fibrosis in patients with early diabetic nephropathy (DN). However, the therapeutic mechanisms involved in vivo remain ambiguous. The goal of this study is to expound the mechanism in vivo of HKC in order to deepen the understanding of its clinical effects, by using the approaches of contrasting the dose-effects of HKC on oxidative stress (OS) in the kidney compared to α-lipoic acid (LA), and then demonstrating whether and how anti-oxidative properties of HKC or LA might be beneficial for the treatment of renal fibrosis in vivo. MATERIALS AND METHODS: Thirty-three rats were divided into 5 groups, a Sham group, a Vehicle group, a L-HKC group, a H-HKC group and a LA group. The different doses of HKC, LA and distilled water were daily administrated for 8 weeks after the induction of DN by the unilateral nephrectomy combined with streptozotocin (STZ) intraperitoneal injections. Rat's general status, biochemical parameters, renal histological changes and OS indicators, as well as the key protein expressions in p38 mitogen-activated protein kinase (p38MAPK)/serine-threonine kinase (Akt) signaling pathways and downstream cytokines including transforming growth factor (TGF)-ß1 and tumor necrosis factor (TNF)-α were examined, respectively. RESULTS: HKC and LA ameliorated body weight, kidney weight, urinary albumin and renal function including blood urea nitrogen and serum uric acid, attenuated renal fibrosis including the cell numbers and extracellular matrix rate in glomerulus, and controlled OS indicators including malondialdehyde, total superoxide dismutase, 8-hydroxy-2'-deoxyguanosine and nicotinamide adenine dinucleotide phosphate oxidase 4, but did not lower blood glucose in DN model rats. Among them, the anti-renal fibrosis effect of H-HKC was better than that of LA. In addition, HKC simultaneously down-regulated the protein expressions of phosphorylated p38MAPK, phosphorylated Akt (p-Akt), TGF-ß1 and TNF-α in the kidney of DN model rats, unlike HKC, LA only down-regulated p-Akt and TNF-α protein expressions. CONCLUSION: We have demonstrated that HKC, similar to LA, is renoprotective via attenuating OS and renal fibrosis in the DN rat model. The potential mechanisms by which HKC and LA exert their therapeutic effects in vivo are respectively through down-regulating the activation of p38MAPK and/or Akt pathways as well as the expressions of TGF-ß1 and/or TNF-α in the kidney. Our findings thus provide the useful information about a clinical combination of HKC and LA in early DN patients.

[Strategies and key technologies for risk control and management in quality of Chinese medicine injections based on integrated pharmacology].

This paper focuses on the quality risk control and management of Chinese medicine (CM) injections. The most important technological requirements are analyzed, and a strategy for integrated pharmacology to study CM mechanism is proposed. A key technology system for quality risk control and management was further constructed. The strategy and technology system was finally applied to Shengmai injection for quality risk control and management.

[Comparative study on pseudoanaphylactoid reactions induced by pulse-activating injection before and after improving technology].

OBJECTIVE: To investigate a possibility to improve the security of pulse-activating injection by comparing the difference of pseudoanaphylactoid reactions (PR) induced by pulse-activating injection before and after improving technology. METHOD: The analysis of vascular permeability of the mice's ears: ICR mouse were divided into different test groups, and intravenously injected with solutions of different concentration of pulse-activating injection before and after improving technology, positive control Compound 48/80 and 5% glucose injection. All test substances were mixed with 0. 4% Evans blue. The reaction and vascular permeability of the ears were observed and measured 30 min after injection. The vascular permeability of the rat's skin: the rats were intravenous injected with 0. 6% Evans blue normal saline solution first, 10 minutes later, the same test substances were intradermal injected into the back of rats, there are 16 injected spots in the back of rat. The rats were sacrificed and the diameter of locus ceruleus and the content of Evans blue leaked out were measured 20 min after injection. RESULT: Pulse-activating injection before improving technology with dose of 16.7 mL x kg(-1) ( in 1.67 times the clinical dose ) caused obvious vascular hyperpermeability in ICR mice. In the group of pulse-activating injection before improving technology with dose of 10 mL x kg(-1) (in clinic equivalent dose), no obvious vascular hyperpermeability in the ears were observed. The degrees of vascular hyperpermeability in the group of pulse-activating injection after improving technology with dose of 16.7 mL x kg(-1) were more lessen than the same dose of injection before improving technology. Pulse-activating injection before improving technology caused obvious exudation, oedema locus ceruleus in the injection site of rat's back, and it showed a certain dose-effect relation. Pulse-activating injection after improving technology caused locus ceruleus in the injection site too, but the diameters of the locus ceruleus were shorter than the diameters in the group of pulse-activating injection before improving technology, and the contents of leaked out Evans blue were fewer. All of these showed that PR of skin induced by pulse-activating injection after improving technology is alleviated. CONCLUSION: Pulse-activating injection before improving technology cause obvious vascular hyperpermeability, but the same dose of pulse-activating injection after improving technology can't cause obvious vascular hyperpermeability. The result indicated that the pulse-activating injection before improving technology can cause PR, improving technology can lessen the degree of PR induced by the injection.

[Experimental study on anaphylactoid reactions induced by different components of shengmai injection (new production process) on Beagle dogs].

OBJECTIVE: To evaluate the sensitization effect of different components of Shengmai injection (new production process) on Beagle dogs. METHOD: Beagle dogs were randomly divided into 7 groups, 3 in each group. Each group was respectively injected with 5% glucose injection, Ginseng Radix et Rhizoma Rubra extract, Ophiopogonis Radix extract, Schisandrae Chinensis Fructus extract, Schisandrae Chinersis Fructus distillate, Shengmaifang, 0.2% tween 80. The changes of each dog were observed from injection before until 24 hours after injection, and the response level was determined according to the severity of the symptoms. Blood samples were collected before injection and at 10 min after injection for measuring histamine content in plasma by ELISA. Sensitization of the injection was comprehensively determined by combined the response level of symptoms and the histamine level. RESULT: One dog of Ginseng Radix et Rhizoma Rubra extract group showed untypical symptoms of anaphylactoid reactions, and serum histamine of two dogs increased more than doubled. The Beagle dogs administrated with 0.2% tween 80 showed typical symptoms of anaphylactoid reactions, while there was no significant increase of serum histamine. Other groups were observed with no typical anaphylactoid reactions. CONCLUSION: The sensitization effect of Shengmai injection (new production process) may be associated with Ginseng Radix et Rhizoma Rubra extract and 0.2% tween 80.

[Experimental study on anaphylactoid reactions induced by shengmai injection (new production process) on Cynomolgus monkey].

OBJECTIVE: To compare the anaphylactoid effect of old Shengmai injection and new Shengmai injection on Cynomolgus monkey. METHOD: Cynomolgus monkeys were randomly divided into 4 groups, and were respectively injected with 5% glucose injection, old Shengmai injection, new Shengmai injection, positive control drug. The changes of each monkey were observed from injection before until 24 hours after injection, and the response level was determined according to the severity of the symptoms. Blood samples were collected before injection and at 10 min after injection for measuring histamine content in plasma. Blood pressure and heart rates were detected before injection and at 10 min after injection. Sensitization of the injection was comprehensively determined by combined the response level of symptoms and the histamine level. RESULT: The Cynomolgus monkeys administrated with old Shengmai injection showed typical symptoms of anaphylactoid reactions and the content of serum histamine is not more than doubled. The Cynomolgus monkeys administrated with new Shengmai injection showed untypical symptoms of anaphylactoid reactions and the content of serum histamine did not rise. CONCLUSION: The old Shengmai injection can induce typical anaphylactoid reactions on Cynomolgus monkeys, and the sensitization ability is strong. The symptoms of anaphylactoid reactions induced by the new Shengmai injection appeared later and showed lesser degree with the sensitization lower.