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For wild natural medicine, unanticipated biodiversity as species or varieties with similar morphological characteristics and sympatric distribution may co-exist in a single batch of medical materials, which affects the efficacy and safety of clinical medication. DNA barcoding as an effective species identification tool is limited by its low sample throughput nature. In this study, combining DNA mini-barcode, DNA metabarcoding and species delimitation method, a novel biological sources consistency evaluation strategy was proposed, and high level of interspecific and intraspecific variations were observed and validated among 5376 Amynthas samples from 19 sampling points regarded as "Guang Dilong" and 25 batches of proprietary Chinese medicines. Besides Amynthas aspergillum as the authentic source, 8 other Molecular Operational Taxonomic Units (MOTUs) were elucidated. Significantly, even the subgroups within A. aspergillum revealed here differ significantly on chemical compositions and biological activity. Fortunately, this biodiversity could be controlled when the collection was limited to designated areas, as proved by 2796 "decoction pieces" samples. This batch biological identification method should be introduced as a novel concept regarding natural medicine quality control, and to offer guidelines for in-situ conservation and breeding bases construction of wild natural medicine.
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Objective To investigate whether Danhong injection can enhance the therapeutic effect of neural stem cell (NSC) transplantation in repairing cerebral ischemia injury by regulating the nuclear factor E2-related factor 2 (Nrf2) signaling pathway. Methods Forty male SD rats were randomly divided into the NSC transplantation group (NSC group), Danhong injection group (DH group), NSC+ Danhong injection group (N+D group), NSC+ Danhong injection group +ML385 group(N+D+M group) and PBS control group (PBS group), 8 rats in each group. All rat models of cerebral ischemia were established by embolization of the middle cerebral artery. Reperfusion was performed at 1.5 h after embolization. All rats in each group received corresponding interventions at 3 d after reperfusion. The neurological function score was evaluated before and 1, 2, 4 weeks after NSC transplantation. All rats were sacrificed at 4 weeks after NSC transplantation. The parameters related to oxidative stress were detected. The expression levels of neuron-specific nuclear protein (NeuN) and von Willebrand factor (vWF) were determined by immunofluorescence staining. Results Before NSC transplantation, the neurological function scores did not significantly differ among different groups (all P > 0.05). At postoperative 1, 2 and 4 weeks, the neurological function scores in the NSC, DH and N+D groups were significantly lower than those in the PBS and N+D+M groups (all P < 0.05). Compared with the PBS and N+D+M groups, the malondialdehyde (MDA) levels were significantly decreased, whereas the superoxide dismutase (SOD) and glutathione peroxidase (GPX) levels were considerably increased in the NSC, DH and N+D groups (all P < 0.05). The GPX level in the N+D+M group was significantly lower than that in the PBS group (P < 0.05). Immunofluorescence staining showed that the transplant NSC in the rat brain migrated to the surrounding area of cerebral infarction and survived, and expressed neuronal marker NeuN and neovascularization marker vWF. However, the number of living NSC in the N+D+M group was significantly lower compared with those in the remaining groups. Conclusions Danhong injection may improve the microenvironment of stem cell transplantation, enhance the survival rate of transplant NSC and improve the therapeutic effect of NSC transplantation for cerebral ischemia injury probably by regulating the Nrf2 signaling pathway.
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OBJECTIVE： To establish the method for simultaneous determination of 69 kinds of pesticide residues in Paeonia tactilora， Astragalus membranaceus， Ranunculus ternatus and Cornus officinalis. METHODS： GC-MS/MS method was adopted. The determination was performed on HP-5MS fused silica capillary column with splitless injecting samples. The injector temperature was set at 240 ℃， and sample size was 1 μL. The carrier gas was high-purity helium， the inlet mode was constant pressure， the pre-column pressure was 146 kPa， and the temperature was programmed. Triple four-pole tandem mass spectrometry was used for the detection， and electron impact ion source was used as ion source. The temperature of the ion source was 230 ℃， ionization energy was 70 eV， and the collision gas was nitrogen. The inlet temperature was 280 ℃ and four-pole temperature was 150 ℃， mass spectrometry monitoring mode was multi-reaction monitoring （MRM）， and solvent delay time was 5 min. RESULTS： The linear range of 69 kinds of pesticide residue was 4.82-399.6 ng/mL （all r＞0.990）. LODs were all in the range of 0.001 7-0.013 3 mg/kg， and LOQs were all in the range of 0.000 5-0.004 mg/kg. RSDs of precision and stability tests were less than 10％ （n＝6）. RSD of reproducibility test was lower than 5％ （n＝6， only pesticide amidine and permethrin were detected）. The recoveries were in the range of 62.9％-123.5％ （all RSD＜10％， n＝6）. Among 12 batches of samples， dichlorvos and diphenylamine were detected in C. officinalis； chlordimeform and permethrin were detected in A. membranaceus； diphenylamine and chlordimeform were detected in P. tactilora； diphenylamine and vinclozolin were detected in R. ternatus. CONCLUSIONS： The method is simple in operation and reproducible for simultaneous determination of 69 kinds of pesticide residue in P. tactilora， A. membranaceus， R. ternatus and C. officinalis.
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Objective To explore the expression levels of HIF-1 α and MUC5AC,MUC5B mRNA in chronic rhinosinusitis(CRS) and their correlation.Methods Eighty cases of CRS with nasal polyps(CRSwNP) treated in our hospital from May 2014 to May 2015,80 cases of CRS without NP(CRSsNP) and 80 healthy people without CRS(control group) were chosen.Nasal sinus mucosa in 3 groups were collected and treated.The expressions of HIF-1 α,MUC5AC and MUC5B mRNA were measured by semiquantitative reverse transcription polymerase chain reaction(RT-PCR),and their correlations were analyzed.Results The relative expression levels of HIF-1 α,MUC5AC and MUC5B mRNA in the CRSwNP group or CRSsNP group were 1.35±0.85,1.35±0.91;0.80±0.55,0.79±0.49 and 1.18±1.01,1.21±1.02 respectively,while which in the control group were 0.42±0.33,0.43±0.36 and 0.47±0.43 respectively,the difference between the CRS groups and control group was statistically significant(P<0.05).The expression of HIF-1αin the CRSwNP group and CRSsNP group was positively correlated with MUC5AC,MUC 5B mRNA(r=0.476,P=0.023,r=0.476,P=0.026,r=0.478,P=0.035,r=0.508,P=0.021).The ROC analysis results showed AUC=0.956 4.Conclusion The expression levels of HIF-1α and MUC5AC,MUC5B mRNA in chronic rhinosinusitis are significantly upregulated,moreover HIF-1α is closely related with MUC5AC and MUC5B mRNA.
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This article was aimed to study the quality control of pineapple leaf in order to further develop the appli-cations of pineapple leave as medicinal resources. The tissue structure of pineapple leaf was observed by tissue biop-sy assay. Thin-layer chromatography (TLC) was employed on the qualitative identification of ananasate in the leave. The p-coumaric acid and ananasate were detected with high-performance liquid chromatography (HPLC) assay in the content determination. The results showed that pineapple leaf can be divided into the upper and lower leaf epidermis with stoma under microscope observation. Ananasate of the leave can be indentified clearly by TLC. HPLC method can accurately detect the content of p-coumaric acid and ananasate of pineapple leave. There are different contents of p-coumaric acid and ananasate of pineapple leave in different origins. In Y unnan province, the contents of p-coumaric acid and ananasate of pineapple leave are the highest; while the contents of p-coumaric acid and ananasate in Guangxi province are the lowest. It was concluded that ananasate can be used in the quality identification. The p-coumaric acid and ananasate can be used in the quantitative determination in the better control of the quality of pineapple leaf.
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AIM:To establish a method for determing loganin and paeonol in Liuwei Dihuang Granules(Radix Rehmannial praeparata,Fructus Corni,Cortex Moutan,Rhizoma Dioscoreae,etc.).METHODS:Loganin was determined using acetonitrile-water(15∶85) as the mobile phase with UV detection at 236 nm,and paeonol was determined using methanol-water(70∶30) as the mobile phase with UV detection at 274 nm by HPLC with Diamonsil C_ 18(150 mm?4.6 mm,3 ?m) column.RESULTS:The linear ranges of loganin and paeonol were in the ranges of 2.34-234 ?g/mL(r=0.999 99),7.07-141.40 ?g/mL(r=0.999 6),respectively.The average recoveries of loganin and paeonol were 100.06%,95.75% with RSD of 3.10%,0.65%,respectively.CONCLUSION:This method is simple,accurate,reproducible and can be used for determining loganin and paeonol in Liuwei Dihuang Granules.
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AIM: To establish a quick,accurate,sensitive method for analysis of theophylline,prednisone acetate and dexamethasone acetate illegally added into traditional Chinese medicinal preparation for antitussive and antiasthmastics by UPLC-MS. METHODS: The UPLC-MS/MS method was used.Electrospay ionization(ESI) source was applied and operated in the positive and negative ion mode.Theophylline,prednisone acetate and dexamethasone acetate illegally added into traditional Chinese medicinal preparations were identified. RESULTS: Both theophylline and prednisone acetate were found in formulations. CONCLUSION: The method is selective and sensitive and can be used to detect the theophylline,prednisone acetate and dexamethasone acetate illegally added into traditional Chinese medicinal preparations.
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AIM: To establish a quick,accurate,sensitive method for analysing sildenafil citrate illegally added into traditional Chinese medicinal preparation. METHODS: The ultra performance liquid chromatography-quadrupole mass spectrometry method was used.Electrospay ionization(ESI) source was applied and operated in the positive and negative ion mode.The compound of sildenafil citrate illegally added into traditional Chinese medicinal preparations were identified according to the spectrum,chromatographic behavior and mass spectral data by comparison with those of reference substance. RESULTS: Sildenafil citrate was found in two formulations of three. CONCLUSION: The method is selective and sensitive and can be used to detect the sildenafil citrate illegally added into traditional Chinese medicinal preparations.
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AIM: To establish the HPLC method for determinating syringin and betaine in Qianglining Tablet(Radix et Rhizoma seu Caulis Acanthopanacis senticosi,Fructus Lycii). METHODS: A Hypersil C_18 column was used for the determination of syringin.The mobile phase for syringin was methanol-0.1 mol/L acetic acid (24∶76),and the detection wavelength was at 270 nm.A Hypersil NH_2 column was used for the determination of betaine.The mobile phase for betaine was acetonitrile-water(25∶75),and ELS detector was used. RESULTS: The linear range of syringin and betaine were 0.121 6-0.608 0 ?g(r=0.999 9) and 0.660 8-3.964 8 ?g(r=0.999 1),respectively.The average recoveries of syringin and betaine were 99.09%(RSD=2.13%) and 96.34%(RSD=1.31%),respectively. CONCLUSION: The method is simple,accurate,and can be used for the quality control of Qianglining Tablet.
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The appearance and changes of Acetylcholinesterase (ACHE) in the brains of 62 rats from early embryos to adults were studied by histochemical method and the brains of 19 rats of corresponding stages were examined with Nissl stain. The main results are as follows:1. AChE appeared in the following sequence: spinal cord, myelencephalon, metencephalon, mesencephalon, diencephalon and finally in telencephalon. In telencephaIon, AChE appeared earlier in paleocortex than in neocortex. The sequence described above showed that the appearance of AChE reflects cephalization of the nervous system in the rat.2. AChE-positive neurons of the rat brain consisted of cholinergic neurons and cholinoceptive ceils, both of which are related to the neurotransmitter ACh.3. The following nuclei of adult rat brain were shown to be AchE-positive in our specimens, but had not been reported in available literature: n. tractus mesencephali, n. anterior dorsalis thalami, n. anterior ventralis thalami, n. lateralis thalami, n. habenulae lateralis, n. reticularis thalami, n. supraopticus, n. paraventricularis, n. parafascicularis and n. ruber. In the cerebral cortex very few positive cells have been noticed.