A multicenter study on the effects of different methods of disinfecting medical external-use ultrasound probes.

BACKGROUND: Microbial contamination of external-use ultrasound probes is a serious and overlooked issue. We assessed the effects of different methods of disinfecting medical external-use ultrasound probes. METHODS: On-site disinfection experiments were conducted at 10 hospitals; the tips and sides of external-use ultrasound probes were sampled before and after disinfection, and 3 disinfection methods were assessed (use of a new ultraviolet [UV] ultrasound probe disinfector, wiping with ordinary paper towels or cleaning with disinfectant wipes). RESULTS: For the new UV probe disinfector, the median microbial death rates for the tips and sides of the external-use ultrasound probe were 93.67% and 97.50%, respectively, which were higher than those for wiping with paper towels and cleaning with disinfectant wipes (12.50% and 10.00% and 20.00% and 21.42%, respectively); the rates of microorganisms exceeding the standard were 15.0% and 13.3%, respectively, which were lower than those for wiping with paper towels and cleaning with disinfectant wipes (53.3% and 60.0% and 46.7% and 38.3%, respectively). The death rates of different species of microorganisms were high, ranging from 87.5% to 100%. CONCLUSIONS: The new UV ultrasound probe disinfector significantly reduced the risk of potential nosocomial infections according to the low microbial death rate for conventional disinfection methods.

Piperlonguminine attenuates renal fibrosis by inhibiting TRPC6.

ETHNOPHARMACOLOGICAL RELEVANCE: Liuwei Dihuang (LWDH) is a classic prescription that has been used to the treatment of "Kidney-Yin" deficiency syndrome for more than 1000 years in China. Recent studies have confirmed that LWDH can prevent the progression of renal fibrosis. Numerous studies have demonstrated the critical role that TRPC6 plays in the development of renal fibrosis. Due to the complex composition of LWDH and its remarkable therapeutic effect on renal fibrosis, it is possible to discover new active ingredients targeting TRPC6 for the treatment of renal fibrosis. AIM OF STUDY: This study aimed to identify selective TRPC6 inhibitors from LWDH and evaluate their therapeutical effects on renal fibrosis. MATERIALS AND METHODS: Computer-aided drug design was used to screen the biologically active ingredients of LWDH, and their affinities to human TRPC6 protein were detected by microcalorimetry. TRPC6, TRPC3, and TRPC7 over-expressed HEK293 cells were constructed, and the selective activities of the compounds on TRPC6 were determined by measuring [Ca2+]i in these cells. To establish an in vitro model of renal fibrosis, human renal proximal tubular epithelial HK-2 cells were stimulated with TGF-ß1. The therapeutic effects of LWDH compounds on renal fibrosis were then tested by detecting the related proteins. TRPC6 was knocked-down in HK-2 cells to investigate the effects of LWDH active ingredients on TRPC6. Finally, a unilateral ureteral obstruction model of renal fibrosis was established to test the therapeutic effect. RESULTS: From hundreds of LWDH ingredients, 64 active components with oral bioavailability ≥30% and drug-likeness index ≥0.18 were acquired. A total of 10 active components were obtained by molecular docking with TRPC6 protein. Among them, 4 components had an affinity with TRPC6. Piperlonguminine (PLG) had the most potent affinity with TRPC6 and blocking effect on TRPC6-mediated Ca2+ entry. A 100 µM of PLG showed no detectable inhibition on TRPC1, TRPC3, TRPC4, TRPC5, or TRPC7-mediated Ca2+ influx into cells. In vitro results indicated that PLG concentration-dependently inhibited the abnormally high expression of α-smooth muscle actin (α-SMA), collagen I, vimentin, and TRPC6 in TGF-ß1-induced HK-2 cells. Consistently, PLG also could not further inhibit TGF-ß1-induced expressions of these protein biomarkers in TRPC6 knocked-down HK-2 cells. In vivo, PLG dose-dependently reduced urinary protein, serum creatinine, and blood urea nitrogen levels in renal fibrosis mice and markedly alleviated fibrosis and the expressions of α-SMA, collagen I, vimentin, and TRPC6 in kidney tissues. CONCLUSION: Our results showed that PLG had anti-renal fibrosis effects by selectively inhibiting TRPC6. PLG might be a promising therapeutic agent for the treatment of renal fibrosis.

[Determination of gypenoside XLVI and LVI in Gynostemma pentaphyllum from Fujian by ultra-high performance liquid chromatography-charged aerosol detector].

Gynostemma pentaphyllum (Thunb.) Makino contains dammarane-type triterpenoid saponins, similar to ginseng, with a host of pharmacological activities. However, its planting resources and chemical composition are quite complex. The chemical constituents of Gynostemma pentaphyllum vary drastically among different origins and varieties. Thus, the corresponding quality control methods also need to be different. Currently, limited information is available about the quality control of Gynostemma pentaphyllum from Fujian. A new method based on ultra-high performance liquid chromatography-charged aerosol detection (UHPLC-CAD) was established for the determination of gypenoside XLVI and LVI in Gynostemma pentaphyllum. The major components of Gynostemma pentaphyllum were characterized using UHPLC-quadrupole time-of-flight-mass spectrometry (UHPLC-Q-TOF/MS) combined with UHPLC-CAD. The results revealed gypenoside XLVI, LVI, and their corresponding malonyl-containing acidic saponins as the main components. However, malonylgypenoside XLVI and LVI can easily remove their malonyl group and convert to gypenoside XLVI and LVI during the application of Gynostemma pentaphyllum. In this study, the samples were pretreated using alkali hydrolysis to transform the acid saponins completely, and the final contents of gypenoside XLVI and LVI were determined via UHPLC-CAD. The optimal alkaline hydrolysis, extraction, and liquid chromatography conditions were established. First, the alkaline hydrolysis conditions were optimized. The effects of the volume of ammonia and reaction time on the contents of gypenoside XLVI, LVI, malonylgypenoside XLVI, and LVI were examined. Malonylgypenoside XLVI and LVI could be transformed completely to gypenoside XLVI and LVI by standing for 24 h in an ethanol-water-ammonia (50∶46∶4, v/v/v) mixture. Furthermore, the extraction conditions were optimized. Next, effects of the different solvents, extraction time, and solid-liquid ratio on the extraction rates of gypenoside XLVI and LVI were investigated. The extraction method for Gynostemma pentaphyllum powder using the ethanol-water-ammonia (50∶46∶4, v/v/v) and a solid-liquid ratio of 1∶150 (g∶mL) for 30 min was established. Finally, a prepared test solution was separated on a Waters ACQUITY UPLC BEH C18 chromatographic column (100 mm×2.1 mm, 1.7 µm). Acetonitrile and 0.1% (v/v) formic acid aqueous solution were used as the mobile phases for gradient elution. The flow rate was set to 0.5 mL/min and column temperature was maintained at 40 ℃. The separation was detected using a charged aerosol detector. Results indicated that the logarithm of the mass concentrations of gypenoside XLVI and LVI had a linear relationship with the logarithm of the peak area in the range of 9.94-318.00 µg/mL and 12.78-409.00 µg/mL, respectively. The correlation coefficients (r) were 0.9993 and 0.9995, respectively. The limit of detection (LOD) and the limit of quantification (LOQ) of gypenoside XLVI were 1.58 µg/mL and 6.36 µg/mL, respectively. The LOD and LOQ of gypenoside LVI were 2.05 µg/mL and 8.18 µg/mL, respectively. The relative standard deviations (RSDs) of precision, repeatability, and 24 h stability were less than 2.0% (n=6). The spiked recoveries of gypenoside XLVI were 100.2%-107.2% and the RSD value was 2.4%. The spiked recoveries of gypenoside LVI were 97.9%-104.2% and the RSD value was 2.6%. The results of 16 batches of Gynostemma pentaphyllum samples indicated that the gypenoside XLVI content was 0.57%-2.57%, and gypenoside LVI content was 0.66%-2.99%. Hence, this method has high sensitivity and good reproducibility. Therefore, it can be used for quality research and quality control of Gynostemma pentaphyllum from Fujian.

[Qualitative analysis of Gynostemma longipes for medicinal usage].

The Qinling-Daba Mountains area is the main producing areas of Gynostemma longipes for medicinal usage, and samples of wild whole plants in Pingli, Shaanxi Province and Qingchuan, Sichuan Province were collected. The ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(UHPLC-Q-TOF-MS~E) was used to profile the chemical compositions and analyze the similarities and differences of G. longipes samples in these areas. Based on the accurate molecular weight and fragment information obtained from Q-TOF-MS~E, the structures of the main components were identified by combining with the mass spectra, chromatographic behaviors of reference standards and related literatures. The results showed that the components of wild G. longipes from different places among Qinling-Daba Mountains area were similar. Forty-five chemical components were identified in the whole plant of G. longipes from Pingli, Shaanxi Province, including 43 triterpenoid saponins and 2 flavonoids which contain all main peaks in its fingerprint. The main components are dammarane-type triterpenoid saponins, such asgypenoside ⅩLⅨ, gypenoside A and its malonylated product of glycosyl.

Identification and Differentiation of Polygonum multiflorum Radix and Polygoni multiflori Radix Preaparata through the Quantitative Analysis of Multicomponents by the Single-Marker Method.

The quantitative analysis of multicomponents by the single-marker (QAMS) method was established and the relationship between F value (the ratio of the sum of the contents of emodin-8-O-ß-D-glucopyranoside and physcion-8-O-ß-D-glucopyranoside to the sum of the contents of emodin and physcion) and the steaming time was found to identify and differentiate Polygonum multiflorum Radix and its processed product. Emodin was considered as the control substance, and the correction factors of physcion, emodin-8-O-ß-D-glucopyranoside, and physcion-8-O-ß-D-glucopyranoside were computed. In addition, the contents of the four components were determined. When the F value is greater than or equal to 1.0, the sample was identified as Polygonum multiflorum Radix, and if the F value was between 0.6 and 1.0, the sample of Polygoni multiflori Radix Preaparata was processed incompletely. The F value of the qualified Radix Polygonum multiflorum should be no more than 0.6. However, the influence of different sample injection volumes and the chromatographic columns and instruments used on the durability of the correction factors and RSD ≤3% hindered accurate identification; therefore, a QAMS method using an external standard value with methodological verification was developed. We redefined the "Polygonum multiflorum rules." The method using "Polygonum multiflorum rules" revised after optimization of the determination results was used, as it was accurate and led to convenient operation and low inspection costs, and moreover, the method could differentiate Polygoni multiflori Radix Preaparata and Polygonum multiflorum Radix medicinal samples and precisely identify samples that were different from the completely processed product Polygoni multiflori Radix Preaparata.

Analysis of the Effect of 60Co-γ Irradiation Sterilization Technology on the Chemical Composition of Saffron Using UPLC and UPLC/Q-TOF-MS.

To evaluate the effect of 60Co-γ irradiation sterilization technology on the chemical composition of saffron, we collected 10 batches of saffron samples and treated them with different irradiation doses. UPLC characteristic chromatogram showed that there was no significant effect of irradiation on 13 common peak areas. The results of cluster analysis and principal component analysis showed that there were no differences in the chemical composition in nonirradiated and irradiated samples. UPLC/Q-TOF-MS identified 40 characteristic components of saffron, and the results showed that all of these were detected in the saffron samples both with and without irradiation. Irradiation doses at or below 10 kGy had no significant effect on the chemical components of saffron. This provides a sound basis for the use of 60Co-γ ray irradiation sterilization technology during the preparation of original powder saffron as a medicinal herb, for the effective destruction of mycotoxin contamination.

[Studies on the chemical constituents of Rhodiola dumulosa (II)].

The rhizoma samples of Rhodiola dumulosa were cut into fragments and extracted by 95% EtOH. This extract was successively extracted with Et2O, EtOAc, n-BuOH. Five compounds were obtained from the EtOAc fraction. The structures were identified as beta-sitosterol glucoside (I), chrysophanol-8-O-beta-D-glucopyranoside (II), rhodionin (III), crenuloside (IV), herbacetin-7-O-(3"-beta-D-glucopyranosyl)-alpha-L-rhamnoside (V). Chrysophanol-8-O-beta-D-glucopyranoside and crenuloside of compounds were obtained from this plant for the first time, and chrysophanol-8-O-beta-D-glucopyranoside was obtained from Rhodiola Linn for the first time.

[Studies on the chemical constituents from Rhodiola dumulosa (I)].

The rhizoma samples of Rhodiola dumulosa were cut into fragments and extracted by 95% EtOH. This extract was successively extracted with Et2O, EtOAc, n-BuOH. Six compounds were obtained from the Et2O fraction. The structures of six compounds were identified. The compounds are beta-sitosterol (I), herbacetin-8-methyl ether (II), kaemperol (III), kaemperol-7-O-alpha-L-rhamnoside (IV), beta-sitosterol glucoside (V), herbacetin-7-alpha-L-rhamnoside (VI). beta-sitosterol, herbacetin-8-methyl ether and kaemperol of compounds were obtained from this plant for the first time.