Objective To explore the optimal extraction process of flavonoids from Epimedii folium by deep eutectic solvent method.Methods Using the extraction rate of epimedin A,epimedin B,epimedin C and icariin in Epimedii folium as evaluation indices,the extraction process of flavonoids in Epimedii folium was optimized based on single factor experiments such as solid-liquid ratio,molar ratio,moisture content,extraction temperature,and extraction time,combined with orthogonal experimental design,and the components in the solvent were recovered using macroporous resin.Results The deep eutectic solvent formed by choline chloride and ethanolamine had the best extraction effect.When the molar ratio of choline chloride to ethanolamine was 1:4,the solid-liquid ratio was 0.5 g:25 mL,the water content was 50％,the extraction temperature was 70 ℃,and the extraction time was 15 minutes,the total extraction rate was(20.07±0.43)mg/g,and the AB-8 macroporous resin could recover flavonoids in the eutectic solvent with an average recovery rate of 88.29％.Conclusion Deep eutectic solvents have great potential in the extraction of active ingredients in traditional Chinese medicine due to their simple preparation and higher extraction efficiency compared to traditional solvents.

OBJECTIVE To establish an UPLC-MS/MS method for the determination of the concentrations of cyperenone and α-cyperone in rat plasma and to study the pharmacokinetics. METHODS Gradient elution was carried out on a Phenomennex C18(150 mm×2.0 mm, 3 μm) column with acetonitrile-water as mobile phase. The column temperature was 30 ℃, injection volume was 1 μL, osthenite was used as the internal standard, electrospray ion source and positive ion mode were used. The m/z values of cyperenone, α-cyperone and osthenite were 219.1/135.1, 219.1/111.0 and 245.0/123.0, respectively. The plasma concentrations of cyperenone and α-cyperone were measured, and the main pharmacokinetic parameters were calculated using DAS 2.0 software. RESULTS The linear relationship of cyperenone was good in the range of 10-500 ng·mL-1(r=0.991 0), and the linear relationship of α-cyperone was good in the range of 2.5-300 ng·mL-1(r=0.994 1), RSDs of intra-day precision were less than 9.45%. RSDs of daytime precision were less than 9.09%. The recoveries were greater than 86.79%. After intragastric administration of essential oil extract(20 mg·kg-1) from Cyperus rotundus L. in SD rats. The pharmacokinetic parameters of Cmax, AUC0-∞ and MRT(0-∞) of cyperenone and α-cyperone were (8 862.59±1 106.81)ng·L-1, (7 060.94±774.25)ng·L-1·h, (3.21±0.72)h and (934.69±106.81)ng·L-1, (792.26±74.52)ng·L-1·h, (4.94± 0.82)h, respectively. CONCLUSION The established method can be used for the rapid and accurate determination of the concentration of cyperenone and α-cyperone in plasma, and can be used for the pharmacokinetic study of cyperenone and α-cyperone in rats in vivo.

Objective:To explore the correlation between CCAAT enhancer binding protein beta (CEBPB) expression and clinical characteristics in ccRCC, and to investigate the effect of CEBPB on proliferation and invasion of ccRCC cells.Methods:Between March 2020 to December 2020, the transcriptome and clinical data of 537 ccRCC cases were downloaded from TCGA database, and the correlation of CEBPB expression with clinical characteristics of ccRCC were analyzed. Univariate and multivariate Cox regression analysis were used to determine the effect of CEBPB expression on the prognosis of ccRCC patients. The correlation between CEBPB expression and immunocyte infiltration in ccRCC was investigated via TIMER database. The expression levels of CEBPB mRNA and protein in human renal tubular epithelial cell line HK2 and ccRCC cell lines (Caki-1, ACHN, 786O, 769P and A498) were determined by real-time PCR and western blot, respectively. After transfected with NC siRNA or CEBPB siRNA for 48 h, the proliferation and invasion of ACHN cells and 786O cells were determined by using MTT assay and invasion assay, respectively.Results:TCGA databases analysis revealed that, compared with normal kidney tissue, the expression of CEBPB mRNA in ccRCC was up-regulated by 2.55-fold ( P<0.05). CEBPB expression was positively correlated with age, tumor grade, tumor stage, lymph node metastasis and distant metastasis ( P<0.05). The tumor grade ( HR=1.703, P=0.040), tumor stage( HR=1.773, P=0.026), distant metastasis ( HR=3.080, P<0.001) and the high expression of CEBPB ( HR=1.874, P=0.003) were independent poor prognostic factors for ccRCC patients. The analysis results by using TIMER database showed that CEBPB expression was positively correlated with infiltrating levels of B cells (Rho=0.168), M2 macrophages (Rho=0.373), Tregs (Rho=0.348), neutrophils (Rho=0.194), and natural killer T cell (Rho=0.421) in ccRCC. The expression level of CEBPB mRNA in Caki-1, ACHN, 786O, 769P and A498 cells was (9.43±1.25)-fold, (5.44±0.82)-fold, (4.50±0.52)-fold, (4.88±0.73)-fold and (7.50 ± 1.04)-fold of HK2 cells, respectively. The expression level of CEBPB protein was (6.22±0.45)-fold, (5.84±0.85)-fold, (6.51±0.55)-fold, (6.23±0.62)-fold and (3.84±0.45)-fold of HK2 cells, respectively ( P<0.05). MTT assay showed that the proliferation rates of ACHN cells and 786O cells at 24, 48, 72, 96 h were (98.4±1.7)% and (99.0±1.4)%, (97.8±2.1)% and (98.5±1.5)%, (101.3±1.2)% and (97.6±1.7)%, (97.5±2.0)% and (99.1±1.3)% in NC siRNA group, and (68.8±5.8)% and (79.5±6.2)%, (57.9 ± 6.1)% and (70.8±5.1)%, (50.9±4.6)% and (66.8±4.9)%, (43.2±5.0)% and (60.5±5.3)% in CEBPB siRNA group. Compared with NC siRNA group, the proliferation activity of ACHN cells and 786O cells was significantly inhibited in the CEBPB siRNA group ( P<0.05). Cell invasion assay showed that the invasion activity of ACHN cells and 786O cells were (95.0±5.2)% and (97.3±4.4)% in NC siRNA group, (35.2±5.4)% and (26.7±3.3)% in CEBPB siRNA group, respectively ( P<0.05). Compared with NC siRNA group, the invasion activity of ACHN cells and 786O cells were significantly inhibited in the CEBPB siRNA group ( P<0.05). Conclusions:CEBPB was highly expressed in ccRCC, which was closely related to the prognosis and immunocyte infiltration of ccRCC patients. Silencing the expression of CEBPB significantly inhibited the proliferation and invasion of ccRCC cells

Objective: To determine total phenylethanoid glycoside and acteoside in Plantago Herba to provide reference for evaluating the quality of medicinal materials.Methods: With acteoside as the control sample, a UV visible spectrophotometric method was used to determine total phenylethanoid glycosides in Plantago Herba.An HPLC method was applied to determine acteoside in Plantago Herba , and the conditions were as follows: an ODS2 C 18 (150 mm× 4.6 mm ,5 μm) chromatographic column was used with acetonitrile-0.1% formic acid (13∶87) as the mobile phase at a flow rate of 1.0 ml·min-1 , the detection wavelength was 332nm, the column temperature was 30℃, and the sample volume was 10 μl.Results: The reference solution and the sample solution had the maximum absorption at 332 nm, and the linear relationship was good within the range of 0.003 1-0.155 0 mg·ml-1 (r=0.999 5).The content of total benzene alcohol glycosides in 3 batches of samples was 2.73% , 2.61% and 2.84% , respectively;acteoside over the range of 0.000 6-0.155 0 mg·ml-1 (r=0.999 1) showed a good linear relationship with peak area,the sample recovery was 98.5% and the RSD was 1.6% (n =6), and the acteoside content in 3 batches of samples respectively was 0.54% , 0.51% and 0.56%.Conclusion: The method is simple, accurate and reproducible, and can be used for the determination of total phenylethanoid glycosides and acteoside in Plantago Herba.

Objective:To study the compatibility and stability of coenzyme A for injection, adenosine disodium triphosphate and inosine injection. Methods:By simulating the clinical medication, the three drugs and 5% glucose injection were mixed together. The contents and relative substances of coenzyme A, adenosine disodium triphosphate and inosine were measured by HPLC. The changes in appearance, pH and insoluble particles were observed or tested at ambient temperature. Results:The mixed solution showed no signifi-cant changes in appearance, pH, number of insoluble particles, contents and relative substances of coenzyme A, adenosine disodium triphosphate and inosine in 4 h, while the mixed solution became turbid and the pH, number of insoluble particles and contents of the three drugs showed significant changes after 24-h storage. Conclusion:The mixed solution of coenzyme A for injection, adenosine dis-odium triphosphate and inosine injection in 5% glucose injection should be used up in 4 h at ambient temperature.

Objective To establish a method for determination of phenylethanoid glycoside and acteoside in Plantago Herba. Methods UV-visible spectrophotometric method was used for the determination of the content of phenylethanoid glycosides compounds in Plantago Herba. HPLC method was used for the determination of acteoside in Plantago Herba. Chromatographic column with C18 ODS2 (4.6 mm × 150 mm, 5 μm) was used. Acetonitrile-0.1%formic acid (13:87) was as mobile phase; the flow rate was 1 mL/min; the detection wavelength was 332 nm; the column temperature was 30 ℃; the sample volume was 10 μL. Results The contents of phenylethanoid glycoside in Plantago Herba from different producing areas were among 1.03%–3.47%. Acteoside with peak area over the 0.0062–1.55 mg range showed a good linear relationship; the sample recovery rate was 98.9%, and the RSD was 1.6%. The contents of acteoside in Plantago Herba from different producing areas was among 0.18%–0.56%. Conclusion The method is simple, stable and reproducible, which can be used for the determination of phenylethanoid glycoside and acteoside in Plantago Herba from different producing areas and provide experimental basis for quality control of Plantago Herba.

Objective To establish an HPLC method for the simultaneous determination of quercetin and kaempferol inKaempferia galanga L..Methods ODS2 C18 (5μm, 4.6 mm×150 mm) was used as chromatographic column; methanol-0.4% phosphate (47:53) was the mobile phase; the flow rate was 1 mL/min; column temperature was 30℃; the detection wavelength was 367 nm; the injection volume was 10μL.Results Quercetin showed good linear relationship in the range of 0.016 5–1.65μg (r=0.999 7). The average recovery rate was 96.8%, RSD=2.02%; kaempferol showed good linear relationship in the range of 0.014 6–1.46μg (r=0.999 5). The average recovery rate was 97.3%, RSD=1.77%.Conclusion The method is simple, accurate, and with good reproducibility, which can be used for content determination of quercetin and kaempferol inKaempferia galanga L..

Objective To study the compatible stability of the carbohydrate-electrolyte injection and commonly used vitamin-electrolyte injections. Methods By simulating clinical use of medicines,the carbohydrate-electrolyte injection and various vitamin-electrolyte injections were mixed respectively.The content of sodium acetate was measured by HPLC,and changes in appearance,pH value and insoluble particles of the injections were observed. Results At room temperature,the compatibility solutions showed no significant changes in appearance,pH value,the number of insoluble particles and the content of sodium acetate within 8 h. Conclusion The carbohydrate-electrolyte injection is compatible with commonly used vitamin-electrolyte injections,and the admixtures are stable within 8 h at room temperature.

Objective To optimize the prescription of GA and GB hydrophilic gel matrix tablets; To study the in vitro release mechanism. Methods On the basis of the results of the mono-factor investigation, mixture uniform design was used to optimize the handicraft molding prescription of GA and GB hydrophilic gel matrix tablets. The release mechanism was investigated by the vitro of the GA and GB hydrophilic gelmatrix tablets to accumulate releasing rate to conduct linear fitting. Results The optimized prescription of GA and GB hydrophilic gel matrix tablets was: powder: HPMC: lactose=23:24:53. Conclusion Mixture uniform design can be used to optimize the prescriptions of GA and GB hydrophilic gel matrix tablets, and the results are accurate. The hydrophilic gelmatrix tablets release medicine by non-Fick mechanism, and the medicine release is in accordance with zero-order.

Objective: To establish an HPLC method for the determination of coenzyme A in coenzyme complex for injection. Methods:The content determination was performed on an Intersil ODS-3 column with methanol-pH 6. 5 phosphate buffer solution (10∶90) as the mobile phase. The detection wavelength was 259 nm and the flow rate was 1. 0 ml·min-1 . The column temperature was 30℃ and the injection volume was 20 μl. Results:The linear range of coenzyme A was 1.624-32.482 u·ml-1(r=0.999 9). The average recovery was 102. 36% and RSD was 1. 14%(n=6). Conclusion: The method is simple, accurate and reproducible, and it can be used for the quality control of coenzyme complex for injection.