The isolation of minor components from complex natural product matrices presents a significant challenge in the field of purification science due to their low concentrations and the presence of structurally similar co...The isolation of minor components from complex natural product matrices presents a significant challenge in the field of purification science due to their low concentrations and the presence of structurally similar compounds.This study introduces an optimized twin-column recycling chromatography method for the efficient and simultaneous purification of these elusive constituents.By introducing water at a small flowing rate between the twin columns,a step solvent gradient is created,by which the leading edge of concentration band would migrate at a slower rate than the trailing edge as it flowing from the upstream to downstream column.Hence,the band broadening is counterbalanced,resulting in an enrichment effect for those minor components in separation process.Herein,two target substances,which showed similar peak position in high performance liquid chromatography(HPLC)and did not exceed 1.8%in crude paclitaxel were selected as target compounds for separation.By using the twin-column recycling chromatography with a step solvent gradient,a successful purification was achieved in getting the two with the purity almost 100%.We suggest this method is suitable for the separation of most components in natural produces,which shows higher precision and recovery rate compared with the common lab-operated separation ways for natural products(thin-layer chromatography and prep-HPLC).展开更多
Glypican-3 is a protein encoded by the Glypican-3 gene located on human X chromosome (Xq26), composed of two subunits, a 40 kDa N-terminal subunit, and a 30 kDa C-terminal subunit. Glypican-3 is a currently potential ...Glypican-3 is a protein encoded by the Glypican-3 gene located on human X chromosome (Xq26), composed of two subunits, a 40 kDa N-terminal subunit, and a 30 kDa C-terminal subunit. Glypican-3 is a currently potential target molecule for liver cancer treatments because of its over-expression and growth effects on hepatocellular carcinoma (HCC). This study examined the expression and purification of a C-terminal subunit of Glypican-3 protein (GPC3-Csub) due to its application in both diagnosis and therapy for hepatocellular carcinoma. The gene encoding for GPC3-Csub was successfully cloned into plasmid pET28a fused with an affinity tag composed of six consecutive histidine residues (His-tag). Recombinant protein GPC3-Csub was expressed in Escherichia coli BL21 (DE3) in the condition of adding 3% ethanol with IPTG induction. GPC3-Csub was extracted using repeated freeze-thaw cycles with lysozyme, and inclusion bodies were solubilized by 8M Urea, SDS 10% in pH 12. His-tag fused GPC3-Csub proteins allowed it to be purified by affinity chromatography method using the Nickel-nitrilotriacetic acid (Ni-NTA) column. High expression of GPC3-Csub was confirmed by Coomassie staining and western-blot. GPC3-Csub could be isolated with a Ni-NTA column and have a purity of about 90%.展开更多
In this study, an optimized high performance liquid chromatography-fluorescence detector (HPLC-FL) method for the determination of benzo[a]pyrene in edible oil was established. HPLC was performed with Thermo Fisher Sc...In this study, an optimized high performance liquid chromatography-fluorescence detector (HPLC-FL) method for the determination of benzo[a]pyrene in edible oil was established. HPLC was performed with Thermo Fisher Scientific C18 column (250 mm×4.6 mm, 5 μm) as the chromatographic column and acetonitrile and water as the mobile phase, and the excitation wavelength and emission wavelength of fluorescence detector were 286 and 430 nm, respectively. The response was high, and the linear range was 0.5-10.0 ng/ml. The lowest limit of detection was 0.11 ng/ml, and the average recovery was 92.5%. This method is suitable for quantitative analysis of benzo[a]pyrene content in edible oil.展开更多
The novel pulsed liquid chromatography radionuclide separation method presented here provides a new and promising strategy for the extraction of uranium from seawater.In this study,a new chromatographic separation met...The novel pulsed liquid chromatography radionuclide separation method presented here provides a new and promising strategy for the extraction of uranium from seawater.In this study,a new chromatographic separation method was proposed,and a pulsed nuclide automated separation device was developed,alongside a new chromatographic column.The length of this chromatographic column was 10 m,with an internal warp of 3 mm and a packing size of 1 mm,while the total separation units of the column reached 12,250.The most favorable conditions for the separation of nuclides were then obtained through optimizing the separation conditions of the device:Sample pH in the column=2,sample injection flow rate=5.698 mL/min,chromatographic column heating temperature=60℃.Separation experiments were also carried out for uranium,europium,and sodium ions in mixed solutions;uranium and sodium ions in water samples from the Ganjiang River;and uranium,sodium,and magnesium ions from seawater samples.The separation factors between the different nuclei were then calculated based on the experimental data,and a formula for the separation level was derived.The experimental results showed that the separation factor in the mixed solution of uranium and europium(1:1)was 1.088,while achieving the initial separation of uranium and europium theoretically required a 47-stage separation.Considering the separation factor of 1.50for the uranium and sodium ions in water samples from the Ganjiang River,achieving the initial separation of uranium and sodium ions would have theoretically required at least a 21-stage separation.Furthermore,for the seawater sample separation experiments,the separation factor of uranium and sodium ions was 1.2885;therefore,more than 28 stages of sample separation would be required to achieve uranium extraction from seawater.The novel pulsed liquid chromatography method proposed in this study was innovative in terms of uranium separation and enrichment,while expanding the possibilities of extracting uranium from seawater through chromatography.展开更多
The low temperature coal tar(CT)is taken as the raw material,and the extraction and column chromatography are used for detailed and accurate characterization in this paper.The n-heptane soluble fraction(CT-HS)and inso...The low temperature coal tar(CT)is taken as the raw material,and the extraction and column chromatography are used for detailed and accurate characterization in this paper.The n-heptane soluble fraction(CT-HS)and insoluble fraction(CT-HI)were obtained by n-heptane Soxhlet extraction.The extraction rate of CT-HS reached 92.79%(mass),which indicated that there are few heavy compounds in it.Further,different solvents(methylbenzene,benzene,ethyl acetate,methylbenzene-ethanol)were used to elute CT-HS by chromatographic column to obtain five fractions(saturates,aromatics,heteroatoms,phenolics and resins,named CT-SA,CT-AR,CT-HE,CT-PH,CT-RE,respectively).The yields of CTSA,CT-AR,CT-HE,CT-PH,CT-RE are 42.12%,10.43%,2.19%,9.50%and 6.63%(mass),respectively.Gas chromatography-mass spectrometry analysis of eluting components show that alkanes are the main components in CT,followed by polycyclic aromatics,and the corresponding fractions are CT-SA and CT-AR,respectively.The relative content of aliphatics in CT-SA is 76.93%,and the relative content of aromatics in CT-AR is 75.05%.This separation technology effectively separates and enriches different components in CT,and the activation energy required for the pyrolysis process of a single eluting fraction is lower than that of CT,which is expected to provide an important reference for the separation,analysis and conversion of complex oil products such as coal-oil co-processing products,coal tar and other complex heavy carbon oil products.展开更多
The purpose of this study was to establish a high-performance liquid chromatography (HPLC) method for the simultaneous determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, a...The purpose of this study was to establish a high-performance liquid chromatography (HPLC) method for the simultaneous determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection. The chromatographic method employed was as follows: the column was a Welch Ultimate XB-C18 column (250 mm × 4.6 mm, 10 μm), the mobile phase was a gradient elution of 0.4% formic acid aqueous solution (A) and acetonitrile (B), the detection wavelengths were 280 nm for sodium danshensu, protocatechuic aldehyde, and salvianolic acid B and 326 nm for 4-coumaric acid and rosmarinic acid, the sample volume was 10 μL, the flow rate was 1.0 mL/min, and the column temperature was 35°C. This method can realize the separation and determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid within 50 minutes. The linear relationships of the five peak areas and their concentrations are good (R<sup>2</sup>> 0.9997). The precision RSD values are all less than 1.0%. The reproducibility RSD values are all less than 1.3%. The stability RSD values are all less than 2.2%. The recovery values ranged from 92.4% to 99.4%. This method is simple, accurate, and reproducible. It can be used for the determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection.展开更多
A sensitive, accurate and robust Liquid Chromatography Tandem Mass Spectrometry method has been developed and validated to measure voriconazole trough levels in human plasma. The plasma samples were mixed with flucona...A sensitive, accurate and robust Liquid Chromatography Tandem Mass Spectrometry method has been developed and validated to measure voriconazole trough levels in human plasma. The plasma samples were mixed with fluconazole as an Internal Standard and directed to protein precipitation and drug extraction. An aliquot of 1 μl was injected into the chromatographic system and separated by the Acquity BEH C18 column at a flow rate of 0.30 ml/min in a gradient mobile phase consisting of acetonitrile, Ultrapure water (UPW), methanol and formic acid. Voriconazole was detected by a Triple Quadrupole Detector (TQD) operating on Multiple Reaction Monitoring (MRM) and a positive ion mode Electrospray ionization (ESI) Q1 mass: 350.1 m/z, Q3 mass: 281.1 m/z. Method linearity of the calibration curve (0.10 - 8.00 μg/ml) indicated a correlation coefficient r ≥ 0.99. The intra and inter-assay accuracy was within 85% - 115% and the intra and inter-assay precision was ≤5.76%. Voriconazole recovery percentage was between 97.69 - 119.62%. The method was successively applied in routine voriconazole TDM.展开更多
BACKGROUND Congenital biliary atresia(CBA)is a serious hepatobiliary disease in children with unknown etiology.Its outcome is often liver transplantation or death.Clarifying the etiology of CBA is of great significanc...BACKGROUND Congenital biliary atresia(CBA)is a serious hepatobiliary disease in children with unknown etiology.Its outcome is often liver transplantation or death.Clarifying the etiology of CBA is of great significance for prognosis,treatment,and genetic counseling.CASE SUMMARY A male Chinese infant at an age of 6 mo and 24 d was hospitalized because of"yellow skin for more than 6 mo".Soon after birth,the patient developed jaundice,which then progressively intensified.A"laparoscopic exploration"indicated"biliary atresia".After coming to our hospital,genetic testing suggested a GPC1mutation[loss 1(exons 6-7)].The patient recovered and was discharged after living donor liver transplantation.After discharge,the patient was followed up.The condition was controlled by oral drugs,and the patient’s condition was stable.CONCLUSION CBA is a complex disease with a complex etiology.Clarifying the etiology is of great clinical importance for treatment and prognosis.This case reports CBA caused by a GPC1 mutation,which enriches the genetic etiology of biliary atresia.However,its specific mechanism needs to be confirmed by further research.展开更多
基金supported by the National Natural Science Foundation of China(22078281)。
文摘The isolation of minor components from complex natural product matrices presents a significant challenge in the field of purification science due to their low concentrations and the presence of structurally similar compounds.This study introduces an optimized twin-column recycling chromatography method for the efficient and simultaneous purification of these elusive constituents.By introducing water at a small flowing rate between the twin columns,a step solvent gradient is created,by which the leading edge of concentration band would migrate at a slower rate than the trailing edge as it flowing from the upstream to downstream column.Hence,the band broadening is counterbalanced,resulting in an enrichment effect for those minor components in separation process.Herein,two target substances,which showed similar peak position in high performance liquid chromatography(HPLC)and did not exceed 1.8%in crude paclitaxel were selected as target compounds for separation.By using the twin-column recycling chromatography with a step solvent gradient,a successful purification was achieved in getting the two with the purity almost 100%.We suggest this method is suitable for the separation of most components in natural produces,which shows higher precision and recovery rate compared with the common lab-operated separation ways for natural products(thin-layer chromatography and prep-HPLC).
文摘Glypican-3 is a protein encoded by the Glypican-3 gene located on human X chromosome (Xq26), composed of two subunits, a 40 kDa N-terminal subunit, and a 30 kDa C-terminal subunit. Glypican-3 is a currently potential target molecule for liver cancer treatments because of its over-expression and growth effects on hepatocellular carcinoma (HCC). This study examined the expression and purification of a C-terminal subunit of Glypican-3 protein (GPC3-Csub) due to its application in both diagnosis and therapy for hepatocellular carcinoma. The gene encoding for GPC3-Csub was successfully cloned into plasmid pET28a fused with an affinity tag composed of six consecutive histidine residues (His-tag). Recombinant protein GPC3-Csub was expressed in Escherichia coli BL21 (DE3) in the condition of adding 3% ethanol with IPTG induction. GPC3-Csub was extracted using repeated freeze-thaw cycles with lysozyme, and inclusion bodies were solubilized by 8M Urea, SDS 10% in pH 12. His-tag fused GPC3-Csub proteins allowed it to be purified by affinity chromatography method using the Nickel-nitrilotriacetic acid (Ni-NTA) column. High expression of GPC3-Csub was confirmed by Coomassie staining and western-blot. GPC3-Csub could be isolated with a Ni-NTA column and have a purity of about 90%.
文摘In this study, an optimized high performance liquid chromatography-fluorescence detector (HPLC-FL) method for the determination of benzo[a]pyrene in edible oil was established. HPLC was performed with Thermo Fisher Scientific C18 column (250 mm×4.6 mm, 5 μm) as the chromatographic column and acetonitrile and water as the mobile phase, and the excitation wavelength and emission wavelength of fluorescence detector were 286 and 430 nm, respectively. The response was high, and the linear range was 0.5-10.0 ng/ml. The lowest limit of detection was 0.11 ng/ml, and the average recovery was 92.5%. This method is suitable for quantitative analysis of benzo[a]pyrene content in edible oil.
基金the Natural Science Foundation of Jiangxi Province,China(No.20202BABL203004)the Opening Project of the State Key Laboratory of Nuclear Resources and Environment(East China University of Technology)(No.2022NRE23)the Opening Project of Jiangxi Province Key Laboratory of Polymer Micro/Nano Manufacturing and Devices(No.PMND202101).
文摘The novel pulsed liquid chromatography radionuclide separation method presented here provides a new and promising strategy for the extraction of uranium from seawater.In this study,a new chromatographic separation method was proposed,and a pulsed nuclide automated separation device was developed,alongside a new chromatographic column.The length of this chromatographic column was 10 m,with an internal warp of 3 mm and a packing size of 1 mm,while the total separation units of the column reached 12,250.The most favorable conditions for the separation of nuclides were then obtained through optimizing the separation conditions of the device:Sample pH in the column=2,sample injection flow rate=5.698 mL/min,chromatographic column heating temperature=60℃.Separation experiments were also carried out for uranium,europium,and sodium ions in mixed solutions;uranium and sodium ions in water samples from the Ganjiang River;and uranium,sodium,and magnesium ions from seawater samples.The separation factors between the different nuclei were then calculated based on the experimental data,and a formula for the separation level was derived.The experimental results showed that the separation factor in the mixed solution of uranium and europium(1:1)was 1.088,while achieving the initial separation of uranium and europium theoretically required a 47-stage separation.Considering the separation factor of 1.50for the uranium and sodium ions in water samples from the Ganjiang River,achieving the initial separation of uranium and sodium ions would have theoretically required at least a 21-stage separation.Furthermore,for the seawater sample separation experiments,the separation factor of uranium and sodium ions was 1.2885;therefore,more than 28 stages of sample separation would be required to achieve uranium extraction from seawater.The novel pulsed liquid chromatography method proposed in this study was innovative in terms of uranium separation and enrichment,while expanding the possibilities of extracting uranium from seawater through chromatography.
基金financed by the project supported by the National Natural Science Foundation of China(22078266,21908180,22178289,22278338)the Key Research and Development Program of Shaanxi(2020ZDLGY11-02,2021GY-136)the Special Fund for High-level Scholars of China(XJ21B10)。
文摘The low temperature coal tar(CT)is taken as the raw material,and the extraction and column chromatography are used for detailed and accurate characterization in this paper.The n-heptane soluble fraction(CT-HS)and insoluble fraction(CT-HI)were obtained by n-heptane Soxhlet extraction.The extraction rate of CT-HS reached 92.79%(mass),which indicated that there are few heavy compounds in it.Further,different solvents(methylbenzene,benzene,ethyl acetate,methylbenzene-ethanol)were used to elute CT-HS by chromatographic column to obtain five fractions(saturates,aromatics,heteroatoms,phenolics and resins,named CT-SA,CT-AR,CT-HE,CT-PH,CT-RE,respectively).The yields of CTSA,CT-AR,CT-HE,CT-PH,CT-RE are 42.12%,10.43%,2.19%,9.50%and 6.63%(mass),respectively.Gas chromatography-mass spectrometry analysis of eluting components show that alkanes are the main components in CT,followed by polycyclic aromatics,and the corresponding fractions are CT-SA and CT-AR,respectively.The relative content of aliphatics in CT-SA is 76.93%,and the relative content of aromatics in CT-AR is 75.05%.This separation technology effectively separates and enriches different components in CT,and the activation energy required for the pyrolysis process of a single eluting fraction is lower than that of CT,which is expected to provide an important reference for the separation,analysis and conversion of complex oil products such as coal-oil co-processing products,coal tar and other complex heavy carbon oil products.
文摘The purpose of this study was to establish a high-performance liquid chromatography (HPLC) method for the simultaneous determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection. The chromatographic method employed was as follows: the column was a Welch Ultimate XB-C18 column (250 mm × 4.6 mm, 10 μm), the mobile phase was a gradient elution of 0.4% formic acid aqueous solution (A) and acetonitrile (B), the detection wavelengths were 280 nm for sodium danshensu, protocatechuic aldehyde, and salvianolic acid B and 326 nm for 4-coumaric acid and rosmarinic acid, the sample volume was 10 μL, the flow rate was 1.0 mL/min, and the column temperature was 35°C. This method can realize the separation and determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid within 50 minutes. The linear relationships of the five peak areas and their concentrations are good (R<sup>2</sup>> 0.9997). The precision RSD values are all less than 1.0%. The reproducibility RSD values are all less than 1.3%. The stability RSD values are all less than 2.2%. The recovery values ranged from 92.4% to 99.4%. This method is simple, accurate, and reproducible. It can be used for the determination of sodium danshensu, protocatechuic aldehyde, rosmarinic acid, salvianolic acid B, and 4-coumaric acid in Danhong injection.
文摘A sensitive, accurate and robust Liquid Chromatography Tandem Mass Spectrometry method has been developed and validated to measure voriconazole trough levels in human plasma. The plasma samples were mixed with fluconazole as an Internal Standard and directed to protein precipitation and drug extraction. An aliquot of 1 μl was injected into the chromatographic system and separated by the Acquity BEH C18 column at a flow rate of 0.30 ml/min in a gradient mobile phase consisting of acetonitrile, Ultrapure water (UPW), methanol and formic acid. Voriconazole was detected by a Triple Quadrupole Detector (TQD) operating on Multiple Reaction Monitoring (MRM) and a positive ion mode Electrospray ionization (ESI) Q1 mass: 350.1 m/z, Q3 mass: 281.1 m/z. Method linearity of the calibration curve (0.10 - 8.00 μg/ml) indicated a correlation coefficient r ≥ 0.99. The intra and inter-assay accuracy was within 85% - 115% and the intra and inter-assay precision was ≤5.76%. Voriconazole recovery percentage was between 97.69 - 119.62%. The method was successively applied in routine voriconazole TDM.
文摘BACKGROUND Congenital biliary atresia(CBA)is a serious hepatobiliary disease in children with unknown etiology.Its outcome is often liver transplantation or death.Clarifying the etiology of CBA is of great significance for prognosis,treatment,and genetic counseling.CASE SUMMARY A male Chinese infant at an age of 6 mo and 24 d was hospitalized because of"yellow skin for more than 6 mo".Soon after birth,the patient developed jaundice,which then progressively intensified.A"laparoscopic exploration"indicated"biliary atresia".After coming to our hospital,genetic testing suggested a GPC1mutation[loss 1(exons 6-7)].The patient recovered and was discharged after living donor liver transplantation.After discharge,the patient was followed up.The condition was controlled by oral drugs,and the patient’s condition was stable.CONCLUSION CBA is a complex disease with a complex etiology.Clarifying the etiology is of great clinical importance for treatment and prognosis.This case reports CBA caused by a GPC1 mutation,which enriches the genetic etiology of biliary atresia.However,its specific mechanism needs to be confirmed by further research.
