Structured Review

Shimadzu Corporation reverse phase hplc
CTSH processing of mature BMP-4 protein. (A) Human cell lines were treated with 5 μM CTSHi for 24 h and then cell lysates were analyzed using Western blot. Proteins (50 μg) from cell lysates were separated on <t>SDS-PAGE</t> and transferred on to PVDF membrane. BMP-4 was detected with anti-BMP-4 N16 antibody (Santa Cruz) and then with secondary antibody labelled with HRP. The molecular mass in kDa is indicated on the left hand side of the blots. Molecular mass of the mature BMP-4 is detected to be approximately 18 kDa. (B) Using Western blot we analyzed the products of the reaction between mature human recombinant BMP-4 and nCTSH. Mature human recombinant BMP-4 was incubated for 1.5 h at 37ºC in CTSH activity buffer (lane 1), with 60 ng of nCTSH in CTSH activity buffer (lane 2) and 60 ng of nCTSH in CTSH activity buffer that was pre-treated for 10 min with 10 μM CTSHi (lane 3). Mature human recombinant BMP-4 has a molecular mass of 13 kDa. (C) Using reverse phase <t>HPLC</t> we analyzed CTSH activity buffer (buffer), mature human recombinant BMP-4 in CTSH activity buffer (BMP-4) and the products of the reaction between mature human recombinant BMP-4 and nCTSH in CTSH activity buffer (BMP-4 + nCTSH). BMP-4 was eluted in the fraction around 17.0 min.
Reverse Phase Hplc, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 92/100, based on 137 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Images

1) Product Images from "Cathepsin H indirectly regulates morphogenetic protein-4 (BMP-4) in various human cell lines"

Article Title: Cathepsin H indirectly regulates morphogenetic protein-4 (BMP-4) in various human cell lines

Journal: Radiology and Oncology

doi: 10.2478/v10019-011-0034-3

CTSH processing of mature BMP-4 protein. (A) Human cell lines were treated with 5 μM CTSHi for 24 h and then cell lysates were analyzed using Western blot. Proteins (50 μg) from cell lysates were separated on SDS-PAGE and transferred on to PVDF membrane. BMP-4 was detected with anti-BMP-4 N16 antibody (Santa Cruz) and then with secondary antibody labelled with HRP. The molecular mass in kDa is indicated on the left hand side of the blots. Molecular mass of the mature BMP-4 is detected to be approximately 18 kDa. (B) Using Western blot we analyzed the products of the reaction between mature human recombinant BMP-4 and nCTSH. Mature human recombinant BMP-4 was incubated for 1.5 h at 37ºC in CTSH activity buffer (lane 1), with 60 ng of nCTSH in CTSH activity buffer (lane 2) and 60 ng of nCTSH in CTSH activity buffer that was pre-treated for 10 min with 10 μM CTSHi (lane 3). Mature human recombinant BMP-4 has a molecular mass of 13 kDa. (C) Using reverse phase HPLC we analyzed CTSH activity buffer (buffer), mature human recombinant BMP-4 in CTSH activity buffer (BMP-4) and the products of the reaction between mature human recombinant BMP-4 and nCTSH in CTSH activity buffer (BMP-4 + nCTSH). BMP-4 was eluted in the fraction around 17.0 min.
Figure Legend Snippet: CTSH processing of mature BMP-4 protein. (A) Human cell lines were treated with 5 μM CTSHi for 24 h and then cell lysates were analyzed using Western blot. Proteins (50 μg) from cell lysates were separated on SDS-PAGE and transferred on to PVDF membrane. BMP-4 was detected with anti-BMP-4 N16 antibody (Santa Cruz) and then with secondary antibody labelled with HRP. The molecular mass in kDa is indicated on the left hand side of the blots. Molecular mass of the mature BMP-4 is detected to be approximately 18 kDa. (B) Using Western blot we analyzed the products of the reaction between mature human recombinant BMP-4 and nCTSH. Mature human recombinant BMP-4 was incubated for 1.5 h at 37ºC in CTSH activity buffer (lane 1), with 60 ng of nCTSH in CTSH activity buffer (lane 2) and 60 ng of nCTSH in CTSH activity buffer that was pre-treated for 10 min with 10 μM CTSHi (lane 3). Mature human recombinant BMP-4 has a molecular mass of 13 kDa. (C) Using reverse phase HPLC we analyzed CTSH activity buffer (buffer), mature human recombinant BMP-4 in CTSH activity buffer (BMP-4) and the products of the reaction between mature human recombinant BMP-4 and nCTSH in CTSH activity buffer (BMP-4 + nCTSH). BMP-4 was eluted in the fraction around 17.0 min.

Techniques Used: Western Blot, SDS Page, Recombinant, Incubation, Activity Assay, High Performance Liquid Chromatography

2) Product Images from "Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis"

Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis

Journal: BMC Proceedings

doi: 10.1186/1753-6561-3-S6-S5

The radioactive signal from oligopeptides being modified with radioactive acetyl CoA . The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with [1- 14 C] acetyl CoA (final 300 μM with specific activity 11.2 mCi/mmol) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC that had been connected to a radioactivity flow detector after the absorbance detector. A clear time dependent increase in the radioactivity signal is observed, verifying that the eluted oligopeptides are labelled with radioactive acetyl groups.
Figure Legend Snippet: The radioactive signal from oligopeptides being modified with radioactive acetyl CoA . The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with [1- 14 C] acetyl CoA (final 300 μM with specific activity 11.2 mCi/mmol) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC that had been connected to a radioactivity flow detector after the absorbance detector. A clear time dependent increase in the radioactivity signal is observed, verifying that the eluted oligopeptides are labelled with radioactive acetyl groups.

Techniques Used: Modification, Incubation, Activity Assay, Purification, High Performance Liquid Chromatography, Radioactivity, Flow Cytometry

MBP-hNaa30p specificity constants (V/K) for selected oligopeptides based on detection of acetylated oligopeptides (215 nm) . Purified MBP-hNaa30p (80 nM) was incubated with selected oligopeptides and acetyl CoA (300 μM) in acetylation buffer for 30 minutes at 37°C. The acetylation kinetics was analysed by reverse phase HPLC. V/K is the V max /K m (oligopeptides) . Error bars indicate S.D. Experiments are performed in triplicates.
Figure Legend Snippet: MBP-hNaa30p specificity constants (V/K) for selected oligopeptides based on detection of acetylated oligopeptides (215 nm) . Purified MBP-hNaa30p (80 nM) was incubated with selected oligopeptides and acetyl CoA (300 μM) in acetylation buffer for 30 minutes at 37°C. The acetylation kinetics was analysed by reverse phase HPLC. V/K is the V max /K m (oligopeptides) . Error bars indicate S.D. Experiments are performed in triplicates.

Techniques Used: Purification, Incubation, High Performance Liquid Chromatography

Reverse phase HPLC absorbance profile at 215 nm for the separation of acetylated and non-acetylated peptides . A ; The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with acetyl CoA (300 μM) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC. The resulting absorbance profile at 215 nm indicate good separation of unacetylated ('a') and acetylated oligopeptides ('b'). B ; An expanded version of the absorbance profile for the formation of acetylated oligopeptide. A clear time dependent increase in the absorption signal is observed.
Figure Legend Snippet: Reverse phase HPLC absorbance profile at 215 nm for the separation of acetylated and non-acetylated peptides . A ; The oligopeptide 1 MLGTE-RRR 24 (200 μM) was incubated with acetyl CoA (300 μM) and purified MBP-hNaa30p (80 nM) in acetylation buffer for 60 minutes at 37°C. Samples were collected at indicated time points and analysed with reverse phase HPLC. The resulting absorbance profile at 215 nm indicate good separation of unacetylated ('a') and acetylated oligopeptides ('b'). B ; An expanded version of the absorbance profile for the formation of acetylated oligopeptide. A clear time dependent increase in the absorption signal is observed.

Techniques Used: High Performance Liquid Chromatography, Incubation, Purification

3) Product Images from "Black ginger extract increases physical fitness performance and muscular endurance by improving inflammation and energy metabolism"

Article Title: Black ginger extract increases physical fitness performance and muscular endurance by improving inflammation and energy metabolism

Journal: Heliyon

doi: 10.1016/j.heliyon.2016.e00115

HPLC chromatogram of KPE for determination of PMFs. PMFs in KPE were separated on a Develosil RPAQUEOUS-AR-5 column. The chromatographic mobile phase consisted of A (acetonitrile: water: acetic acid = 35: 62.5: 2.5, v/v) and B (acetonitrile: acetic acid = 97.5: 2.5, v/v). The gradient program was set as 0–20 min (solvent A: 99–1%). Use of 263 nm as a selective wavelength allowed identification of the 8 known PMFs. Peak 1 , 5-hydroxy-3,7-dimethoxyflavone; 2 , 5-hydroxy-7-methoxyflavone; 3 , 5-hydroxy-3,7,4'-trimethoxyflavone; 4 , 5-hydroxy-3,7,3',4'-tetramethoxyflavone; 5 , 3,5,7,3',4'-pentamethoxyflavone; 6 , 5,7,4'-trimethoxyflavone; 7 , 3,5,7,4'-tetramethoxyflavone; 8 , 5,7-dimethoxyflavone.
Figure Legend Snippet: HPLC chromatogram of KPE for determination of PMFs. PMFs in KPE were separated on a Develosil RPAQUEOUS-AR-5 column. The chromatographic mobile phase consisted of A (acetonitrile: water: acetic acid = 35: 62.5: 2.5, v/v) and B (acetonitrile: acetic acid = 97.5: 2.5, v/v). The gradient program was set as 0–20 min (solvent A: 99–1%). Use of 263 nm as a selective wavelength allowed identification of the 8 known PMFs. Peak 1 , 5-hydroxy-3,7-dimethoxyflavone; 2 , 5-hydroxy-7-methoxyflavone; 3 , 5-hydroxy-3,7,4'-trimethoxyflavone; 4 , 5-hydroxy-3,7,3',4'-tetramethoxyflavone; 5 , 3,5,7,3',4'-pentamethoxyflavone; 6 , 5,7,4'-trimethoxyflavone; 7 , 3,5,7,4'-tetramethoxyflavone; 8 , 5,7-dimethoxyflavone.

Techniques Used: High Performance Liquid Chromatography

Related Articles

High Performance Liquid Chromatography:

Article Title: The structure and function of a soybean ?-glucan-elicitor-binding protein
Article Snippet: .. The digested peptides were chromatographed by reverse-phase HPLC, and the amino acid sequences were determined with a gas-phase sequencer (model PSQ-2, Shimadzu) ( , ). .. Protein was determined by DC protein assay kit (Bio-Rad), and purified GEBP was estimated by the density of the silver-stained SDS/PAGE band.

Article Title: An aqueous extract of Ammi visnaga fruits and its constituents khellin and visnagin prevent cell damage caused by oxalate in renal epithelial cells
Article Snippet: .. The samples were analyzed using reverse-phase HPLC with diode array detector Shimadzu VP series HPLC system (Kyoto, Japan) with diode array detection. ..

Article Title: Development of dual casein kinase 1δ/1ε (CK1δ/ε) inhibitors for treatment of breast cancer
Article Snippet: .. Some of the final products were isolated by reverse-phase HPLC using Shimadzu Prep LC system with photodiode array detector, Waters SunFire C18 OBD Prep Column, 100Å, 10 μm, 30 mm × 250 mm. .. Compounds were eluted using a gradient elution of 90/10 to 0/100 A/B over 10 min at a flow rate of 50.0 mL/min, where solvent A was water (+0.1 % TFA) and solvent B was acetonitrile/methanol (1:1).

Article Title: Black ginger extract increases physical fitness performance and muscular endurance by improving inflammation and energy metabolism
Article Snippet: .. The contents of all PMFs in powdered KPE were determined by reverse-phase HPLC using a Prominence HPLC system (Shimadzu, Kyoto, Japan) equipped with a photodiode array detector (Model SPD-M20A) and a Develosil RPAQUEOUS-AR-5 column (4.6 × 150 mm, 5 μm particle size, Nomura Chemical Co., Ltd., Japan). ..

Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis
Article Snippet: .. Separation of non acetylated and acetylated oligopeptides using reverse phase HPLC The acetylation activity was analysed by a reverse phase HPLC system, consisting of a LC-20AB solvent delivery module, an SPD-M20A photodiode array detector and a SIL-20AC autosampler (Shimadzu Prominence), and a 250 mm × 3 mm Nucleosil C18 HD column (Macherey-Nagel) reverse phase HPLC column. .. In addition, a radioactivity flow detector (LB 509 – Berthold) and a peristaltic pump were connected down stream of the HPLC absorbance detector (Figure ).

Article Title: Activation of Human Peripheral Basophils in Response to High IgE Antibody Concentrations without Antigens
Article Snippet: .. Reverse-phase HPLC was sourced from Shimadzu (Kyoto, Japan). .. Tetramethylrodamin B isothiocyanate (TRITC)-phalloidin and adenosine were sourced from Sigma-Aldrich Japan (Tokyo, Japan).

Article Title: Cathepsin H indirectly regulates morphogenetic protein-4 (BMP-4) in various human cell lines
Article Snippet: .. Samples were analyzed using 12% SDS-PAGE followed by Western blot or reverse-phase HPLC (Shimadzu Coorporation, Japan) using a Discovery BIO Wide Pore C5 column (Sigma, St.Louis, MO, USA) with UV-VIS detector. .. Statistical analysis SPSS PC software (Release 13.0) was used for statistical analysis.

Article Title: Transcriptional analysis of Amorphotheca resinae ZN1 on biological degradation of furfural and 5-hydroxymethylfurfural derived from lignocellulose pretreatment
Article Snippet: .. Analytical methods Residual glucose in the medium was monitored with a SBA-40D glucose analyzer (Shandong Academy of Sciences, Jinan, China).Furan compounds were analyzed using reverse-phase HPLC (LC-20AT, SPD-20A UV detector, Shimadzu, Kyoto, Japan) with a YMC-Pack ODS-A column (YMC, Tokyo, Japan) at the column temperature of 35 °C [ ]. ..

Isolation:

Article Title: Development of dual casein kinase 1δ/1ε (CK1δ/ε) inhibitors for treatment of breast cancer
Article Snippet: .. Some of the final products were isolated by reverse-phase HPLC using Shimadzu Prep LC system with photodiode array detector, Waters SunFire C18 OBD Prep Column, 100Å, 10 μm, 30 mm × 250 mm. .. Compounds were eluted using a gradient elution of 90/10 to 0/100 A/B over 10 min at a flow rate of 50.0 mL/min, where solvent A was water (+0.1 % TFA) and solvent B was acetonitrile/methanol (1:1).

Activity Assay:

Article Title: Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis
Article Snippet: .. Separation of non acetylated and acetylated oligopeptides using reverse phase HPLC The acetylation activity was analysed by a reverse phase HPLC system, consisting of a LC-20AB solvent delivery module, an SPD-M20A photodiode array detector and a SIL-20AC autosampler (Shimadzu Prominence), and a 250 mm × 3 mm Nucleosil C18 HD column (Macherey-Nagel) reverse phase HPLC column. .. In addition, a radioactivity flow detector (LB 509 – Berthold) and a peristaltic pump were connected down stream of the HPLC absorbance detector (Figure ).

SDS Page:

Article Title: Cathepsin H indirectly regulates morphogenetic protein-4 (BMP-4) in various human cell lines
Article Snippet: .. Samples were analyzed using 12% SDS-PAGE followed by Western blot or reverse-phase HPLC (Shimadzu Coorporation, Japan) using a Discovery BIO Wide Pore C5 column (Sigma, St.Louis, MO, USA) with UV-VIS detector. .. Statistical analysis SPSS PC software (Release 13.0) was used for statistical analysis.

Western Blot:

Article Title: Cathepsin H indirectly regulates morphogenetic protein-4 (BMP-4) in various human cell lines
Article Snippet: .. Samples were analyzed using 12% SDS-PAGE followed by Western blot or reverse-phase HPLC (Shimadzu Coorporation, Japan) using a Discovery BIO Wide Pore C5 column (Sigma, St.Louis, MO, USA) with UV-VIS detector. .. Statistical analysis SPSS PC software (Release 13.0) was used for statistical analysis.

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    Shimadzu Corporation reversed phase hplc determinations
    <t>HPLC</t> chromatograms obtained with method B and 250-fold SPE preconcentration for the aerated natural samples γ-irradiated with different doses. a Chromatograms of Vistula River water spiked with 10 μg/L concentration of each analyte, obtained for sample prior to the irradiation ( a ), after γ-irradiation with dose 100 Gy ( b ), and 250 Gy ( c ). b Chromatograms recorded in the same conditions for sample of wastewater from hospital spiked with 10 μg/L each analyte prior to γ-irradiation ( a ), after irradiation with 100 ( b ), 250 ( c ), and 500 ( d ) Gy dose. In both chromatograms: 1-carbamazepine, 2-bisphenol A, <t>3-DCF,</t> 4-ibuprofen
    Reversed Phase Hplc Determinations, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 88/100, based on 104 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reversed phase hplc determinations/product/Shimadzu Corporation
    Average 88 stars, based on 104 article reviews
    Price from $9.99 to $1999.99
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    93
    Shimadzu Corporation hplc system
    Biophysical Characterization and Stability of Gcg-XTEN. Gcg-XTEN was produced recombinantly in E. coli and purified to homogeneity using three column steps (see methods ). (A) SDS-PAGE analysis of the purified protein product (lane 2). Molecular weight markers are shown in lane 1 with relevant size markers labeled at the left. Note that the true molecular weight of the molecule is 16305 daltons (confirmed by mass spectrometry; not shown). Slow migration in SDS-PAGE relative to globular protein standards is typical of XTEN fusion proteins due to differences in primary amino acid composition. (B) Glucagon receptor (GcgR) Ca 2+ -flux assay comparing the efficacy of Gcg-XTEN to unmodified glucagon. Calculated EC50 values for each curve fit are shown. (C) Reverse phase <t>C18</t> <t>HPLC</t> analysis and (D) Size exclusion chromatography HPLC analysis of the purified Gcg-XTEN construct at the time of production. (E) Reverse phase C18 HPLC analysis and (F) Size exclusion chromatography HPLC analysis of Gcg-XTEN after 6 months storage at either −80°C (black), 2–8°C (blue), or 25°C (red). Note the scale is expanded in panel E to better illustrate the appearance of minor peaks at 25°C.
    Hplc System, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 93/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hplc system/product/Shimadzu Corporation
    Average 93 stars, based on 13 article reviews
    Price from $9.99 to $1999.99
    hplc system - by Bioz Stars, 2020-11
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    88
    Shimadzu Corporation reverse phase hplc chemical fingerprint analysis
    Characterization of <t>BHJRST.</t> (a) Confirmation of the identity of the ingredients. The macroscopic and microscopic appearance of the 5 ingredients used to prepare BHJRST was examined. (b) <t>HPLC</t> chromatograms of water extracts of BHJRST and three of the five single ingredients. Three major peaks were identified in the classical BHJRST formula (top panel) by comparison of their retention times with those of peaks in extracts of the single ingredients Rhizoma Anemarrhenae (Zhi-Mu), Ginseng Radix (Ren-Shen), or Radix Glycyrrhizae Preparata (Zhi-Gan-Cao).
    Reverse Phase Hplc Chemical Fingerprint Analysis, supplied by Shimadzu Corporation, used in various techniques. Bioz Stars score: 88/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/reverse phase hplc chemical fingerprint analysis/product/Shimadzu Corporation
    Average 88 stars, based on 1 article reviews
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    HPLC chromatograms obtained with method B and 250-fold SPE preconcentration for the aerated natural samples γ-irradiated with different doses. a Chromatograms of Vistula River water spiked with 10 μg/L concentration of each analyte, obtained for sample prior to the irradiation ( a ), after γ-irradiation with dose 100 Gy ( b ), and 250 Gy ( c ). b Chromatograms recorded in the same conditions for sample of wastewater from hospital spiked with 10 μg/L each analyte prior to γ-irradiation ( a ), after irradiation with 100 ( b ), 250 ( c ), and 500 ( d ) Gy dose. In both chromatograms: 1-carbamazepine, 2-bisphenol A, 3-DCF, 4-ibuprofen

    Journal: Environmental Science and Pollution Research International

    Article Title: Analytical, toxicological and kinetic investigation of decomposition of the drug diclofenac in waters and wastes using gamma radiation

    doi: 10.1007/s11356-015-5236-6

    Figure Lengend Snippet: HPLC chromatograms obtained with method B and 250-fold SPE preconcentration for the aerated natural samples γ-irradiated with different doses. a Chromatograms of Vistula River water spiked with 10 μg/L concentration of each analyte, obtained for sample prior to the irradiation ( a ), after γ-irradiation with dose 100 Gy ( b ), and 250 Gy ( c ). b Chromatograms recorded in the same conditions for sample of wastewater from hospital spiked with 10 μg/L each analyte prior to γ-irradiation ( a ), after irradiation with 100 ( b ), 250 ( c ), and 500 ( d ) Gy dose. In both chromatograms: 1-carbamazepine, 2-bisphenol A, 3-DCF, 4-ibuprofen

    Article Snippet: Analytical procedures The reversed-phase HPLC determinations of DCF were carried out using Shimadzu LC-10 AT chromatograph with the diode array UV–Vis detector model SPD-M10A VP.

    Techniques: High Performance Liquid Chromatography, Irradiation, Concentration Assay

    Chromatographic examination of the effect of selected scavengers of radicals on the a yield of DCF radiolytic decomposition and b on release of chloride in function of different absorbed dose. a Examined using the HPLC-UV method A. b Examined using the suppressed ion-chromatography with the conductivity detection. Gamma-irradiated aqueous solution of DCF 50 mg/L were saturated with air

    Journal: Environmental Science and Pollution Research International

    Article Title: Analytical, toxicological and kinetic investigation of decomposition of the drug diclofenac in waters and wastes using gamma radiation

    doi: 10.1007/s11356-015-5236-6

    Figure Lengend Snippet: Chromatographic examination of the effect of selected scavengers of radicals on the a yield of DCF radiolytic decomposition and b on release of chloride in function of different absorbed dose. a Examined using the HPLC-UV method A. b Examined using the suppressed ion-chromatography with the conductivity detection. Gamma-irradiated aqueous solution of DCF 50 mg/L were saturated with air

    Article Snippet: Analytical procedures The reversed-phase HPLC determinations of DCF were carried out using Shimadzu LC-10 AT chromatograph with the diode array UV–Vis detector model SPD-M10A VP.

    Techniques: High Performance Liquid Chromatography, Ion Chromatography, Irradiation

    Crude RP-HPLC analytical chromatograms at 214 nm of 3 (mass of 961.6) in a gradient from 10% to 90% MeCN or MeOH in water containing 0.1 % trifluoroacetic acid at a flow rate of 1.5 mL/min over 35 minutes (5 to 20 minutes are shown) using an analytical Vydac C18 column (Vydac 218TP104). (A) Analytical HPLC trace in MeCN of crude peptide 3 after a three hour cleavage which shows only one major peak. A major impurity peak (mass of 685.4) is masked in this chromatogram. (B) Analytical HPLC trace in MeOH of crude peptide 3 after a three hour cleavage which identifies both the desired product and an impurity peak masked in MeCN chromatogram. (C) Co-injection of crude 3 from three hour cleavage with purified 1 (mass of 685.4) increases the intensity of the impurity peak demonstrating similar retention times. (D) A shorter cleavage time of 1.5 hours diminishes degradation product giving better crude peptide purity.

    Journal: Journal of medicinal chemistry

    Article Title: An in vitro and in vivo investigation of bivalent ligands that display preferential binding and functional activity for different melanocortin receptor homodimers

    doi: 10.1021/acs.jmedchem.5b01894

    Figure Lengend Snippet: Crude RP-HPLC analytical chromatograms at 214 nm of 3 (mass of 961.6) in a gradient from 10% to 90% MeCN or MeOH in water containing 0.1 % trifluoroacetic acid at a flow rate of 1.5 mL/min over 35 minutes (5 to 20 minutes are shown) using an analytical Vydac C18 column (Vydac 218TP104). (A) Analytical HPLC trace in MeCN of crude peptide 3 after a three hour cleavage which shows only one major peak. A major impurity peak (mass of 685.4) is masked in this chromatogram. (B) Analytical HPLC trace in MeOH of crude peptide 3 after a three hour cleavage which identifies both the desired product and an impurity peak masked in MeCN chromatogram. (C) Co-injection of crude 3 from three hour cleavage with purified 1 (mass of 685.4) increases the intensity of the impurity peak demonstrating similar retention times. (D) A shorter cleavage time of 1.5 hours diminishes degradation product giving better crude peptide purity.

    Article Snippet: A purity of greater than 95% was confirmed by RP-HPLC in two diverse solvent systems (10% MeCN in 0.1 % TFA/water and a gradient to 90% MeCN over 35 min; and 10% MeOH in 0.1 % TFA/water and a gradient to 90% MeOH over 35 minutes).

    Techniques: High Performance Liquid Chromatography, Flow Cytometry, Injection, Purification

    Biophysical Characterization and Stability of Gcg-XTEN. Gcg-XTEN was produced recombinantly in E. coli and purified to homogeneity using three column steps (see methods ). (A) SDS-PAGE analysis of the purified protein product (lane 2). Molecular weight markers are shown in lane 1 with relevant size markers labeled at the left. Note that the true molecular weight of the molecule is 16305 daltons (confirmed by mass spectrometry; not shown). Slow migration in SDS-PAGE relative to globular protein standards is typical of XTEN fusion proteins due to differences in primary amino acid composition. (B) Glucagon receptor (GcgR) Ca 2+ -flux assay comparing the efficacy of Gcg-XTEN to unmodified glucagon. Calculated EC50 values for each curve fit are shown. (C) Reverse phase C18 HPLC analysis and (D) Size exclusion chromatography HPLC analysis of the purified Gcg-XTEN construct at the time of production. (E) Reverse phase C18 HPLC analysis and (F) Size exclusion chromatography HPLC analysis of Gcg-XTEN after 6 months storage at either −80°C (black), 2–8°C (blue), or 25°C (red). Note the scale is expanded in panel E to better illustrate the appearance of minor peaks at 25°C.

    Journal: PLoS ONE

    Article Title: Gcg-XTEN: An Improved Glucagon Capable of Preventing Hypoglycemia without Increasing Baseline Blood Glucose

    doi: 10.1371/journal.pone.0010175

    Figure Lengend Snippet: Biophysical Characterization and Stability of Gcg-XTEN. Gcg-XTEN was produced recombinantly in E. coli and purified to homogeneity using three column steps (see methods ). (A) SDS-PAGE analysis of the purified protein product (lane 2). Molecular weight markers are shown in lane 1 with relevant size markers labeled at the left. Note that the true molecular weight of the molecule is 16305 daltons (confirmed by mass spectrometry; not shown). Slow migration in SDS-PAGE relative to globular protein standards is typical of XTEN fusion proteins due to differences in primary amino acid composition. (B) Glucagon receptor (GcgR) Ca 2+ -flux assay comparing the efficacy of Gcg-XTEN to unmodified glucagon. Calculated EC50 values for each curve fit are shown. (C) Reverse phase C18 HPLC analysis and (D) Size exclusion chromatography HPLC analysis of the purified Gcg-XTEN construct at the time of production. (E) Reverse phase C18 HPLC analysis and (F) Size exclusion chromatography HPLC analysis of Gcg-XTEN after 6 months storage at either −80°C (black), 2–8°C (blue), or 25°C (red). Note the scale is expanded in panel E to better illustrate the appearance of minor peaks at 25°C.

    Article Snippet: Reverse-Phase Chromatography Reverse phase C18 chromatography (RPC18) was performed using a Phenomenex Gemini 5 µm C18, 100 Å, 4.6×100 mm column (Phenomenex) connected to an HPLC system equipped with an autosampler and UV/VIS detector (Shimadzu).

    Techniques: Produced, Purification, SDS Page, Molecular Weight, Labeling, Mass Spectrometry, Migration, Flux Assay, High Performance Liquid Chromatography, Size-exclusion Chromatography, Construct

    Characterization of BHJRST. (a) Confirmation of the identity of the ingredients. The macroscopic and microscopic appearance of the 5 ingredients used to prepare BHJRST was examined. (b) HPLC chromatograms of water extracts of BHJRST and three of the five single ingredients. Three major peaks were identified in the classical BHJRST formula (top panel) by comparison of their retention times with those of peaks in extracts of the single ingredients Rhizoma Anemarrhenae (Zhi-Mu), Ginseng Radix (Ren-Shen), or Radix Glycyrrhizae Preparata (Zhi-Gan-Cao).

    Journal: Evidence-based Complementary and Alternative Medicine : eCAM

    Article Title: Bai-Hu-Jia-Ren-Shen-Tang Decoction Reduces Fatty Liver by Activating AMP-Activated Protein Kinase In Vitro and In Vivo

    doi: 10.1155/2015/651734

    Figure Lengend Snippet: Characterization of BHJRST. (a) Confirmation of the identity of the ingredients. The macroscopic and microscopic appearance of the 5 ingredients used to prepare BHJRST was examined. (b) HPLC chromatograms of water extracts of BHJRST and three of the five single ingredients. Three major peaks were identified in the classical BHJRST formula (top panel) by comparison of their retention times with those of peaks in extracts of the single ingredients Rhizoma Anemarrhenae (Zhi-Mu), Ginseng Radix (Ren-Shen), or Radix Glycyrrhizae Preparata (Zhi-Gan-Cao).

    Article Snippet: Reverse-Phase HPLC Chemical Fingerprint Analysis of BHJRST All HPLC fingerprint analyses of the water extracts of BHJRST and the single remedies of BHJRST were performed on a HPLC instrument equipped with a Shimadzu 10A system controller (both from Shimadzu Corporation, Kyoto, Japan).

    Techniques: High Performance Liquid Chromatography