1,1′-Carbonyldiimidazole (CDI), dextranase (Penicillium sp.), and 3,5-dinitrosalicylic acid (DNS) were purchased from Sigma Chemical Co. (St Louis, MO, USA). Succinic anhydride and 5-benzyl N-(Boc)-glutamate were purchased from Tokyo Chemical Industry (Tokyo, Japan). Dextran (molecular weight: 15–20 kDa) was purchased from Fluka (Sigma Chemical Co.). Celecoxib was ether-extracted from Celebrex capsules (Pfizer, Inc., New York, NY, USA). All other chemicals were reagent grade, commercially available products. Buffer solutions (pH 1.2 and 6.8) were prepared as described in USP XXIII. Thin layer chromatographys (TLCs) were performed on Merck Kieselgel 60 F254. The high-performance liquid chromatography (HPLC) system consisted of a Model 306 pump, a Model 117 UV detector, a Model 234 autoinjector, and a Model 805 manometric module from Gilson (Middleton, WI, USA). A symmetry column C18 (Waters, Milford, MA, USA) (250×4.6 mm) with a guard column (Waters, 3.9×20 mm) was used. Six-week-old male Sprague Dawley rats (Samtako Bio Korea, Kyeong-gi-do, South Korea) were housed in the university animal facility with controlled temperature, humidity, and dark/light cycle. The animal protocol used in this study has been reviewed and approved by the Pusan National University-Institutional Animal Care and Use Committee (PNU-IACUC) on their ethical procedures and scientific care.
Model 306 pump
Manufactured by Gilson
Sourced in United States
The Gilson Model 306 pump is a reliable and versatile laboratory instrument designed for fluid handling applications. It features a compact design and delivers consistent flow rates to meet the requirements of various laboratory procedures.
Automatically generated - may contain errors
Lab products found in correlation
Model 234 autoinjector, by Gilson (5 mentions)
Guard column, by Waters Corporation (4 mentions)
Symmetry c18 column, by Waters Corporation (2 mentions)
Model 805 manometric module, by Gilson (2 mentions)
Omnic software, by Gilson (2 mentions)
Model uv170 detector, by Gilson (2 mentions)
Symmetry r18 column, by Waters Corporation (2 mentions)
Kieselgel 60 f254, by Merck Group (1 mentions)
3 5 dinitrosalicylic acid, by Merck Group (1 mentions)
Succinic anhydride, by Tokyo Chemical Industry (1 mentions)
Celecoxib, by Pfizer (1 mentions)
Lichrospher cn column, by Merck Group (1 mentions)
Trifluoroacetic acid, by Merck Group (1 mentions)
151 variable uv detector, by Gilson (1 mentions)
7 protocols using model 306 pump
1
Dextran-based Celecoxib Conjugate Synthesis
2
HPLC Analysis of Drug Compounds
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Separation was carried out on a LiChrospher®CN column (125 × 4.0 mm, 5 μm) from Merck. The column was housed in a column heater set at 22 °C. The mobile phase consisted of 0.1% formic acid, 10 mM ammonium formate and acetonitrile (55 : 15 : 30, v/v/v). The flow rate of the mobile phase was 1.4 mL min−1. Chromatography was performed with a model 306 pump with a loop Rheodyne (20 μL) and a model UV170 detector from Gilson (Middleton, USA) set at 228 nm for quantitative measurements, and additively at 254 and 290 nm to detect degradation products. The system was controlled by Omnic software from Gilson.
3
HPLC Analysis of Celecoxib and Metabolites
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The HPLC system consisted of a model 306 pump, a 117 variable ultraviolet detector, a model 234 autoinjector, and a Model 805 manometric module from Gilson Inc. (Middleton, WI, USA). A C18 symmetry column (Waters, Milford, MA, USA) (250 × 4.6 mm) with a guard column (Waters, 3.9 × 20 mm) was used. The samples were filtered through a membrane filter (0.45 μm), and the filtrate (20 μL) was injected on a symmetry C18 column (Waters), which was eluted with a mobile phase at a flow rate of 1 mL/min. The mobile phase consisted of 60% ACN (Merck, Darmstadt, German) in 0.067 M phosphate buffer (pH 4.5) containing 0.1% trifluoroacetic acid (Sigma-Aldrich), which was filtered through a 0.45 μm membrane filter (Waters) before use. The eluate was monitored at 273 nm, and the detection limit was about 0.2 μg/mL under our experimental conditions. Accuracy and relative standard deviations were 98.7% and 0.43%, respectively Trilution® LC V4 software (Gilson, Middleton, WI, USA) was used for data analysis. The retention times of celecoxib, A1C, G1C, and N-GA1C were 10.68, 3.11, 3.00, and 2.43 min, respectively.
4
HPLC Analysis of 5-ASA and MCP
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The HPLC system consisted of a model 306 pump, a 151 variable UV detector, and a model 234 autoinjector from Gilson (Middleton, WI, USA). We used a Symmetry R18 column (250×4.6 mm, 5 μm; Waters, Milford, MA, USA) with a guard column (20×4.6 mm; Waters). The samples prepared from each experiment were filtered using a 0.45 μm membrane filter. HPLC analysis was conducted at a flow rate of 1 mL/min using mobile phases consisting of acetonitrile and 3.0 mM pH 3.0 phosphate buffer (2:8, v/v) for MCP and acetonitrile, and 5.0 mM pH 7.4 phosphate buffer (2:8, v/v) for 5-ASA. The eluate was monitored at 323 nm (for 5-ASA) and 275 nm (for MCP) using a UV detector measuring the absorption with a sensitivity of 0.01 absorbance units full scale. The retention times of 5-ASA and MCP were 7.5 and 9.4 minutes, respectively.
5
Quantitative HPLC Analysis of 5-ASA and ASP
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The HPLC system consisting of a model 306 pump, a 151 variable UV detector, and a model 234 autoinjector from Gilson (Middleton, WI, USA) was used for experiments. A Symmetry R18 column (250 × 4.6 mm, 5 μm) and a guard column (20 × 4.6 mm, 5 μm) were purchased from Waters (Milford, MA, USA). Samples prepared for HPLC analysis were filtered through a syringe filter (0.45 μm). HPLC analysis was performed at a flow rate of 1 mL/min using a mobile phase comprising acetonitrile and 10 mM pH 4.0 phosphate buffer (1.5:8.5, v/v). The eluate was monitored at 323 nm (for 5-ASA) and 225 nm (for ASP) with the UV detector, which measured the absorption with a sensitivity of AUFS 0.01. The retention times of 5-ASA and ASP were 7.1 and 5.5 min, respectively.
6
HPLC Analysis of Pharmaceutical Compounds
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The high-performance liquid chromatography (HPLC) system consisted of a Gilson model 306 pump, 151 variable UV detector, and model 234 autoinjector (Gilson, Middleton, WI, USA). Chromatographic separation was conducted using a symmetric C18 column (Hector, Theale, Berkshire, UK; 250 × 4.6 mm, 5 μm). Before injection for HPLC analysis, samples were filtered through membrane filters (0.45 μm, Revodix Inc., Gyeong-gi-do, Hanam, Korea). Mobile phases were prepared as follows: mobile phase A consisted of distilled water and acetonitrile (7:3, v/v), and mobile phase B consisted of acetonitrile and 1 mM phosphate buffer (pH 7.4) with 0.5 mM tetrabutylammonium chloride (1.5:8.5, v/v). The HPLC analysis was conducted at a flow rate of 1 mL/min. The eluate was monitored at 295 nm (for DpS), 367 nm (for AS-DpS-AS) and 330 nm (for 5-ASA) using a UV detector that measured the absorption with a sensitivity of AUFS 0.01. The retention times of DpS and AS-DpS-AS using mobile phase A were 9.8 min and 7.1 min, respectively, and that of 5-ASA using mobile phase B was 10.1 min.
7
Chromatographic Validation Protocol
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Chromatography was performed with a model 306 pump with a loop Rheodyne (20 μl) and a model UV170 detector controlled by OMNIC software (all from Gilson Inc., Middleton, WI, USA). The columns were housed in a column heater set at 25°C. The developed methods were validated according to the ICH and FDA guidelines for their specificity, linearity, sensitivity, accuracy, precision, and robustness [27 , 28 ].
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