The analysis of 5-FU was carried out at room temperature (25 ± 1°C) using Waters HPLC system (Waters, USA) equipped with 1515 LC pump, 717 autosamplers, quaternary LC-10A VP pumps, a programmable UV-visible variable-wavelength detector, SPD-10AVP column oven, an SCL 10AVP system controller (Shimadzu, Japan) and an inline vacuum degasser was used [22 (link)]. The software utilized for data analysis was Millennium (version 32). All analysis was performed using a Lichrosphere (150 mm × 4 mm) RP Nucleodur C18 column (Macherey Nagel, Germany) having a 5 μm particle size. The mobile phase used for this analysis was a binary mixture of methanol and water (80:20% v/v). The mobile phase has flowed with a flow rate of 1.0 mL/min with UV detection at 254 nm. The sample volume was 10 μL for each analysis. The same instrument, same chromatographic conditions were used for the analysis of 5-FUC except for the mobile phase. The mobile phase used for the analysis of 5-FUC was a binary mixture of methanol and ethyl acetate (70:30% v/v) [24 ].
Spd 10avp column oven
Manufactured by Shimadzu
Sourced in Japan
The SPD-10AVP column oven is a temperature-controlled device designed for use in high-performance liquid chromatography (HPLC) systems. It is responsible for maintaining a consistent temperature within the column, which is crucial for accurate and reproducible separation of analytes. The core function of the SPD-10AVP is to provide a stable and controlled environment for the chromatographic column, ensuring optimal performance and reliable results.
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4 protocols using spd 10avp column oven
1
HPLC Analysis of 5-FU and 5-FUC
2
HPLC Quantification of NRG
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NRG was assayed by HPLC using a system consisting of a quaternary LC-10AT VP pump equipped with UV/VIS detector, SPD-10AVP column oven (Shimadzu), a Rheodyne injector, and chromatography data system software (CLASS-VP Ver 6.14 SP1). Chromatographic separation was carried out on column (LiChrospher®100 RP-18 (5 μm), Merck, Darmstadt, Germany) with a mobile phase consisting of a 7:3 ratio of methanol and water with 0.1 % glacial acetic acid at ambient temperature (25 ± 0.5°C) having a flow rate of 1 ml/min. Samples (20 μl) were filtered through microfilters having 0.45 μm pore size and Rheodyne injector was used for injecting the samples which were then examined at 289 nm [22 (link)].
3
In Vitro Drug Release Kinetics of ZP-LPS
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The drug release behavior of ZP-LPS was determined by a dialysis method using PBS (pH 7.4) containing 1% Tween 20 as dissolution medium. A comparison was done with ZP suspension. ZP-LPS (equivalent to 20 mg ZP) or ZP (20 mg) was suspended in 2.0 mL of dissolution medium contained in a dialysis bag (MWCO: 12000 g/mol). The dialysis bag was then suspended in a 200 mL glass beaker containing 100 mL of dissolution medium under mild agitation in a water bath at 37°C. At pre-designated time points for up to 30 days, the dissolution medium (0.5 mL) was withdrawn following the addition of the same volume of the fresh medium. The analyses of the samples were done using the previously published HPLC method following slight modification and re-validation.15 (link) Analysis was done using 5µm RP 18 (C18), Lichrospher®100, (250×4.6 mm) attached to an HPLC equipped with variable wavelength programmable UV/VIS detector, quaternary LC-10 AT VP pump, SCL 10A VP system controller (Shimadzu), SPD-10 AVP column oven (Shimadzu), and a Rheodyne injector with a 20µL loop. For the drug analysis class-VP 5.032 software was used. Process parameters used for separation were as follows: temperature, 25 ± 2 °C; mobile phase, phosphate buffer and acetonitrile at 65:35 (v/v); flow rate, 1.0 mL/min;pH, 3; and detection wavelength of ZP, 318 nm.All experiments were performed in triplicate.
4
HPLC Analysis of Plant Extracts
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Chromatographic experiments were conducted on a Shimadzu (Shimadzu Corporation, Kyoto, Japan) HPLC instrument comprising quaternary LC-10A VP pumps, a variable wavelength programmable UV-visible detector, an SPD-10A VP column oven (Shimadzu Corporation, Kyoto, Japan), and a SCL 10A VP system controller. The instrument was controlled by use of Class VP 5.032 software (Shimadzu Corporation, Kyoto, Japan) installed with the equipment. Dried plant extract (10 mg) obtained by a rotatory evaporator was dissolved in 10 mL of HPLC grade methanol and sonicated for dissolving. This sample solution was filtered through a 0.22 µL syringe and then injected using a Rheodyne injector fitted with a 20 µL fixed loop. The separation was achieved using a column with dimensions of 15 × 4.6 mm, a particle size of 5 µm, and a C18 reverse phase column. The mobile phase used consisted of acetonitrile and water in the ratio of 50:50 v/v. All the analyses were performed at room temperature and chromatograms were monitored at wavelengths of 220, 254, and 300 nm using a UV visible detector (Shimadzu Corporation, Kyoto, Japan).
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