Electrospray time-of-flight mass spectrometry (ESI-TOF-MS) was carried out using an Agilent 1200 HPLC system equipped with an Agilent 6210 time-of-flight (ESI-TOF) mass spectrometer (Santa Clara, CA, USA). The nWPH and pWPH samples were separated using gradient conditions on an Agilent Eclipse C18 column (3 × 50 mm; 1.8 μm) (Agilent Technologies, CA, USA) heated to 60 °C. Elution was achieved using solvent A (water + 0.1% formic acid (FA)) and B (100% acetonitrile + 0.1% FA). Gradient conditions were: 5% B at 1 min to 45% B at 25 min, to 95% B at 25.5 min and 95% B at 28 min to 5% B at 28.1 min and 5% B at 32 min with a flow rate of 0.3 mL/min and 5 μL of sample was injected. Accurate mass data were obtained using a dual ESI source in both positive and negative mode (injected in two different methods): data was acquired over a mass range of m/z 100–1000. The source was operated with the following parameters: temperature 350 °C; gas flow 12 L/min; nebulizer 50 psi; capillary voltage 4000 V; fragmentor 100 V; skimmer voltage 60 V. Reference masses (internal calibration of high resolution spectra) were: positive mode: m/z 121.050873, 922.009798; negative mode: m/z 119.03632, 966.000725.
Agilent eclipse c18 column
Manufactured by Agilent Technologies
Sourced in United States
The Agilent Eclipse C18 column is a high-performance liquid chromatography (HPLC) column designed for the separation and analysis of a wide range of organic compounds. It features a stable and versatile C18 stationary phase that provides effective retention and separation of both polar and non-polar analytes.
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2 protocols using agilent eclipse c18 column
1
Electrospray Ionization Time-of-Flight Mass Spectrometry
2
HPLC Analysis of Antiretroviral and Antifungal Drugs
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An Agilent
1260 Infinity series HPLC (Agilent Technologies, Santa Clara, CA)
equipped with an Agilent Eclipse C18 column (particle size: 4.6 μm;
length: 150 mm) was used. Analyses were performed at a flow rate of
1 mL/min using an isocratic elution method. A run time of 8 min was
used. All drugs were detected at 210 nm with an injection volume of
5 μL for donor samples and 100 μL for receiver samples.
For atazanavir, a mobile phase of 40% acetonitrile and 60% water containing
0.1% (v/v) trifluoracetic acid was used. Calibration curves covering
the concentration ranges of 10–80 and 50–400 μg/mL
(5 μL injection volume) as well as 0.1–1 μg/mL
(100 μL injection volume) were established. For lopinavir, a
mobile phase containing 60% acetonitrile and 40% water was used, and
calibration curves were established over the concentration ranges
of 5–120 μg/mL (5 μL injection volume) and 0.025–5
μg/mL (100 μL injection volume). For clotrimazole, the
mobile phase used consisted of 45% acetonitrile and 55% water containing
0.1% (v/v) trifluoracetic acid, and calibration curves of 0–20
and 50–400 μg/mL (5 μL injection volume) as well
as 0.1–5 μg/mL (100 μL injection volume) were used.
The retention time for ATZ, LPV, and CTZ was observed as 4.7, 5.3,
and 5 min, respectively.
1260 Infinity series HPLC (Agilent Technologies, Santa Clara, CA)
equipped with an Agilent Eclipse C18 column (particle size: 4.6 μm;
length: 150 mm) was used. Analyses were performed at a flow rate of
1 mL/min using an isocratic elution method. A run time of 8 min was
used. All drugs were detected at 210 nm with an injection volume of
5 μL for donor samples and 100 μL for receiver samples.
For atazanavir, a mobile phase of 40% acetonitrile and 60% water containing
0.1% (v/v) trifluoracetic acid was used. Calibration curves covering
the concentration ranges of 10–80 and 50–400 μg/mL
(5 μL injection volume) as well as 0.1–1 μg/mL
(100 μL injection volume) were established. For lopinavir, a
mobile phase containing 60% acetonitrile and 40% water was used, and
calibration curves were established over the concentration ranges
of 5–120 μg/mL (5 μL injection volume) and 0.025–5
μg/mL (100 μL injection volume). For clotrimazole, the
mobile phase used consisted of 45% acetonitrile and 55% water containing
0.1% (v/v) trifluoracetic acid, and calibration curves of 0–20
and 50–400 μg/mL (5 μL injection volume) as well
as 0.1–5 μg/mL (100 μL injection volume) were used.
The retention time for ATZ, LPV, and CTZ was observed as 4.7, 5.3,
and 5 min, respectively.
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