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Separation was achieved with an isocratic gradient of 0.01 N H2SO4 at a flow-rate of 0.6 ml/min and the temperature of the column oven was set to 60 °C.
The mobile phase was methanol and H2O with an isocratic gradient (50/50).
The ovorubin peak was then further purified by size exclusion chromatography (Superdex 200 HR 10/20, Amersham-Pharmacia, Uppsala, Sweden) using an isocratic gradient of sodium phosphate buffer 50 mM, 150 mM NaCl, pH 7.6.
The mobile phase consisted of an isocratic gradient system as described by Ferreres et al. [ 13] with some modifications.
An isocratic gradient of 33% B was used to elute unconjugated material, followed by a linear gradient of 41 95% B to elute mono- and diconjugated species.
In order to elute the samples, an isocratic gradient of 3.2 mM NaOH with a flux rate of 0.15 ml/min was applied for 20 min.
Similar(41)
Precision was excellent for both isocratic and gradient analyses.
Finally the retention is adjusted using isocratic or gradient elution.
The different elements involved in these methodologies, for both isocratic and gradient elution, are given.
The transferability of retention data among isocratic and gradient RPLC elution modes is studied.
The new suppressor has been shown to perform well under both isocratic and gradient elution conditions.
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