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Different mobile phase additives were compared, including acetic acid, formic acid, ammonium formate, and a range of primary amines.
Chaotropic agents recently gained popularity as interesting and useful mobile phase additives in liquid chromatography due to their effect on analytes retention, peak symmetry and separation efficiency.
The aim of this paper is to present a development of liquid chromatographic method when chaotropic salts are used as mobile phase additives following the QbD principles.
The effects of the organic modifiers, modifier composition, mobile phase additives, and temperature were investigated for the separation of these amides on the IC or the OD-H column.
HPLC-grade solvents (acetonitrile, methanol, water), glass fiber filters (Type A/E, 90 mm, 1 µm), Gelman Acrodisc® CR PTFE syringe filters (13 mm, 0.45 µm), and mobile phase additives, ACS grade formic acid (98%) and trifluoroacetic acid (99%) were also used.
These compounds are commonly used mobile phase additives in LC/MS of proteins and peptides.
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In a first step, the parameters potentially influencing the separation, such as addition of an alcohol to the polar organic solvent or the type of mobile phase additive(s), were examined by means of experimental designs.
The LC/MS experiments revealed that ascorbic acid is a convenient mobile phase additive.
Solid-phase extraction was used for sample preparation, after which hydroxypropylbetadex was complexed with a mobile phase additive (1-napthol).
Formic acid ∼98%% (FA), and ammonium formate (NH4FA), as LC-MS grade mobile-phase additives, were purchased from Fluka.
A limitation of ESI is the occurrence of ion-suppression (matrix) effects, which are typically caused by mobile-phase additives (such as acids and bases) and coeluting impurities from biological samples [54,55].
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