Lyophilized peptides (Lifetein LLC) were dissolved in 10% dimethyl sulfoxide (DMSO), mixed with 127 nM C. albicans Bar1 or Bar1(D232A) protein in 50 µM potassium phosphate solution (pH 6.5) for 1 h at 37°C, and then passed through a centrifugal filtration unit to separate peptide fragments from the peptidase. Samples were analyzed using a Shimadzu high-performance liquid chromatograph and Thermo LCQ Deca XP Max ion trap mass spectrometer system.
High performance liquid chromatograph
Manufactured by Thermo Fisher Scientific
Sourced in United States
A high-performance liquid chromatograph (HPLC) is an analytical instrument used to separate, identify, and quantify components in a liquid mixture. It functions by pumping the sample mixture through a column packed with a stationary phase, where the components are separated based on their interactions with the stationary phase and the mobile phase.
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Lab products found in correlation
2 protocols using high performance liquid chromatograph
1
Peptide Characterization by Mass Spectrometry
2
Antioxidant Enzyme Activities and AsA Assay
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The methods of extraction and measurement have been illuminated in detail in our previous study (Yan et al., 2010) . SOD activity (U g À1 FW) was determined by the inhibition on nitro blue tetrazolium reduction. APX (mmol AsA min À1 g À1 FW) and CAT (mmol H 2 O 2 min À1 g À1 FW) activities were, respectively, estimated by AsA oxidation rate and H 2 O 2 decomposing rate. AsA content (mg g À1 FW) was detected by using a high performance liquid chromatograph (Thermo, USA) with a hypersil C18 column (5.0 mm particles size, 4.6 Â 150 mm). Dithiothreitol was used to completely reduce the oxidized AsA for detecting total AsA content and calculating AsA redox state. MDA content was reckoned by the reaction with thiobarbituric acid.
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