Reverse-phase LC-MS/MS was performed as previously described [32] , using an Agilent 1200 SL system (Agilent Technologies, Dorset, United Kingdom) connected to a 6410 triple quadrupole LC-MS/MS with electrospray ionization on the negative mode, with multiple reaction monitoring (Agilent Technologies, Santa Clara, CA, USA) with a Kinetex C18 column (2.6 μm, 150×2.1 mm; Phenomenex, Cheshire, United Kingdom). As green tea catechin conjugates (sulfate and glucuronide) are not commercially available, catechin conjugates were analyzed using peak area, and free forms were analyzed relative to known standards, relative to the internal standard EG (2 μg/ml). A total of 55 metabolites (green tea catechins, metabolites and conjugated forms) were investigated, and the MS transitions are as previously reported [32] .
Agilent 1200 sl system
Manufactured by Agilent Technologies
Sourced in United Kingdom, United States
The Agilent 1200 SL system is a high-performance liquid chromatography (HPLC) instrument designed for analytical applications. It features a compact and modular design, enabling flexible configurations to meet various laboratory requirements. The system includes components such as a pump, autosampler, and detector, providing reliable and efficient separation and detection of chemical compounds.
Automatically generated - may contain errors
3 protocols using agilent 1200 sl system
1
Comprehensive LC-MS/MS Analysis of Green Tea Metabolites
2
Validating loxB Deletion and Overexpression in A. fumigatus
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To confirm the functionality of our deletion and overexpression loxB strains, Costar® 12-well cell culture plates (Corning, Corning, NY, USA) containing 3 mL of GMM media were inoculated with spores of wild type, ΔloxB and OE::loxB A. fumigatus mutants to a final concentration of 1 × 106 spores/mL (n = 3). Plates were covered with AeraSealTM Breathable Sealing Film (Excel Scientific, Victorville, CA, USA) and incubated for 3 days at 37°C and 185 rpms. The antioxidant cocktail (0.2 mg/mL BHT, TPP, and EDTA) were added to prevent oxylipin degradation.
The culture supernatants were extracted using solid phase extraction protocol as described previously (Yang et al., 2009 (link)). The reconstituted solutions were injected onto a UPLC/MS/MS system (Agilent 1200 SL system, Santa Clara, CA, USA) coupled to AB Sciex 4000 Qtrap MS/MS system (Foster City, CA, USA). The detailed parameters were described in a previous paper (Zivkovic et al., 2012 (link)). Limit of quantification (LOQ) for 13-HODE is presented in Supplementary TableS2 .
Levels of 13-HODE between wild type, ΔloxB, and OE::loxB strains were analyzed for significance using students T-test via GraphPad Prism version 6.00 for Windows, GraphPad Software. (La Jolla, CA, USA2 ). Values in figures are expressed as mean (n = 3) ± SEM.
The culture supernatants were extracted using solid phase extraction protocol as described previously (Yang et al., 2009 (link)). The reconstituted solutions were injected onto a UPLC/MS/MS system (Agilent 1200 SL system, Santa Clara, CA, USA) coupled to AB Sciex 4000 Qtrap MS/MS system (Foster City, CA, USA). The detailed parameters were described in a previous paper (Zivkovic et al., 2012 (link)). Limit of quantification (LOQ) for 13-HODE is presented in Supplementary Table
Levels of 13-HODE between wild type, ΔloxB, and OE::loxB strains were analyzed for significance using students T-test via GraphPad Prism version 6.00 for Windows, GraphPad Software. (La Jolla, CA, USA
3
HPLC Characterization of Polyphenols
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The major polyphenols in the fruit samples and green tea were characterised using HPLC as described previously (27) . An Agilent 1200 SL system (Agilent Technologies) equipped with a diode array detector (DAD) was used. It comprised of a binary pump, degasser, column oven (35°C) and a well plate autosampler (5°C). A Zorbax Eclipse plus C18 column (1•8 µm, 100 × 2•1 mm) and Agilent-Zorbax eclipse XDB-C18 (1•8 µm, 50 × 4•6 mm), both from Agilent Technologies, were used for green tea and fruit extracts, respectively. Other parameters were 5-µl injection volume, at 0•5 ml/min flow rate with needle wash in the flush point for 3 s. For all analyses, ultrapure, nucleasefree water (≥18•2 MΩ cm at 25°C) from a Millipore Q water purifying system (Millipore) was used. For sample preparation, a Genevac (EZ-2 plus model, Fisher Scientific Ltd) was used for centrifugal evaporation. Polyphenols were identified by their retention times compared with authentic standards, and standard curves were used for quantification.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!