CH4 in the headspace of slurries was analyzed by gas chromatography as described previously [23 (link)]. Volatile fatty acids from centrifuged (10,000× g, 10 min) samples of the sediment slurries were analyzed by HPLC as described previously [23 (link)]. Data analyses were performed using ChromQuest (Thermo Scientific, Waltham, MA, USA) and Chromeleon software (Thermo Scientific, Waltham, MA). Sulfate and sulfide were quantified as described in [24 (link)].
Chromquest
Manufactured by Thermo Fisher Scientific
Sourced in United States
ChromQuest is a software suite designed for chromatography data analysis and reporting. It provides a comprehensive solution for managing and interpreting chromatographic data from various analytical instruments.
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6 protocols using chromquest
1
Biogeochemical Analysis of Sediment Slurries
2
Quantitative LC-UV Analysis of Ascorbic Acid
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We used a liquid chromatographic separation with ultraviolet light detection (LC–UV) method adapted and re-validated from established LC–UV protocols [10 (link),11 ,12 (link)]. The LC system consisted of a solvent delivery pump (Model P4000; Thermo Separation Products; San Jose, CA, USA), which pumped a mixture of 5% methanol and 95% 0.05 M phosphoric acid (HPLC grade, cat # P3786; Sigma Aldrich, Oakville, ON, Canada) through a 15 cm × 4.6 mm reversed-phase 5 µm column (Zorbax SB-CN; Agilent Technologies Canada Inc., Mississauga, ON, Canada) at 1.0 mL/min. Three microliters of each prepared sample, quality control or standard was injected directly onto the LC column using an autoinjector (Ultra WISP 715; Waters Scientific, Toronto, ON, Canada) in duplicate.
The column effluent was monitored with a variable wavelength UV detector (UV 6000 Thermo Separation Products; San Jose, CA, USA). The signal from the detector at 246 nm was integrated and recorded with a chromatography data system (Chrom Quest, version 5.0, ThermoFisher Scientific Inc., Nepean, ON, Canada). The area under the AA peak at 246 nm was subjected to least squares linear regression and the actual AA concentration in each sample determined by interpolation from the standard curve.
The column effluent was monitored with a variable wavelength UV detector (UV 6000 Thermo Separation Products; San Jose, CA, USA). The signal from the detector at 246 nm was integrated and recorded with a chromatography data system (Chrom Quest, version 5.0, ThermoFisher Scientific Inc., Nepean, ON, Canada). The area under the AA peak at 246 nm was subjected to least squares linear regression and the actual AA concentration in each sample determined by interpolation from the standard curve.
3
Lipid Extraction and Fatty Acid Analysis
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We used the method of Bligh and Dyer (18 (link)) on a reduced-scale to extract total lipids from the gastrocnemius muscle. FA methyl esters (FAME) were prepared by ultrasound-assisted total lipid methylation, as described by Santos et al. (19 (link)). FAME was separated by gas chromatography. Retention times and peak areas were determined using the Chrom-Quest™ software (Thermo Scientific™, USA). FA contents in the muscles were reported as mg/g of total fat.
4
Quantitative Analysis of Phenolic Compounds
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ChromQuest (version 3.1.6) software, Thermo Fisher Scientific, Waltham, MA, USA and 4880 software (Unicam, Lisbon, Portugal) were used for data acquisition and treatment of HPLC-DAD and ED analyses, respectively. The identification of pCA and FA was performed by comparison with standard solutions using commercially available standards and the identification of the main dehydrodiferulic acids and hydroxycinnamic acid amides was based on results previously reported [11 (link)]. For quantitative data analyses, the limit of significance was set at p < 0.05. Paired-samples t-tests, independent-samples t-tests, ANOVA followed by post hoc Tukey tests, principal component analyses (PCA) and Pearson’s coefficient correlations were obtained using the software SPSS version 21 (IBM, NY, USA).
5
Biogeochemical Analyses of Sediment Slurries
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CH4 in the headspace of slurries was analyzed by gas chromatography as described previously [20 (link)]. Volatile fatty acids from centrifuged (10,000× g, 10 min) samples of the sediment slurries were analyzed by HPLC as described previously [20 (link)]. Data analyses were performed using ChromQuest (Thermo Scientific, Waltham, MA, USA) and Chromeleon software (Thermo Scientific, Waltham, MA, USA). Sulfate concentrations were analyzed by Ion Chromatography system equipped with an AS22 column (4 × 250 mm) and ED 40 electrochemical detector (Dionex, Sunnyvale, CA, USA). The eluents were 1.7 mM NaHCO3 and 1.8 mM Na2CO3. The analyses were conducted with a flow rate of 1.2 mL min−1 at 35 °C. Sodium bromide was used as internal standard. Sulfide measurements were done using the methylene blue method [18 (link)].
6
HPLC-DAD-MS/MS for Compound Identification
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ChromQuest (Thermo Fisher Scientific, Waltham, MA, USA) and MassLynx (Waters, Dublin, Ireland) software were used to control analytical conditions and collect data from HPLC-DAD and HPLC-DAD-MS/MS, respectively. For compounds identification purposes, mass and UV spectra were compared with spectra already published in the literature. When standards were commercially available, the identification was based on the comparison of their fragmentation patterns and retention times.
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