Supernatant from the BSG fermentation (100 μl) was obtained by centrifugation (8,000 ×g, 5 min, 4°C), followed by dansylation at 80°C for 40 min after addition of 1 M sodium carbonate–sodium bicarbonate buffer (pH 10.0; 200 μl), dansyl chloride dissolved in acetone at 20 mg/ml (100 μl), and distilled water (600 μl). Then, 10%(v/v) acetic acid (100 μl) was added to terminate the reaction. After centrifugation, the reaction solution was filtered with a 0.2-μm syringe filter (Merck Millipore, USA) and injected into a high-performance liquid chromatography (HPLC) system for analysis at 254 nm and 30°C [29 (link)]. A JASCO HPLC system (JASCO LC-2000 Plus Series, Japan) with a photodiode array detector, an autosampler (JASCO AS-2055 Plus), and an AccQ-Tag C18 column (3.9 × 150 mm, 4 μm; Waters Corporation, UK) was used. Eluent A was prepared by mixing tetrahydrofuran, methanol, and 50 mM sodium acetate–acetic acid buffer (pH 6.2) at a ratio of 1:15:84 (v:v:v), and eluent B was methanol. Eluents A and B were applied at a flow-rate of 1 ml/min for 5 min, 16 min, and 4 min at ratios of 80:20, 45:55, and 0:100, respectively [29 (link)]. The GABA content was calculated using calibration curves determined at five concentrations with GABA standard material (Sigma-Aldrich, USA). All experiments were conducted three times.
Accq tag c18 column
Manufactured by Waters Corporation
Sourced in United Kingdom, United States
The AccQ-Tag C18 column is a high-performance liquid chromatography (HPLC) column designed for the analysis of amino acids. The core function of this column is to separate and detect amino acids in various samples using HPLC techniques.
Automatically generated - may contain errors
Lab products found in correlation
0.2 μm syringe filter, by Merck Group (1 mentions)
Lc 2000 plus series, by Jasco (1 mentions)
Accq tag ultra derivatization kit, by Waters Corporation (1 mentions)
Acquity uplc h class system, by Waters Corporation (1 mentions)
6 aminoquinolyl n hydroxysuccinimidyl carbamate, by Waters Corporation (1 mentions)
Binary hplc pump, by Waters Corporation (1 mentions)
Ods a column, by YMC (1 mentions)
Accq tag, by Waters Corporation (1 mentions)
1260 infinity series hplc system, by Agilent Technologies (1 mentions)
5 protocols using accq tag c18 column
1
GABA Quantification from BSG Fermentation
2
Amino Acid Profiling of Pegfilgrastim
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Amino acid composition was estimated using AccQ Tag Ultra Derivatization kit (Waters). Pegfilgrastim samples (200 μg) were hydrolyzed at 110°C for 24 h in vacuum hydrolysis tubes. After hydrolysis, samples were labeled with the dye 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (Waters) by incubating at 55°C for 10 min. The labeled samples were injected into a Waters Acc Q Tag C18 column (130 Å, 2.1 × 100 mm). The chromatographic column was equilibrated with 10% of solvent A (proprietary of Waters) and 90% of solvent B (proprietary of Waters). The peaks were separated using Waters ACQUITY UPLC H-Class System by applying a combination of a gradient of solvent A (10% to 4% in 8.5 min), solvent B (0% to 80% in 5.2 mins, 80% to 15.6 in 1.6 min), solvent C (90.1% to 36.3% in 8.3 mins) and solvent D (0% to 59.7% in 8.6 min) and detected at 260 nm keeping the column temperature at 43°C with a flow rate of 0.7 mL/min. Water was used as solvent C and solvent B was used as solvent D.
The amino acids eluted between 2 to 9 min. The mole % of each amino acid was calculated based on the calibration curve generated for respective amino acid standards.
The amino acids eluted between 2 to 9 min. The mole % of each amino acid was calculated based on the calibration curve generated for respective amino acid standards.
3
Amino Acid Composition Analysis of Silkworm Pupae
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Determination of amino acids composition in silkworm pupae protein was performed according to AccQ Tag method using HPLC (Waters, 2690, Waters Corp., Milford, MA, USA) with a Waters AccQ Tag C18 column (3.9 mm × 150 mm) at the temperature of 37 °C. The samples of 10 µL were injected and their peaks were compared to standard amino acids.
4
Quantifying Polyphenols, Amino Acids, and Flavonoids
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Assessment of total polyphenol content was performed according to the modified Singleton method using gallic acid as standard [23 ]. The free amino acid and GABA contents of water extracts of Z. jujuba and D. longan were measured using the AccQ·Tag method (Bae et al., 2023). The high-performance liquid chromatography (HPLC) instrument used was a Waters product consisting of a 1525 pump (Binary HPLC Pump) and a 474 fluorescence detector (Scanning Fluorescence Detector). In addition, an ACCQ-Tag C18 column (3.9 mm × 150 mm I.D., 4 µm) was purchased from Waters and used, and the excitation wavelength of the fluorescence detector was set to 250 nm and the measurement wavelength was set to 390 nm. The flow rate was set at 1 mL per minute and 10 µL was injected. Mobile phase A was used by diluting 100 mL of AccuQTag Eluent A with 1 L of water, and mobile phase B was analyzed in gradient mode using 60% acetonitrile (water: acetonitrile, 40:60, v/v). The quercetin-3-glucuronide (Q3G) content of lettuce (L. sativa) water extract was detected at a wavelength of 350 nm using a YMC-Pack ODS-A column (250 mm × 4.6 mm, 5 μm). The mobile phase consisted of 0.5% formic acid in water (A) and 0.5% formic acid in acetonitrile (B). The gradient of the mobile phase was 80% A for 0 min, 77% A for 5 min, 73% A for 20 min, and 80% A for 25–30 min [24 (link)].
5
GABA Quantification in Plant Extracts
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The GABA production in aqueous extracts from FM was quantified by reversed-phase high-performance liquid chromatography in a Series 1260 Infinity HPLC system (Agilent Technology, Waldbronn, Germany). Aliquots of 2 mL were centrifuged (Eppendorf, Hauppauge, NY, USA) at 12000×g for 10 min at room temperature to remove solids and bacterial cells and were then ultrafiltered through membranes with a pore size of < 3 kDa (Pall Life Sciences, Port Washington, NY, USA) (Wu and Shah 2016) . Afterwards, a derivatization of the extracts was made in a precolumn following the methodology of the 6-aminoquinolyl-n-hydroxy succinimidyl carbamate commercial kit AccQ-Tag (Waters Corporation, Ciudad de México, México).
Chromatographic separation was performed using an AccQ-Tag C18 column (1.7 μm, 2.1 × 100 mm) maintained at a constant temperature (37 °C). Mobile phase A consisted of an AccQ-Tag ultra-eluent A, and phase B consisted of AccQ-Tag ultra-eluent B. Finally, phase C consisted of an eluent of Milli-Q water. The gradient separation times were set as follows: (A:B:C), 0 min, (100:0:0), 0.5 min (100:0:0), 18 min (95:5:0), 19 min (91: 9:0), 29.5 min (83:17:0), 38 min (0:60:40), and 41 min (100:0:0). Detection was performed by UV absorbance at 254 nm to quantify GABA (Diana et al. 2014b ).
Chromatographic separation was performed using an AccQ-Tag C18 column (1.7 μm, 2.1 × 100 mm) maintained at a constant temperature (37 °C). Mobile phase A consisted of an AccQ-Tag ultra-eluent A, and phase B consisted of AccQ-Tag ultra-eluent B. Finally, phase C consisted of an eluent of Milli-Q water. The gradient separation times were set as follows: (A:B:C), 0 min, (100:0:0), 0.5 min (100:0:0), 18 min (95:5:0), 19 min (91: 9:0), 29.5 min (83:17:0), 38 min (0:60:40), and 41 min (100:0:0). Detection was performed by UV absorbance at 254 nm to quantify GABA (Diana et al. 2014b ).
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