A total of 1 ml of blood was obtained from the mouse's tail. Then the blood was separated by centrifugation at 2,000 x g for 10 min at 4°C. The concentration of ketamine and norketamine in the serum and urine (1 ml) was determined using high-performance liquid chromatography. Briefly, samples were collected on the day before the mice were euthanized. The final mobile phase was prepared via mixing ammonium bicarbonate solution (5 mM) adjusted with concentrated ammonia to a pH of 11.3 and acetonitrile in a ratio of 70:30. A Phenomenex LUX® AMP (Phenomenex) 3 µm, 150x4.6 mm column served as the stationary phase. All chemicals were of analytical grade. Chiral separation experiments were carried out with an Agilent 1260 Series Liquid Chromatograph (Agilent Technologies Inc.), equipped with an autosampler and a diode array detector. Each analysis was performed at ambient column temperature or a column temperature of 40°C. The measurements were performed under isocratic conditions with a flow rate of 0.5 ml/min and an injection volume of 1 µl. Ultraviolet detection was performed at 200 nm. Data evaluation was performed via a ChemStation for LC 3D Systems Rev. C. 01.07SR2 software (Agilent Technologies GmbH).
Agilent 1260 series liquid chromatograph
Manufactured by Agilent Technologies
Sourced in United States
The Agilent 1260 series liquid chromatograph is a high-performance liquid chromatography (HPLC) system designed for analytical and preparative applications. It is capable of performing separations and quantitative analysis of a wide range of chemical compounds.
Automatically generated - may contain errors
Lab products found in correlation
Agilent 6410 triple quadrupole mass spectrometer, by Agilent Technologies (1 mentions)
Diamonsil c18 column, by Dikma Technologies (1 mentions)
Agilent 6460 triple quadrupole mass spectrometer, by Agilent Technologies (1 mentions)
Deoxycholic acid dca, by Merck Group (1 mentions)
Sephadex lh 20, by Cytiva (1 mentions)
Gel ods a, by YMC (1 mentions)
Milli q system, by Merck Group (1 mentions)
Hplc grade methanol, by Merck Group (1 mentions)
Lichrospher 60 rp select b, by Merck Group (1 mentions)
Folin ciocalteus s phenol reagent, by Solarbio (1 mentions)
Chemstation, by Agilent Technologies (1 mentions)
Oxyresveratrol, by Yuanye Bio-Technology (1 mentions)
Mulberroside a, by Yuanye Bio-Technology (1 mentions)
C18 column, by Agilent Technologies (1 mentions)
9 protocols using agilent 1260 series liquid chromatograph
1
Quantification of Ketamine and Norketamine
2
HPLC-MS/MS Analysis of Analytes
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The analytes were analyzed on a Diamonsil C18 column (250 × 4.6 mm, 5 μm i.d., Dikma Technologies Inc., CA, USA) using an Agilent 1260 series liquid chromatograph (Agilent Technologies, CA, USA) comprised of an auto injector, a quaternary pump, a diode array detector (DAD), and a column counterpart. The elution was performed with mobile phase composed of 0.1% formic acid in acetonitrile (solvent A) and 0.1% formic acid in water (solvent B) using the following gradient program at the flow rate of 1 mL/min: 0–12 min, linear gradient 15–65% (A); 12-13 min, linear gradient 65–100% (A); 13–25 min, isocratic gradient 100-100% (A). The column temperature was 20°C and the injection volume was 20 μL.
MS/MS analysis was acquired using an Agilent 6410 triple quadrupole mass spectrometer (Agilent Technologies, CA, USA) equipped with an electrospray ionisation (ESI) in the negative mode. Other analytical conditions included the drying gas temperature of 300°C, the drying gas flow of 11 L/min, the nebulizer pressure of 35 psi, and the capillary voltage of 4 kV. To get the highest abundance, the fragmentor voltage and collision energy were optimized. Data acquisition and processing were accepted by Agilent Mass Hunter workstation software version B.05.00 using multiple reaction monitoring (MRM). The quantification was performed using external standard method.
MS/MS analysis was acquired using an Agilent 6410 triple quadrupole mass spectrometer (Agilent Technologies, CA, USA) equipped with an electrospray ionisation (ESI) in the negative mode. Other analytical conditions included the drying gas temperature of 300°C, the drying gas flow of 11 L/min, the nebulizer pressure of 35 psi, and the capillary voltage of 4 kV. To get the highest abundance, the fragmentor voltage and collision energy were optimized. Data acquisition and processing were accepted by Agilent Mass Hunter workstation software version B.05.00 using multiple reaction monitoring (MRM). The quantification was performed using external standard method.
3
Simultaneous Quantitative Analysis of Pharmaceutical Ingredients
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Standards: methylephedrine hydrochloride, bupropion hydrochloride, amphetamine sulfate, fenfluramine, fluoxetine hydrochloride, sibutramine hydrochloride, bixarcomidine, bumetanide, lovastatin, simvastatin and fenofibrate were purchased from the National Institutes for Food and Drug Control (Beijing, China), and rimonabant was purchased from QCC (USA), and the purity was 100%. Chromatographically pure acetonitrile, methanol, formic acid, and acetic acid were purchased from Dima Technology (Beijing, China). Instruments: Agilent 1260 series liquid chromatograph (Agilent), Agilent 6460 triple quadrupole mass spectrometer (Agilent), Synergy UV water purifier (Merck, Germany), TM-1F vortex instrument (Wiggens, Germany), high-speed centrifuge (Hunan Xiang Yi Laboratory Instrument Development Co., Ltd.), SPE-M 96 solid phase extractor device (Agela Technologies), NV-G-S nitrogen blowing device (Agela Technologies), 96-well protein precipitation plate (Nunc, Thermofisher, Pittsburgh, PA).
4
Phytochemical Analysis by LC-MS/MS
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The phytochemical compound in the enriched extract were directly analyzed by liquid chromatography and tandem mass spectrometry (LC-MS/MS) according to the conditions previously described by a previous study [25 (link)]. LC-MS/MS was performed using a Quadrupole/Time-Of-Flight Mass Spectrometer (QTOF LC-MS/MS; Model-6540 UHD Agilent Technologies, Santa Clara, CA, USA) using a Dual ESI ion source attached to an Agilent 1260 series liquid chromatograph. A 150 × 4.6 mm, particle size 5 μm C18 reversed phase column (VertiSep™ AQS—Vertical Chromatography Co., Ltd., Nonthaburi, Thailand) was used at a flow rate of 0.5 mL/min and 40 min of the total run time. The HPLC gradients were consisted of eluent A; 0.1% formic acid in water and eluent B 0.1% formic acid in acetonitrile. The system was run with the following gradient program: 0 min: 95% of linear gradient until 5% of A in 40 min and post run for 5 min. The mass spectral data were acquired with the following ESI inlet conditions: the scanning mass-to-charge (m/z) ranging from 100 to 1700 with a scan rate of 4.00 spectra s−1, the capillary voltage of 3500 V (positive mode) and the fragmentor of 100 V. The pressure of the nebulizer was set at 30 psi, the drying gas temperature at 350 °C, and the continuous gas flow to 10 L min−1.
5
Fecal Bile Acid Quantification by LCMS
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The liquid chromatography-mass spectrometry (LCMS) method was applied to measure fecal bile acid concentration. As reference standards, cholic acid (CA), chenodeoxycholic acid (CDCA), and lithocholic acid (LCA) were purchased from Aladdin, with α-muricholic acid (α-MCA) and β-muricholic acid (β-MCA) from Toronto Research Chemicals, and deoxycholic acid (DCA) from Sigma. And they were added to fecal samples for preliminary measurement by an external standard method. Each fecal sample was suspended in 5 ml of chromatographic ethanol and then ultrasonically extracted for 60 min at 30°C. After 10 minutes of centrifugation (10,000 rpm, 4°C), the supernatant (4 ml) was aspirated and dried under nitrogen. The samples were redissolved with methanol and went through a 0.22 μm filter. Finally, bile acids were analyzed using the Agilent 1260 Series liquid chromatograph combined with a 6120B mass spectrometer. The concentrations of bile acids were determined based on the peak areas [31 (link), 32 (link)].
6
Comprehensive Analytical Characterization of Compounds
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HPLC was performed on an Agilent 1260 series liquid chromatograph (Agilent, Santa Clara, CA, USA) equipped with an evaporation light scattering detector (G4260B), and an Agilent C 18 column [5 μm, 250 mm × 4.6 mm] was also employed. Bruker AVANCE Ⅲ 400 MHz and 600 MHz spectrometers (Bruker, Billerica, MA, USA) recorded the corresponding nuclear magnetic resonance (NMR) spectra. HPLC grade methanol was purchased from Merck (Darmstadt, Germany). Water was purified using a Milli-Q system from Millipore (Bedford, MA, USA). All other reagents were of analytical grade and obtained from conventional commercial sources (Sinopharm Chemical Reagent Co. Ltd., Shanghai, China) . Column chromatography (CC) was performed on macroporous resin (AB-8, 0.3-1.2 mm, Shanghai yuanye Bio-Technology Co., Ltd, Shanghai, China), silica gel Yantai, China) , Sephadex LH-20 (Amersham Pharmacia Biotech AB, Staffanstorp, Sweden) and YMC-Gel ODS-A (50 μm; YMC, Milford, MA, USA). Pre-coated SiO 2 plates (HSGF 254 ; Yantai, China) were used for analytical thin layer chromatography (TLC). Spots were detected on the TLC under UV light or by heating after spraying with vanillin-sulfuric acid reagent.
7
HPLC-MS Analysis of K048 Pharmacokinetics
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Analytical column LiChrospher ® 60 RP-select B (5 µm) LiChroCART ® 125-4 (Merck, Damstadt, Germany) with guard column LiChrospher ® 60 RP-select B (5 µm) was used for analysis. The mobile phase contained 56.2 mmol L -1 Na 2 HPO 4 , 47.9 mmol L -1 citric acid, 0.027 mmol L -1 Na 2 EDTA, 0.925 mmol L -1 octane sulphonic acid, and 75:950 acetonitrile:aqueous phase. The flow rate of the mobile phase was 1 mL min -1 . The pH was adjusted to 3.5 with H 3 PO 4 . Sample volume was 20 µL. All chromatograms were obtained at 40 °C. Samples were analysed with UV detection. The maximum wavelength of K048 was 275 nm.
All pharmacokinetic analyses were performed on an Agilent 1260 Series liquid chromatograph (Palo Alto, CA, USA) composed of degasser, quaternary pump, light-tight autosampler unit set, thermostated column compartment, and MS detector LCQ Fleet (Thermo Finnigan, San Jose, CA, USA). Agilent ChemStation software and statistical software Prism4 (GraphPad Software, San Diego, CA, USA) were used for analysis of results.
All pharmacokinetic analyses were performed on an Agilent 1260 Series liquid chromatograph (Palo Alto, CA, USA) composed of degasser, quaternary pump, light-tight autosampler unit set, thermostated column compartment, and MS detector LCQ Fleet (Thermo Finnigan, San Jose, CA, USA). Agilent ChemStation software and statistical software Prism4 (GraphPad Software, San Diego, CA, USA) were used for analysis of results.
8
Phytochemical Analysis and Antioxidant Evaluation
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Folin-Ciocalteus’s phenol reagent was procured from Beijing Solarbio Co. Ltd, PR China. 1, 1-diphenyl-2-picrylhydrazyl (DPPH), 2, 2-azino-bis (3-ethyl-benzothiazoline-6-sulphonic acid) diammonium salt (ABTS), 2, 4, 6-Tripyridyl-s-triazine (TPTZ), 6-Hydroxy-2, 5, 7, 8-tetramethylchroman-2-carboxylic acid (Trolox), and seven chemical standards (rutin, quercetin, kaempferol, gallic acid, chlorogenic acid, protocatechuic acid, ferulic acid) were provided by Sigma-Aldrich Co., St. Louis, Missouri, USA. Acetic acid, chromatographic-scale methanol and acetonitrile were obtained from Tianjin Bodi Chemical Holding Co. Ltd., China. All solutions involved in this work were filtered with a 0.22 mm nylon filter before use. Analytical-grade reagents were dissolved using deionized water (18 MΩcm). T-400B high-speed multi-function pulverizer was acquired from Dingshuai Hardware Products Co., Ltd., Yongkang City, Zhejiang Province. R-1001VN rotary evaporator was provided from Changcheng Science and Industry Co., Ltd., Zhengzhou City, Henan Province. The HPLC was carried out by an Agilent Series 1260 liquid chromatograph equipped with a quaternary gradient pump system and variable-wavelength detector (VWD) system with a reversed-phase (RP) SB-C18 column (5 μm, 4.6 ×250 mm, Agilent Technologies Inc., USA). Data collection was performed using ChemStation (Agilent Technologies Inc., USA).
9
Quantification of Mulberry Polyphenols by HPLC
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Mulberry samples were grinded
into powder in liquid nitrogen and
lyophilized overnight. A portion (0.5 mL) of absolute methanol was
added to a 1.5 mL EP tube containing 10 mg of dried sample, followed
by sonication for 60 min and centrifugation at 12 000g for 20 min to precipitate plant debris. The supernatants
were then filtered through a 0.22 μm nylon filter and used for
HPLC analysis. The standards of mulberroside A, oxyresveratrol, and
moracin M were purchased from Yuanye Bio-Technology (Shanghai, China),
and the others (moracin C, moracin N, morachalcone A, and chalcomoracin)
were isolated and identified by our lab.40 (link)A 10 μL aliquot of extracted samples and the mixed standard
solution (10 μM for each compound in methanol) was used for
HPLC analysis on an Agilent Series 1260 liquid chromatograph (Agilent)
with a C18 column (250 mm × 4.6 mm, inner dimension: 5 mm, Agilent).
Acetonitrile (solvent A) and 0.1% formic acid in water (solvent B)
were used as the mobile phase with a flow rate of 1.0 mL min–1 at 40 °C. The gradient conditions were optimized as follows:
0–25 min, 10∼50% A; 25–40 min, 50∼95%
A; 40–45 min, 95% A; 45–47 min, 95∼100% A; 47–52
min, 100% A. Spectra were measured at a wavelength of 320 nm.
into powder in liquid nitrogen and
lyophilized overnight. A portion (0.5 mL) of absolute methanol was
added to a 1.5 mL EP tube containing 10 mg of dried sample, followed
by sonication for 60 min and centrifugation at 12 000g for 20 min to precipitate plant debris. The supernatants
were then filtered through a 0.22 μm nylon filter and used for
HPLC analysis. The standards of mulberroside A, oxyresveratrol, and
moracin M were purchased from Yuanye Bio-Technology (Shanghai, China),
and the others (moracin C, moracin N, morachalcone A, and chalcomoracin)
were isolated and identified by our lab.40 (link)A 10 μL aliquot of extracted samples and the mixed standard
solution (10 μM for each compound in methanol) was used for
HPLC analysis on an Agilent Series 1260 liquid chromatograph (Agilent)
with a C18 column (250 mm × 4.6 mm, inner dimension: 5 mm, Agilent).
Acetonitrile (solvent A) and 0.1% formic acid in water (solvent B)
were used as the mobile phase with a flow rate of 1.0 mL min–1 at 40 °C. The gradient conditions were optimized as follows:
0–25 min, 10∼50% A; 25–40 min, 50∼95%
A; 40–45 min, 95% A; 45–47 min, 95∼100% A; 47–52
min, 100% A. Spectra were measured at a wavelength of 320 nm.
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