Agilent 1100 series instrument

Manufactured by Agilent Technologies

Sourced in United States, Germany

The Agilent 1100 series instrument is a high-performance liquid chromatography (HPLC) system designed for analytical and preparative applications. It features advanced technology for precise and reliable separation and detection of chemical compounds. The system includes components such as a solvent delivery module, an autosampler, a column compartment, and a variety of detection modules to meet the diverse needs of laboratory workflows.

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12 protocols using agilent 1100 series instrument

Oxidative Stress Biomarkers in Blood

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Blood samples were collected into heparinized tubes and were examined in the biochemistry laboratory of the study center. Serum levels of oxidized/reduced glutathione (GSH) and malondialdehyde (MDA) concentrations were determined by high-performance liquid chromatograph (HPLC) in the isocratic phase in the Agilent 1100 series instrument with fluorescent detection (Ex:515, Em:553 nm for MDA; Ex:385, Em:515 for GSH) using a kit from Chromsystems Chemicals GmbH (glutathione kits, Chromsystems Chemicals, Munich, Germany; malondialdehyde kits, Chromsystems Chemicals, Munich, Germany). The results were evaluated as µmol/g hemoglobin for GSH and µmol/L hemoglobin for MDA.
Serum levels of glutathione peroxidase were determined using glutathione peroxidase kits (glutathione peroxidase assay kits, Cayman Chemicals, Ann Arbor, USA). ELISA microanalyzer was used for absorbance assays (ChemWell 2910, Awareness Technology Inc., Palm City, USA). The results were evaluated as U/g hemoglobin for glutathione peroxidase.

Tuncer A.A., Bozkurt M.F., Koken T., Dogan N., Pektaş M.K, & Baskin Embleton D. (2016). The Protective Effects of Alpha-Lipoic Acid and Coenzyme Q10 Combination on Ovarian Ischemia-Reperfusion Injury: An Experimental Study. Advances in Medicine, 2016, 3415046.

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General Organic Chemistry Techniques

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All reagents were purchased from
commercial suppliers and used without further purification. Reactions
were carried out in 4 mL screw neck glass vials furnished with screw
caps equipped with poly(tetrafluoroethylene) (PTFE)/rubber septa,
and stir bars under ambient atmosphere unless otherwise noted. Silica
gel 60 Å (40–60 μm, 230–400 mesh) was used
for column chromatography. All NMR spectra were recorded in CDCl₃ using a Bruker AVANCE II 400 MHz or Bruker Avance 500 MHz.
Chemical shifts are given in ppm relative to the residual solvent
peak (¹H NMR: CDCl₃ δ 7.26, ¹³C NMR: CDCl₃ δ 77.16) with multiplicity (br = broad,
s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet),
coupling constants (in hertz), and integration. Kinetic data was analyzed
by Agilent 1260 Infinity Quaternary LC (Eclipse Plus 18C column, 3.5
μm, 4.6 × 100 mm²; UV detector, 265 nm) with
a gradient of acetonitrile and 0.1% formic acid in Milli-Q water at
a flow rate of 1 mL/min. The analytes were calibrated using a five-point
calibration curve with threefold dilution between each sample in the
series. HPLC with a chiral stationary phase was performed on an Agilent
1100 series instrument. High-resolution mass spectrometry analyses
were performed by Thermo Scientific Q Exactive HF Hybrid Quadrupole-Orbitrap
HESI or Bruker microTOF ESI, and low-resolution mass analyses by Bruker
Daltonics amaZon speed no 06052 ESI.

Villo P., Dalla-Santa O., Szabó Z, & Lundberg H. (2020). Kinetic Analysis as an Optimization Tool for Catalytic Esterification with a Moisture-Tolerant Zirconium Complex. The Journal of Organic Chemistry, 85(11), 6959-6969.

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Validated HPLC Method for Quantifying Thymoquinone

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For this study, an Agilent 1100 series instrument (Agilent Technologies, Santa Clara, CA, USA) coupled with UV detection (DAD) and HP Chemstation software V. 32 was used to validate the HPLC method. To analyze the concentration of TQ, a mobile phase of 80% acetonitrile and 20% water was pumped through an Agilent Eclipse XDB-C18 5 µm, 250 × 4.6 mm column with an injection volume of 20 µL and flow rate of 1.0 mL/min. The column temperature was set to 23 °C with UV detection of 250 nm used, with a retention time of 4.2 min. At a concentration of 0.39–100 µg/mL, the method was linear with an R² value of 0.99. Intra- and inter-day precision and accuracy of the method showed a % CV of 0.01 and 0.1, respectively, which is lower than the requirements of 2%.

Haq A., Kumar S., Mao Y., Berthiaume F, & Michniak-Kohn B. (2020). Thymoquinone-Loaded Polymeric Films and Hydrogels for Bacterial Disinfection and Wound Healing. Biomedicines, 8(10), 386.

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Cecal Short-Chain Fatty Acid Quantification

Cited 1 time

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Contents of cecum were extracted as described previously and the concentration of acetate and butyrate in each extract was measured via high-performance liquid chromatography. Briefly, distilled water was added to 20-50 mg of cecal content to a final weight of 200 mg. Following incubation at 80°C for 15 minutes, each sample was centrifuged at 13 000 rpm for 10 minutes, after which the supernatant was filtered through a membrane filter (pore size 0.45 μm). Acetate and butyrate in each sample was then separated and measured using an Agilent 1100 series instrument (Agilent Technologies, Inc, Santa Clara, CA, USA) equipped with a C18 column (ZORBAX Eclipse XDB-C18, analytical 4.6 × 150 mm, 5-Micron; Agilent) and a UV detector (210 nm). The mobile phase consisted of 90% 10 mM KH²PO^{4 (link)} and 10% acetonitrile.

Choi S.I., Son J.H., Kim N., Kim Y.S., Nam R.H., Park J.H., Song C.H., Yu J.E., Lee D.H., Yoon K., Min H., Kim Y.R, & Seok Y.J. (2021). Changes in Cecal Microbiota and Short-chain Fatty Acid During Lifespan of the Rat. Journal of Neurogastroenterology and Motility, 27(1), 134-146.

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SE-HPLC Analysis of Polio Vaccine

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SE-HPLC was performed with an Agilent 1100 series instrument (Agilent Technologies; catalog no. G1380-90000) equipped with a quaternary pump, a degasser, a temperature controlled autosampler, a UV/Vis diode array detector (DAD), and an Agilent 1200 series fluorescence detector (FLD). A TSKgel G6000PW_XL column (7.8 mm by 30 cm) or a TSKgel G3000SW_XL column (7.8 mm by 30 cm) purchased from Tosoh Bioscience (King of Prussia, PA) was used. At a flow rate of 0.8 ml/min, 100 µl of sample was injected per analysis. The mobile phase contained 50 mM sodium phosphate–140 mM NaCl at pH 6.7. The FLD was used as the primary detector and was set to acquire data at an excitation wavelength of 280 nm and an emission wavelength of 336 nm. For polio vaccine peak identification, the elution volume and the corresponding retention time of the FLD signal were calculated on the basis of a standard calibration curve. To calculate polio vaccine recovery, the area under the curve of formulations before lyophilization was compared to the area under the curve of formulations after lyophilization.

Shin W.J., Hara D., Gbormittah F., Chang H., Chang B.S, & Jung J.U. (2018). Development of Thermostable Lyophilized Sabin Inactivated Poliovirus Vaccine. mBio, 9(6), e02287-18.

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Biodistribution of MTX and LDH-MTX in Cervical Cancer

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The biodistribution of free MTX and LDH-MTX in the orthotopic cervical cancer model was compared by administering an ip injection of each drug PBS, MTX (60 mg/kg), or LDH-MTX (143 mg/kg, corresponding to 60 mg/kg MTX; n=3 per group), and the mice were sacrificed at precisely 30 and 60 minutes after the ip injection. The samples were prepared for the biodistribution study by harvesting the organs and tumor tissues, homogenizing with four times the volume of 1 M NaOH solution, and centrifugation at 3,000 rpm for 10 minutes. Two volumes of acetonitrile were added to the ground samples and were vigorously mixed to obtain the protein precipitates. After centrifugation at 10,000 rpm for 3 minutes, four volumes of chloroform were added to the supernatant in order to remove the lipids. Finally, the supernatants were collected via centrifugation at 2,000 rpm for 10 minutes and were analyzed via HPLC (Agilent 1100 series Instrument, Agilent Technologies, Santa Clara, CA, USA). The MTX content in the cells was also quantified via HPLC.

Choi G., Piao H., Alothman Z.A., Vinu A., Yun C.O, & Choy J.H. (2016). Anionic clay as the drug delivery vehicle: tumor targeting function of layered double hydroxide-methotrexate nanohybrid in C33A orthotopic cervical cancer model. International Journal of Nanomedicine, 11, 337-348.

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Biodistribution of Free MTX vs MTX-LDH

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To compare the biodistribution of free MTX and MTX-LDH, 2 × 10⁶ MCF7/mot cells were injected into the mammary pad of 7-week-old female athymic nu/nu mice (Orient bio, Korea). When the tumor volume reached approximately 300 mm³, PBS, LDH, MTX (60 mg/kg), or MTX-LDH (containing 60 mg/kg MTX) (n = 3 per group) was injected intraperitoneally (i.p.), and the mice were sacrificed immediately after 30 min, 1 hr, and 2 hrs. And the tumor and liver were harvested, homogenized, and dissolved in nitric acid. Two volumes of acetonitrile were added to the ground organ and mixed vigorously to precipitate proteins. After centrifugation at 10000 × g for 3 min, four volumes of chloroform were added to the supernatant to remove lipids. The supernatants were collected by centrifugation at 2000 × g for 10 min and analyzed by high-performance liquid chromatography (HPLC; Agilent 1100 series Instrument, USA). The MTX content in the cells was quantified by HPLC with an Agilent 1100 series HPLC system using a C18 column (Nucleosil, 250 × 4.6 mm, Alltech, Nicholasville, KY). The mobile phase was 0.05 M KH₂PO₄, 10% acetonitrile, and pH 6.6, and the flow rate was 1 mL/min, with a column temperature of 40°C. MTX was detected by the UV absorption band at 304 nm.

Choi G., Kwon O.J., Oh Y., Yun C.O, & Choy J.H. (2014). Inorganic Nanovehicle Targets Tumor in an Orthotopic Breast Cancer Model. Scientific Reports, 4, 4430.

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Microwave-Assisted Organic Synthesis Protocols

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All solvents and reagents were purchased from commercial sources and were of the highest grade available unless otherwise noted. Flash chromatography was performed with silica gel (230/400 mesh, Fisher Scientific) on Biotage Sp1 purification system. All anhydrous reactions were carried out under positive pressure of nitrogen or argon. All microwave reactions were carried out on Biotage microwave initiator 2.5 system with power range 0-400 W at 2.45 GHz. HPLC-MS analyses were performed on Agilent 1100 series instrument with Zorbax C18 reverse phase column unless otherwise noted. HRMS results were obtained on an apex-Qe instrument. All ¹H-NMR and ¹³C-NMR spectra were recorded on a Bruker 300 MHz or DRX 500 MHz NMR spectrometer, using deuterated solvents. The spectra are reported in ppm and referenced to deuterated DMSO (2.49 ppm for ¹H, 39.5 ppm for ¹³C) or referenced to deuterated chloroform (7.26 ppm for ¹H, 77 ppm for ¹³C).

Miranti C.K., Moore S., Kim Y., Chappeta V.R., Wu K., De B., Gokhale V., Hurley L.H, & Reyes-Reyes E.M. (2020). Nucleolin represses transcription of the androgen receptor gene through a G-quadruplex. Oncotarget, 11(19), 1758-1776.

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Quantification of Intestinal Short-Chain Fatty Acids

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Contents of ileum and cecum were extracted as described previously [15 (link)] and the concentrations of acetic acid and butyric acid in each extract were measured via high-performance liquid chromatography. Briefly, distilled water was added to 20 to 50 mg of each intestinal contents to a final weight of 200 mg. Following incubation at 80°C for 15 minutes, each sample was centrifuged at 13,000 rpm for 10 minutes, after which the supernatant was filtered through a membrane filter (pore size 0.45 μm). Acetic acid and butyric acid in each sample were then separated and measured using an Agilent 1100 series instrument (Agilent, Santa Clara, CA, USA) equipped with a C18 column (ZORBAX Eclipse XDB-C18, analytical 4.6 × 150 mm, 5-Micron; Agilent) and a UV detector (210 nm). The mobile phase consisted of 90% 10 mM KH₂PO₄ and 10% acetonitrile.

Choi S.I., Kim N., Lee S.M., Nam R.H., Kang S.R., Song C.H., Park Y.T., Min H., Kim Y.R, & Seok Y.J. (2019). Rat Intestinal Acetic Acid and Butyric acid and Effects of Age, Sex, and High-fat Diet on the Intestinal Levels in Rats. Journal of Cancer Prevention, 24(1), 20-25.

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Peptidoglycan Digestion Analysis by HPLC-MS

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HPLC-MS experiments were focused on the separation and identification of individual products of enzymatic digestion of the peptidoglycan. Chromatographic analyses were carried out on an Agilent 1100 series instrument (Agilent Technologies, Santa Clara CA, USA) coupled with a diode-array detector (Agilent Technologies, Santa Clara CA, USA) and an ion-trap mass spectrometer amaZon SL equipped with an ESI ion source (Bruker Daltonics, Fremont, CA, USA). The chromatographic separation was performed on Poroshell 120 SB-AQ column (150 × 2.1 mm; 2.7 μm) (Agilent Technologies, Santa Clara CA, USA) at 35 °C. The mobile phase consisted of eluent A: 0.1% formic acid in water and eluent B: 0.1% formic acid in methanol at a flow rate of 0.2 mL min⁻¹. The gradient started at 1% of B, increased at a constant rate until 70% of B in 20 min, then held constant until 25 min, and then back to the initial 1%, and held constant until 40 min. The injected volume was 2.5 µL. The MS conditions were as follows: spray voltage, −5.0 kV; the pressure of nebulizer gas, 25 psi; the flow of dry gas, 10 L/min; the temperature of dry gas, 300 °C. The measurements were performed in Ultrascan mode in the range of m/z 70–2200. Structural analyses were based on MS data obtained in negative ion mode.

Vacek L., Kobzová Š., Čmelík R., Pantůček R, & Janda L. (2020). Enzybiotics LYSSTAPH-S and LYSDERM-S as Potential Therapeutic Agents for Chronic MRSA Wound Infections. Antibiotics, 9(8), 519.

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