All moisture-sensitive reactions were carried out in an oven dried flask under argon atmosphere. All chemicals and reagents were purchased from commercial suppliers and used without further purification. Anhydrous solvents were obtained as follows: anhydrous tetrahydrofuran, diethyl ether, and benzene were distilled from sodium metal under argon. Anhydrous dichloromethane, toluene, methanol, and acetonitrile were dried via distillation from CaH2 under argon. All other solvents were HPLC grade. 1H NMR and 13C NMR spectra were recorded on Varian INOVA300-1, Bruker Avance ARX-400 and Bruker DRX-500 spectrometers. NMR data are reported as: δ value (chemical shift, J-value (Hz), integration, where s=singlet, d=doublet, t=triplet, q=quartet, br= broad). Low resolution mass analyses were performed on a Agilent 1290 Infinity II spectrometer. High-resolution mass spectrometric analyses were performed at the Purdue University Campus-wide Mass Spectrometry Center. TLC analysis was carried out with SiliCycle 60A-F254 plates. Flash chromatography was performed using SiliCycle 230–400 mesh, 60 Å pore diameter silica gel. HPLC analysis was performed on a Agilent 1260 Infinity instrument. All test inhibitors showed purity >95% by HPLC analysis.
Agilent 1260 infinity instrument
Manufactured by Agilent Technologies
Sourced in Germany, United States
The Agilent 1260 Infinity instrument is a high-performance liquid chromatography (HPLC) system designed for a wide range of analytical applications. It provides reliable and accurate separation and detection of various chemical compounds.
Automatically generated - may contain errors
Lab products found in correlation
Inova300 1, by Bruker (1 mentions)
Avance arx 400, by Bruker (1 mentions)
Drx 500, by Bruker (1 mentions)
Millex gv, by Merck Group (1 mentions)
Xbridge c18 column, by Waters Corporation (1 mentions)
Nanospect ct system, by Bioscan (1 mentions)
Jupiter proteo c12, by Phenomenex (1 mentions)
Astra version 6, by Wyatt Technology (1 mentions)
5 protocols using agilent 1260 infinity instrument
1
Moisture-Sensitive Organic Synthesis Protocols
2
RP-HPLC-MS Analysis of Organic Compounds
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RP-HPLC-MS analysis was performed on an Agilent 1260 Infinity instrument (Agilent Technologies, Waldbronn, Germany) coupled to a variable wavelength detector (VWD) (set to 214 nm) and a 6120 Agilent Quadrupole MS containing an electrospray ionisation (ESI) source operating in positive ionization mode in a m/z range of 200 to 2000. As HPLC column a Poroshell 120 EC-C18 1.8 µm (3.0 × 50 mm, 2.5 µm) RP column from Agilent was used. The mobile phases A and B were 95/5 H2O/MeCN, 0.1% formic acid (A) and 5/95 H2O/MeCN, 0.1% formic acid (B). Samples were analyzed at a flow rate of 0.4 mL/min using a linear gradient from 100% A to 50% B in a time range of 30 min at 25 °C. UV and MS spectral analysis were performed within the OpenLab ChemStation software for LC/MS from Agilent Technologies (Waldbronn, Germany).
3
Peptide Stability Following Radiotherapeutic Administration
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Blood samples were collected in polypropylene tubes containing ethylenediaminetetraacetic acid at 5, 15, 30 and 60 min after administration of 177Lu-PP-F11N. The tubes were placed on ice. Plasma was separated by centrifugation (4000 g for 10 min) and protein precipitation followed using cold methanol (v/v 1/2). The mixture was stirred to favor precipitation of proteins and then centrifuged at 4000 g for 10 min. The supernatant was collected and filtered through a 0.22 µM filter (Millex®-GV, Merck Millipore, Bedford, MA, USA). The clear solution was diluted with H2O (1:1) and then analyzed by radio-high performance liquid chromatography (HPLC) to determine the percentage of intact peptide over time. HPLCs were performed on the Agilent 1260 infinity instrument (Agilent, Santa Clara, United States) connected to a GABI radioactivity-HPLC-flow-monitor γ-spectrometer (Elysia-raytest, Straubenhardt, Germany). Radioligands were analyzed using Phenomenex Jupiter Proteo C12 (90 Å, 250 × 4.6 mm) column (Phenomenex, Torrance, USA) using the gradient 5–50% B in 15 min (A = H2O [0.1% trifluoroacetic acid], B = ACN [0.1% trifluoroacetic acid]) with a flow rate of 2 mL/min.
4
Radiolabeled Peptide Characterization
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AMD3100 was purchased from Toronto
Research Chemicals (Canada).
All other reagents and solvents were purchased from Acros Organics
(Belgium) and Merck (Darmstadt, Germany) and used without further
purification. DOTA-tris(tBu)ester was purchased from CheMatech (France).
Purity of the peptides was >95% as assessed by liquid chromatography
mass spectrometry (LC-MS) on a LCMS-2020 SHIMADZU (Japan) system equipped
with a Waters XBridge C-18 column (4.6 mm × 150 mm, 5 μm
particle size). The gradient used was 15–65% solvent B in 15
min (A = H2O [0.1%TFA], B = ACN [0.1% TFA]) at a flow rate of 1.0 mL/min. Radio-HPLC was performed
on an Agilent 1260 infinity instrument (Agilent) connected to a GABI
radioactivity-HPLC-flow-monitor γ-spectrometer (Elysia-raytest,
Germany). Radioligands were analyzed using Phenomenex Jupiter Proteo
C12 (90 Å, 250 mm × 4.6 mm) column using the gradient 5–50% B in 15 min (A = H2O [0.1%TFA], B = ACN [0.1% TFA]) with a flow rate of 2 mL/min. Quantitative
γ-counting was carried out on a COBRA 5003 γ-system well
counter from Packard Instruments. SPECT/CT images were acquired using
a dedicated nanoSPECT/CT system (Bioscan, Mediso, Hungary).
Research Chemicals (Canada).
All other reagents and solvents were purchased from Acros Organics
(Belgium) and Merck (Darmstadt, Germany) and used without further
purification. DOTA-tris(tBu)ester was purchased from CheMatech (France).
Purity of the peptides was >95% as assessed by liquid chromatography
mass spectrometry (LC-MS) on a LCMS-2020 SHIMADZU (Japan) system equipped
with a Waters XBridge C-18 column (4.6 mm × 150 mm, 5 μm
particle size). The gradient used was 15–65% solvent B in 15
min (A = H2O [0.1%TFA], B = ACN [0.1% TFA]) at a flow rate of 1.0 mL/min. Radio-HPLC was performed
on an Agilent 1260 infinity instrument (Agilent) connected to a GABI
radioactivity-HPLC-flow-monitor γ-spectrometer (Elysia-raytest,
Germany). Radioligands were analyzed using Phenomenex Jupiter Proteo
C12 (90 Å, 250 mm × 4.6 mm) column using the gradient 5–50% B in 15 min (A = H2O [0.1%TFA], B = ACN [0.1% TFA]) with a flow rate of 2 mL/min. Quantitative
γ-counting was carried out on a COBRA 5003 γ-system well
counter from Packard Instruments. SPECT/CT images were acquired using
a dedicated nanoSPECT/CT system (Bioscan, Mediso, Hungary).
5
Gel Permeation Chromatography Analysis
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GPC analysis was performed using an Agilent 1260 Infinity instrument (Agilent Technologies, Santa Clara, CA, USA) equipped with a double detector with the light scattering configuration. Two mixed C columns at 35 °C were employed, using tetrahydrofuran (THF) as the mobile phase with a flow rate of 1 mL/min. GPC samples were prepared in HPLC-grade THF and filtered before injection. The analysis was carried out using ASTRA version 6.1 (Wyatt Technology Corporation, Santa Barbara, CA, USA) software. The number and weight average molecular weight (Mn and Mw, respectively) and polydispersity (Ð) were calculated using narrow standards of PMMA for the calibration curve. The GPC standards used were purchased from Agilent as the InfinityLab EasiVial PMMA pre-weighed calibration kit with samples covering the range from Mn ~600 to ~2,000,000.
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