Analytical HPLC was performed using a LaChrom HPLC system by Merck (Darmstadt, Germany), consisting of an L-7100 pump (4.0 ml/min), an L-7200 autosampler, and an L-7400 UV-detector (254 nm) using a Chromolith SpeedROD RP-18 column (4.6 × 50 mm); gradient elution, 0 to 100 % solvent B (MeCN-H2O-TFA 90:10:0.1) in a 1:1 mixture of H2O-MeCN over 3.5 min. Data were acquired and processed using the EZChromElite Software by Hitachi.
Lachrom hplc system
Manufactured by Merck Group
Sourced in Germany
The LaChrom HPLC system is a versatile high-performance liquid chromatography instrument designed for analytical and preparative separation of a wide range of compounds. The system features a robust and reliable design, offering precise control of flow rates, solvent gradients, and temperature, ensuring accurate and reproducible results. The LaChrom HPLC system is a core laboratory tool for the separation, identification, and quantification of various chemical and biological samples.
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4 protocols using lachrom hplc system
1
Analytical HPLC of Organic Compounds
2
HPLC Analysis of Enantiomeric Alcohol Products
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HPLC analysis was performed using a Merck-Hitachi LaChrom HPLC system equipped with a Merck L-7490 RI detector, an L-7400 UV-detector, a reversed phase Chiralpak® AD-RH column (from Daicel, obtained at Sigma Aldrich, Vienna, Austria) and a thermostat column oven (40 °C). The mobile phase was composed of 25% acetonitrile in ddH2O at a flowrate of 30 mL/min. HPLC standard curves were prepared using racemic product at 0.1, 0.5, 1, 5, and 10 mM concentrations. Peak areas at corresponding retention times were used to calculate the concentrations. The enantiomeric excess of the major product (S)-2-phenylpropanol was calculated using the formula ee = (S-alcohol – R-alcohol)/(S-alcohol + R-alcohol). The retention times and chromatograms of authentic standards (main products ((S)-2-phenylpropanol, (R)-2-phenylpropanol), by-products (acetophenone, phenylethanol) were summarized in the Additional file 1 : Sect. 4. The aldehydes gave too broad peaks on HPLC and were quantified by GC-FID.
3
Quantifying Docetaxel Loading in Nanoparticles
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The amount of loaded DTX was directly quantified by high-performance liquid chromatography (HPLC) in order to determine the association efficiency (AE) and loading degree (LD) of the developed formulations by dissolving the nanoparticles in DMF. The quantification of DTX was performed using a LaChrom ® HPLC system (Merck, US) with a Symmetry reverse-phase C18 column (5 μm, 125 × 4 mm; Milford, US) and a LiChrospher 100 RP-18 guard column (Merck, US). The HPLC method employed a gradient elution with a flow rate of 1 mL/min. The mobile phase composition varied as follows: 10:90 ACN: H 2 O from 0 to 5 min, 60:40 ACN: H 2 O from 5 to 14 min, and 10:90 ACN: H 2 O from 14 to 17 min. The drug was retained at around 9.60 min. 50 µL of each sample was injected and the detection was performed at 227 nm using a UV detector [23] . The AE (%) was determined as a percentage of the experimental drug payload to the theoretical drug payload. DL (%) was calculated as a percentage of the quantified DTX mass to the total mass of the nanoparticles.
4
Cellodextrins Analysis by TLC and HPLC
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The cellodextrins were analyzed by thin‐layer chromatography (TLC). A mobile phase of ethyl acetate, acetic acid, and water (50:25:25, by volume) was used. Staining was with thymol reagent (thymol/ethanol/H2SO4, 0.5:95:5, w/v/v) at 95°C. The cellodextrins were additionally analyzed by high‐performance liquid chromatography (HPLC) on a Hitachi LaChrom HPLC system (Merck, Darmstadt, Germany) using a Luna 5 µm NH2 column (100 Å, 250 × 4.6 mm; Phenomenex, Aschaffenburg, Germany) operated at 40°C. Acetonitrile‐water (67.5:32.5, by volume) was used as eluent at a flow rate of 1.5 ml/min. Cellobiose was analyzed by HPLC using an Aminex HPX‐87H Column (300 × 7.8 mm; Bio‐Rad Laboratories, Vienna, Austria) operated at 60°C. Sulfuric acid (5 mM) was used as eluent at a flow rate of 0.5 ml/min. Refractive index detection was used to quantitate cellodextrins and cellobiose. Calibration was with authentic standards (CarboSynth). αGlc1‐P was determined enzymatically using a continuous coupled enzyme assay (Eis & Nidetzky, 1999 ).
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