The analytical method has been reported previously.8 (link) The program EZchrom Elite (VWR) was used for data requisition and analysis.
Ezchrom elite
Manufactured by Avantor
Sourced in Germany
EZchrom Elite is a data acquisition and analysis software for chromatography. It provides an interface for collecting and processing data from various chromatography instruments.
Automatically generated - may contain errors
Lab products found in correlation
L 2130, by Avantor (1 mentions)
L 2455, by Avantor (1 mentions)
Chromabond sb, by Macherey-Nagel (1 mentions)
L 2200 autosampler, by Hitachi (1 mentions)
L 2130 hta pumps, by Hitachi (1 mentions)
L 2450 diode array detector, by Hitachi (1 mentions)
Lichrocart, by Merck Group (1 mentions)
Gemini c18, by Phenomenex (1 mentions)
5160 pump, by Avantor (1 mentions)
5260 autosampler, by Avantor (1 mentions)
5310 column oven, by Avantor (1 mentions)
Gemini c18 guard column, by Phenomenex (1 mentions)
L 2200 autosampler, by Avantor (1 mentions)
L 2450 diode array detector, by Avantor (1 mentions)
6 protocols using ezchrom elite
1
Analytical Method for Data Analysis
2
Stability Studies Apparatus and Methods
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The following apparatuses were used for the stability studies:
The High-Performance Liquid Chromatography (HPLC) system consisted of an ELITE LaChromVWR/ Hitachi plus autosampler, a VWR photodiode array detector L- 2455, and a VWR L-2130 HPLC pump. Data were acquired and integrated using EZChrom Elite (VWR, Agilent).
pH meter (Bioblock Scientific model 93313).
PAMAS particle counter, Rutesheim, Germany.
3
Simultaneous HPLC Determination of Nitrite and Nitrate
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Two hundred microliters of plasma (baseline and standing samples) were diluted with double-distilled water (1:2, vol/vol) and loaded on pre-conditioned anion exchange columns (Chromabond SB, Macherey-Nagel, Düren, Germany). After a washing step with double-distilled water nitrite and nitrate were eluted with 1 mL 0.5 mol/L sodium chloride. In the eluates, nitrite and nitrate were determined simultaneously by means of HPLC analysis according to a previously published method (Romitelli et al., 2007 (link)) but with some modifications. Briefly, the HPLC consisted of a L-2200 autosampler, two L-2130 HTA pumps, and a L-2450 diode array detector (all: VWR Hitachi, VWR, Vienna, Austria). Separation was performed on a Hypersil ODS column (5 μm; 250 × 4 mm I.D.) with 10.0 min isocratic elution (buffer A: 0.1 mol/L NaH2PO4, pH = 5.5, containing 5.9 mmol/L tetrabutylammonium hydrogensulphate) followed by a linear gradient to 20% buffer B (buffer B: 0.1 mol/L NaH2PO4, pH = 5.5, containing 5.9 mmol/L tetrabutylammonium hydrogensulphate/acetonitrile, 3:1, vol/vol) within another 10 min. The injection volume of standard and sample solutions was 40 μL. The absorbance at 205 nm was recorded. Data acquisition and subsequent analysis was done with the EZchrom Elite (VWR) program. Retention time was ~7.80 min for nitrite and ~14.5 min for nitrate.
4
Phenolic Profiling of Cactus Seed Oil
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The analysis of phenolic compounds extracted from cold pressed cactus seed oil was performed using a HPLC-DAD system (VWR, Hitachi, Darmstadt, Germany), equipped with a reversed phase C18 column (250 × 4 mm, i.d. 5 μm, LichroCART, Lichrospher, Merck, Darmstadt, Germany). During the analysis, the column temperature was set to 40 °C. The mobile phase degassed by ultrasonic treatment was A: water/phosphoric acid (99.5/0.5, (v/v)) and B: methanol/acetonitrile (1/1, (v/v)). The composition of the gradient was: 5% of B at the beginning (0 min) and then changed to obtain 30%, 38%, 45%, 52.5%, 100% and 5% B at 15, 30, 40, 45, 50, and 60.1 min, respectively, in a total run time of 65 min.
The flow rate was 1 mL/min and the injection volume was 20 μL. The detection was conducted on a diode array detector L-2455 (Merck-Hitachi, Darmstadt, Germany) at the wavelengths 280, 310, and 335 nm. EZ Chrom Elite (VWR International GmbH, Darmstadt, Germany) was used as software for the acquisition and evaluation of the data.
The flow rate was 1 mL/min and the injection volume was 20 μL. The detection was conducted on a diode array detector L-2455 (Merck-Hitachi, Darmstadt, Germany) at the wavelengths 280, 310, and 335 nm. EZ Chrom Elite (VWR International GmbH, Darmstadt, Germany) was used as software for the acquisition and evaluation of the data.
5
Analysis of Secondary Metabolites via HPLC-DAD
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For analyses of the SMs, strains were grown in submerged cultures as described above. After 7 days, mycelia were removed from the culture by filtration through Mirachloth (Calbiochem, Merck KGaA, Darmstadt, Germany). Small particulates were removed from the culture filtrates using 0.45 μm syringe filters (BGB®, Schloßböckelheim, Germany) which were then analyzed by high-pressure liquid chromatography with a diode array detector [HPLC-DAD; Hitachi Chromaster LC equipped with a 250 mm × 4.60 mm i.d., 5 μm, Gemini® C18 with a 4 mm × 3 mm Gemini® C18 guard column (Phenomenex, Aschaffenburg, Germany)], a with 5160 pump, 5260 autosampler, 5310 column oven, and 5430 DAD (VWR International GmbH, Darmstadt, Germany). For data analyses, the software EZChrom Elite (VWR, Darmstadt, Germany) was used.
6
Quantitative Assessment of Platelet Aggregation
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The aggregability of platelets can be assessed by quantitative determination of ATP exocytosis [21 (link)]. WB samples containing low, physiological or high levels of albumin were centrifuged at 150 g for 12 min in order to obtain platelet rich plasma (PRP) samples. Platelet aggregation was induced by addition of collagen (2 μg mL-1, final concentration) to 500 μL of the PRP samples. After two minutes, the PRP samples were centrifuged at 1,500 g for two minutes and proteins in the supernatant were precipitated with 0.4 M perchloric acid. After centrifugation at 12,000 g 100 μL of the supernatant were neutralized by addition of 10–12 μL of 2 M K2CO3 at 4°C. The supernatant obtained after centrifugation was used for HPLC analysis (injection volume: 40 μL). Separation of adenine nucleotides was performed on a Hypersil ODS column (5 μm, 250 × 4 mm I.D., equipped with a precolumn; Thermo Electron Corp. Runcorn, Cheshire, UK) using a L-2200 autosampler, two L-2130 HTA pumps, and a L-2450 diode array detector (all VWR International, West Chester, PA, USA) as previously described [22 (link)]. Detector signals (absorbance at 254 nm) were recorded and the program EZchrom Elite (VWR) was used for data acquisition and analysis.
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