Chromatography HPLC diode array detector

Comparison of Retention Times (Minutes) for Lignan Aglycons by High-Performance Liquid Chromatography (HPLC) Diode Array Detector (DAD) with Those of the Lignan Trimethylsilyl Ether (TMS) Derivatives by Gas Chromatography-Mass Spectrometry (CC-MS)... 

Notes LOD, limit of detection MeOH, methanol EtOH, ethanol ACN, acetonitrile EtAC, ethyl acetate SPE, solid phase extraction HLB (hydrophilic lipophilic balanced) TFA, trifluoroacetic acid GC, gas chromatography TMS, trimethylsilyl MS, mass spectrometry HPLC, high-performance liquid chromatography DAD, diode array detector NMR, nuclear magnetic resonance ESI, electrospray ionization APCI, atmospheric pressure chemical ionization CE, capillary electrophoresis ECD, electrochemical detector CD, conductivity detector TLC, thin layer chromatography PDA, photodiode array detector. 

Note A amperometry BI bead injection CL chemiluminescence F fluorescence FAT flow analysis technique HPLC/DAD high-performance liquid chromatography with diode array detector LOD detection limit MCFIA multicommutated flow injection analysis MPFS multipumping flow systems MSFIA multisyringe FIA P potentiometry RSD relative standard deviation SF sampling frequency SFA stopped-flow analysis SP spectrophotometry. 

FIGURE 13.9 The HPLC diode array UV absorbance detector. When a mixture component elutes from the column, not only the chromatography peak but the entire UV absorption spectrum for that component can be recorded. 

Liquid chromatography/mass spectrometry analyses were performed with an ion trap mass spectrometer (LCQ, Thermo Fisher Scientific Inc., MA) equipped with an HPLC system (Agilent, CA Model 1100) connected with a diode-array detector (DAD, G1315A). The sample solution (1-5 p,L) was applied on an Inertsil ODS-3 column (2.1 x 150 mm, 3 p,m, GL... 

High-Performance Liquid Chromatography. A Varian 5060 delivery system was used for this work with detection by UV absorption. Either a Varian UV-50 variable wavelength detector or a Hewlett Packard 1040A scanning diode array detector was used. All HPLC columns were packed in our laboratory (10) with 5-/um particle size Spherisorb-ODS, Spherisorb-CN (Phase Separations), or 8-pm particle size Zorbax-CN (Dupont Ltd). HPLC columns (20 or 25 cm X 4.6 mm i.d.) were coupled via short lengths of stainless steel capillary tubing (5 cm X 0.25 mm i.d.). Separation conditions were as follows ... 

J,m poly(tetrafluoroethylene) (PTFE) syringe-tip filters High-performance liquid chromatography (HPLC) system equipped with Quaternary pump Diode array detector Vacuum degasser... 

Because polyphenolics show chemical complexities and similar structures, isolation and quantification of the individual polyphenolic compounds have been challenging. Many traditional techniques (paper chromatography, thin-layer chromatography, column chromatography) have been used. HPLC, with its merits of exacting resolution, ease of use, and short analysis time, has the further advantage that separation and quantification occur simultaneously. A reversed-phase HPLC apparatus equipped with a diode array detector makes possible the easy isolation and separation of many polyphenolics. For enhanced performance of HPLC separation, the polyphenolics should first be isolated into several fractions to effectively separate the individual polyphenolics (Jaworski and Lee, 1987 Oszmianski and Lee, 1990). 

Ostermeyer (87) described the rapid and sensitive determination of acesulfame, saccharin, aspartame, SA, and BA in fishery products. Aspartame is determined directly in the aqueous extract of samples. For the other additives, cleanup of the extract is performed by anion-exchange chromatography. Ostermeyer used a Nucleosil 100-5 C lg (Macherey-Nagel) (250 X 4 mm, 5 /xm ) column and 0.02 mol/L KH2P04 acetonitrile as mobile phase and 219 nm detection using a diode array detector. Recoveries of spiked samples at concentrations of 50-400 mg sweetener/kg and 50-4000 mg preservative/kg were in the range 84-102% for all analyzed substances (87). The HPLC conditions for the determination of OAs are summarized in Table 5. 

Spectroscopic detectors measure partial or complete energy absorption, energy emission, or mass spectra in real-time as analytes are separated on a chromatography column. Spectroscopic data provide the strongest evidence to support the identifications of analytes. However, depending on the spectroscopic technique, other method attributes such as sensitivity and peak area measurement accuracy may be reduced compared to some nonselective and selective detectors. The mass spectrometer and Fourier transform infrared spectrometer are examples of spectroscopic detectors used online with GC and HPLC. The diode array detector, which can measure the UV-VIS spectra of eluting analytes is a... 

Reversed-phase high-performance liquid chromatography (RP-HPLC) is the usual method of choice for the separation of anthocyanins combined with an ultraviolet-visible (UV-Vis) or diode-array detector (DAD)(Hebrero et al., 1988 Hong et ah, 1990). With reversed-phase columns the elution pattern of anthocyanins is mainly dependent on the partition coefficients between the mobile phase and the Cjg stationary phase, and on the polarity of the analytes. The mobile phase consists normally of an aqueous solvent (water/carboxylic acid) and an organic solvent (methanol or acetonitrile/carboxylic acid). Typically the amount of carboxylic acid has been up to 10%, but with the addition of a mass spectrometer as a detector, the amount of acid has been decreased to as low as 1 % with a shift from trifluoroacetic acid to formic acid to prevent quenching of the ionization process that may occur with trifluoroacetic acid. The acidic media allows for the complete displacement of the equilibrium to the fiavylium cation, resulting in better resolution and a characteristic absorbance between 515 and 540 nm. HPLC separation methods, combined with electrochemical or DAD, are effective tools for anthocyanin analysis. The weakness of these detection methods is a lack of structural information and some nonspecificity leading to misattribution of peaks, particularly with electrochemical... 

Among the three advantages that diode array detector provides to the high-performance liquid chromatography (HPLC) analyst, peak identification is the one that allows the identification of unknown constituents in a sample whose matrix is complex. 

High performance liquid chromatography (HPLC). All HPLC analyses were performed with a model HP1090 analytical HPLC equipped with a diode array detector (Hewlett Packard, Mountain, View, CA). A Protein PAK 3000 SW column (Waters, Bedford, MA) was used for each size exclusion experiment. The column was equilibrated with ten column volumes (100 ml) of elution buffer (2.0 M GuHCl, 50 mM Tris sulfate, 5 mM EDTA, pH 7.5) prior to operation. A sample volume of 25 gl was applied to the column and eluted at a flow rate of 1.0 ml/min to facilitate rapid separation. For equilibrium experiments, each sample was equilibrated for three to eight hours before column separation. 

GC—gas chromatography HCA—hierarchical cluster analysis HELP—heuristic evolving latent projections HHM—horse heart myoglobin HPLC—high performance liquid chromatography HPLC-DAD—high performance liquid chromatography-diode array detector... 

The prepared solutions were monitored by liquid chromatography coupled with a diode array detector (DAD) and with an electrospray mass spectrometry (ESI-MS) detector. Isolation of the formed pigments was achieved by HPLC at the semipreparative scale. 

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