Chromatography HPLC fluorescence detectors

QF = quartz fiber, GF = glass fiber, PUF = polyurethane foam, FA = filter-adsorbent, FFA = filter-filter-adsorbent, DFA = denuder-filter-adsorbent, DDF = denuder-denuder-filter, DCM = dichloromethane, ACN = acetonitrile, HEX = n-hexane, GC-MS = gas chromatography-mass spectrometry, GC-ECD = gas chromatography-electron-capture detector, HPLC = high-performance liquid chromatography, FLD = fluorescence detector, SFE = supercritical fluid extraction, EA = electrostatic precipitator. 

Milbemectin consists of two active ingredients, M.A3 and M.A4. Milbemectin is extracted from plant materials and soils with methanol-water (7 3, v/v). After centrifugation, the extracts obtained are diluted to volume with the extraction solvent in a volumetric flask. Aliquots of the extracts are transferred on to a previously conditioned Cl8 solid-phase extraction (SPE) column. Milbemectin is eluted with methanol after washing the column with aqueous methanol. The eluate is evaporated to dryness and the residual milbemectin is converted to fluorescent anhydride derivatives after treatment with trifluoroacetic anhydride in 0.5 M triethylamine in benzene solution. The anhydride derivatives of M.A3 and M.A4 possess fluorescent sensitivity. The derivatized samples are dissolved in methanol and injected into a high-performance liquid chromatography (HPLC) system equipped with a fluorescence detector for quantitative determination. 

As seen in Chapter 9.C.2, a very wide variety of organics are found in particles in ambient air and in laboratory model systems. The most common means of identification and measurement of these species is mass spectrometiy (MS), combined with either thermal separation or solvent extraction and gas chromatographic separation combined with mass spectrometry and/or flame ionization detection. For larger, low-volatility organics, high-performance liquid chromatography (HPLC) is used, combined with various detectors such as absorption, fluorescence, and mass spectrometry. For applications of HPLC to the separation, detection, and measurement of polycyclic aromatic hydrocarbons, see Wingen et al. (1998) and references therein. 

Currently, high-performance liquid chromatography (HPLC) methods have been widely used in the analysis of tocopherols and tocotrienols in food and nutrition areas. Each form of tocopherol and tocotrienol can be separated and quantified individually using HPLC with either a UV or fluorescence detector. The interferences are largely reduced after separation by HPLC. Therefore, the sensitivity and specificity of HPLC methods are much higher than those obtained with the colorimetric, polarimetric, and GC methods. Also, sample preparation in the HPLC methods is simpler and more efficiently duplicated than in the older methods. Many HPLC methods for the quantification of tocopherols and tocotrienols in various foods and biological samples have been reported. Method number 992.03 of the AOAC International Official Methods of Analysis provides an HPLC method to determine vitamin E in milk-based infant formula. It could probably be said that HPLC methods have become dominant in the analysis of tocopherols and tocotrienols. Therefore, the analytical protocols for tocopherols and tocotrienols in this unit are focused on HPLC methods. Normal and reversed-phase HPLC methods are discussed in the separation and quantification of tocopherols and tocotrienols (see Basic Protocol). Sample... 

Direct determination of urea pesticides by high-performance liquid chromatography has been widely reported in the literature (10,32-36,127-130). Ultraviolet detection has often been used (32,33,35,36,60,127) with usually acceptable sensitivity, although this detector is nonspecific and the sensibility is, in general, low. To overcome this problem, several techniques have been assayed, such as precolumn enrichment (60), postcolumn derivatization (34,10), and the use of other detection techniques such as the electrochemical (129), photoconductivity (128,130), and fluorescence detectors (9,10,34). Table 9 summarizes representative papers using these techniques in HPLC analysis. 

Notes LOD, limit of detection MeOH, methanol EtOH, ethanol ACN, acetonitrile MTBE, methyl tert-butyl ether DCM, dichloromethane THF, tetrahy-drofuran KOH, potassium hydroxide SFE, supercritical fluid extraction MS, mass spectrometry HPLC, high-performance liquid chromatography DAD, diode array detector PDA, photodiode array detector FD, fluorescence detector ECD, electrochemical detector ESI, electrospray ionization APCI, atmosphere pressure chemical ionization TLC, thin layer chromatography FAB, fast atom bombardment NMR, nuclear magnetic resonance BHT, butylated hydroxytoluene SPE, solid phase extraction. 

The concentrates are analysed by High Performance Liquid Chromatography (HPLC) coupled to a fluorescence detector. Elution is performed using a gradient of methanol in a phosphate buffer on a Cj g radial compression column. In order to render the amino acids amenable to fluorescence detection ( X Xgjjj = 455 nm), it is first necessary to derivatize them using orthophtaldialdehyde (O.P.A.) in the presence of mercapto-2 ethanol. Up to 20 amino acids can thus be analysed and quantified, and the complete analysis takes only 35 minutes. Table 2 presents the most commonly determined amino acids during the course of this study. 

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