Chromatogram detection

Some analytical instruments produce a table of raw data which need to be processed into the analytical result. Hyphenated measurement devices, such as HPLC linked to a diode array detector (DAD), form an important class of such instruments. In the particular case of HPLC-DAD, data tables are obtained consisting of spectra measured at several elution times. The rows represent the spectra and the columns are chromatograms detected at a particular wavelength. Consequently, rows and columns of the data table have a physical meaning. Because the data table X can be considered to be a product of a matrix C containing the concentration profiles and a matrix S containing the pure (but often unknown) spectra, we call such a table bilinear. The order of the rows in this data table corresponds to the order of the elution of the compounds from the analytical column. Each row corresponds to a particular elution time. Such bilinear data tables are therefore called ordered data tables. Trilinear data tables are obtained from LC-detectors which produce a matrix of data at any instance during the... 

Fig. 3.90. HPLC chromatograms (detection wavelength 225 nm) of the reaction products of RB15 (left, a-c) and RB38 (right, d-f). (a,d) Initial dye solutions (50 mg/1) (b,c) after incubation with MnP (c,f) after incubation of 200 mg/1 dye in cultures of B. adusta. The parent compounds RB15 and RB38 are not detected under these chromatographic conditions. Reprinted with permission from A. Heinfling-Weidtmann et al. [153].

A gSe of two Waters ultrastyragel columns, designated 10 A and 10 A and a Waters pump (Model 590) for HPLC were used in this study. The elution solvent was tetrahydrofuran (THE) which was distilled in the presence of a small amount of CaH in order to remove the peroxide. The flow rate was maintained at 1 ml/min. The sample injection volume was -30 pi. The chromatogram detected by the differential refractometer (Waters R401) was recorded on a strip chart recorder. All experiments were performed at room temperatures with concentrations below the over-loading condition. 

SIM with three compound specific ions can be applied, as for instance demonstrated by Wang and Budde [101] for sixteen carbamate, urea and thioiuea pesticides and herbicides, by Rodriguez and Orescan [51] for selected sulfonylurea, imidazolinone, and sulfonamide herbicides, and by Yu et al. [17] for 52 carbamates, thiocaibamates, and phenylureas. In the latter case, computer-controlled optimization of the MS measurement conditions is performed. The most prominent ion, either a protonated or an ammoniated molecule, is used for quantitation. The dwell time for each ion in SIM was 0.02 s with an inter-charmel delay of 0.02 s, i.e., 2 s are needed for each data point in the chromatogram. Detection limits ranged from 0.09 to 19 pg/1 with 50-pl injections. 

Fig. 3 HPTLC bioluminescence image of the same plate as in Fig. 2. Note that the different numbers of zones in the chromatograms detected in some of the lanes by the two methods provide complementary information on the samples. Source Photograph supplied by Camag, Muttenz, Switzerland.

Campbell, A. Chejlava, M. Sherma, J. Use of a modified flatbed scanner for documentation and quantification of thin layer chromatograms detected by fluorescence quenching. J. Planar Chromatogr.-Mod. TLC 2001, 16 (3), 244-246. 

Figure 3.52 Isocratic separation of common inorganic anions on lonPac AS18-4pm at different flow rates. Column dimensions 150 mm X 0.4 mm i.d. column temperature 30 °C eluent 30 mmol/L KOH (EG) flow rate see chromatograms detection suppressed...

Fig. 3. Combined headspace sampiing, themnai desorption and purge and trap injection system with exampie headspace chromatogram, (a) injection system (b) Headspace chromatogram. Detection of solvents In blood sample by headspace analysis as part of an Industrial hygiene study. Sample held at 60°C. Column UCON LB 550, 25 m, at 40°C. Produced tom D.W. Grant, Capillary Gas Chromatography, 1996. John Wiiey Sons Ltd. Reproduced with permission.

The chromatogram can finally be used as the series of bands or zones of components or the components can be eluted successively and then detected by various means (e.g. thermal conductivity, flame ionization, electron capture detectors, or the bands can be examined chemically). If the detection is non-destructive, preparative scale chromatography can separate measurable and useful quantities of components. The final detection stage can be coupled to a mass spectrometer (GCMS) and to a computer for final identification. 

This experiment focuses on developing an HPLG separation capable of distinguishing acetylsalicylic acid, paracetamol, salicylamide, caffeine, and phenacetin. A Gjg column and UV detection are used to obtain chromatograms. Solvent parameters used to optimize the separation include the pH of the buffered aqueous mobile phase, the %v/v methanol added to the aqueous mobile phase, and the use of tetrabutylammonium phosphate as an ion-pairing reagent. 

Schematic diagram showing the injection of a mixture of four substances (A, B, C, D) onto a GC column, foliowed by their separation into individuai components, their detection, and the dispiay (gas chromatogram) of the separated materiais emerging at different times from the coiumn.

Further, peak overlap results in nonlinear detector response vs concentration. Therefore, some other detection method must be used in conjunction with either of these types of detection. Nevertheless, as can be seen from Figure Ilf, chiroptical detection can be advantageous if there is considerable overlap of the two peaks. In this case, chiroptical detection may reveal that the lea ding and tailing edges of the peak are enantiomerically enriched which may not be apparent from the chromatogram obtained with nonchiroptical detection (Fig. He). 

Fig. 11. Simulated chromatograms of chiral separations obtained using nonchiroptical detection (a, c, e) and chiroptical detection (b, d, f) illustrating the...

Fig. 3. (a) Flame ionization detector (fid) response to an extract of commercially processed Valencia orange juice, (b) Gas chromatography—olfactometry (geo) chromatogram of the same extract. The abscissa in both chromatograms is a normal paraffin retention index scale ranging between hexane and octadecane (Kovats index). Dilution value in the geo is the -fold that the extract had to be diluted until odor was no longer detectable at each index. 

Aroclor 1248, Aroclor 1254, and Aroclor 1260. Quantitation is by comparison of chromatograms with standard concentrations of pure compounds treated in an identical manner. The phenoxy acid herbicides (2,4-dichlorophenoxy)acetic acid (2,4-D), sUvex, and (2,4,5-trichlorophenoxy)acetic acid (2,4,5-T) can be deterrnined by electron-capture detection after extraction and conversion to the methyl esters with BF.-methanol. The water sample must be acidified to pH <2 prior to extraction with chloroform. 

HPLC method with amperometric detection was applied for detenuination of phenols in sea sediment and some dmg preparation. Peaks of phenol, guaiacol, cresols, hydroquinon and resorcinol were identified on chromatogram of birch tai. The HPLC method with electrochemical detectors was used for detenuination of some drug prepai ation of aminophenol derivate. So p-acetaminophenol (paracetamol) was determined in some drug. 

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