Control Densitometer

In practice, the majority of TLC separations are qualitative or semiquantitative (visual comparison) in nature. However, modern computer-controlled densitometers are now available that scan sample and calibrator chromatograms in tracks on HPTLC plates and provide quantitative capabilities. Clinically relevant analytes that have been measured by TLC include amino acids, bile acids, carbohydrates, drugs, fipids, glycolipids, phospholipids, porphyrins, prostaglandins, steroid hormones, purines, pyrimidines, derivatives of nucleic acid, and urinary organic acids. The advantages of TLC include simphcity, rapidity, versatility, ability to process a large number of samples... 

The calibration and calculation operations in a quantitative TLC analysis are handled automatically by the software in modern computer-controlled densitometers. Chromatographic and measurement errors in scanning densitometry were discussed by Allwohn and Ebel (1989), Ebel and Glaser (1979), and Poliak (1989). 

It looks as if this procedure would be complicated, but it is not when a computer-controlled densitometer is used. The analyst applies samples and standards with an applicator, scans the undeveloped plate, and then develops it and rescans it. All calculations are performed automatically. While running, the program suggests the steps necessary to improve the quality of the data and their evaluation. When the times comes when digital cameras will be used, these procedures will become even simpler. After the application, the basic image of a spotted plate will be taken, and after the development, the plate will be scanned again. The data from both pictures will be compared and processed, and, because of the large volume of collected information, it is expected that most of errors will be detected and reduced. 

Today, modern computer-controlled densitometers allow quantitative determination of almost all chemicals, both colorless and colored, which have UV-Vis absorbance or are capable of measuring fluorescence. Densitometric detection can be performed in a wide wavelength range, from 200 to 800 nm, with high sensitivity and accuracy. The method is rapid and offers a possibility of analyzing simultaneously a large number of samples along with consecutive qualitative analysis (absorption spectra, R values). 

Guidelines for uniform density and dot gain limits have been estabHshed. These criteria ate the standards used to control the press. A densitometer is used to measure these standards and measurements ate made on sheets removed as the press is mnning. The operator then uses these measurements to guide press adjustments, primarily ink flows, to correct the printed result to standard (8). 

Figure 7.20 Computer-controlled automated scanning densitometer for quantitative TLC.

Figure 7. Flow and instrumentation diagram for high temperature, balanced pressure bench-scale unit. LR, level recorder LT, level transmitter TR, temperature recorder FR, pressure recorder FR, flow recording RD, densitometer PC, pressure controller TC, temperature controller GM, gas meter F, filter VSD, variable speed drive PCV, pressure regulator CV, control valve SV, solenoid valve S, gamma ray source.

Film and Photomultiplier Detector Systems. In the original ultracentrifuge built by Svedberg and co-workers (3,4) and also in the commercial instrument, the light intensity pattern was recorded on film, requiring a densitometer to obtain the absorption profile. Schachman and co-workers ( 5,6) constructed a mechanically scanned photomultiplier system in the early 1960 s. In addition to a commercial version now available (7), a number of other systems with computer controlled gathering of data have been constructed (8, , 10, 11, 12). 

In the above method, an autotitrator repeatable to +0.5% relative error (or 0.01 wt%) was employed to measure H2O2 wt%, independent of the relative amounts of DI water and slurry. A densitometer with an accuracy of + 0.0005 g/cc was used to estimate the percentage volume of slurry in the mixture (repeatability = +3% slurry by volume), for a specific concentration of H2O2. In the above blend, the measured H2O2 wt% concentration was used to control the speed (or strokes/batch) of the W2000 slurry pump, whereas the density, obtained using a coriolis mass flow meter, controlled the speed of the DI water pump. On the basis of the property curves obtained from bench... 

Computer-controlled systems were originally developed by Ebel and his team in order to achieve the most optimum positioning of any spot on a sample track in the measuring slit. Furthermore scanning during a standstill of the plate was considered an advantage, since errors which result from the time constants of densitometers and integrators could be avoided 28,29). 

Recently Kaiser and collaborators have developed new possibilities by means of a computer-controlled system 4i,44). They have coupled a desk computer (Apple II) via an AD-converter to the signal output of a Camag-scanner. Additionally, the computer is connected to the control electronics of the scanning table of the densitometer. Thus the transmission of the raw measuring data and the computer-control of the automatic scan is possible. 

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