Paper chromatography determinations

The amino group is readily dia2oti2ed in aqueous solution, and this reaction forms a basis for the assay of sulfas. Aldehydes also react to form anils, and the yellow product formed with 4-(dimethylamino)hen2a1dehyde can be used for detection in thiu-layer and paper chromatography. Chromatographic retention values have been deterrnined in a number of thiu layer systems, and have been used as an expression of the lipophilic character of sulfonamides (23). These values have corresponded well with Hansch lipophilic parameters determined in an isobutyl alcohol—water system. 

Specifications, Analysis, and Toxicity. Dicyandiamide is identified quaHtatively by paper chromatography and quantitatively by ultraviolet spectrometry of the chromatogram. More commonly, total nitrogen analysis is used as a purity control or the dicyandiamide is converted by hydrolysis to guanylurea, which is determined gravimetrically as the nickel salt (50). Methods based on the precipitation of silver dicyandiamide picrate are sometimes used (51). Dicyandiamide can also be titrated with tetrabutylammonium hydroxide ia pyridine solution. Table 4 gives a typical analysis of a commercial sample. Dicyandiamide is essentially nontoxic. It may, however, cause dermatitis. 

The sulfate is obtained by evaporating the aqueous layer in vacuo. The hydrochloride can be obtained in the same way but using HCl instead of H2SO4. SAM-HCl has a solubility of 10% in H2O. The salts are stable in the cold at pH 4-6 but decompose in alkaline media. [Cantoni Biochem Prep 5 58 1957.] The purity of SAM can be determined by paper chromatography [Cantoni J Biol Chem 204 403 1953 Methods Enzymol 3 601 1957], and electrophoretic methods or enzymic analysis [Cantoni and Vignos J Biol Chem 209 647 1954]. 

Palladium, D. of as dimethylglyoximate. (g) 463 as nioximate, (g) 474 by EDTA, (ti) 329 Paper chromatography 229 see Thin layer chromatography Parallax errors due to, 85 Parallel determinations 132 Partial ionic equations 850, (T) 851 Partition chromatography 13. 217 Partition coefficient 162 Patton and Reeder s indicator 317, 328 Peptisation 419. 421... 

Many methods of determining individual colorants and blended compositions have been selected, critically reviewed, and published. Combined methods such as paper chromatography, TLC, HPLC, spectrophotometry, voltammetry, and more recently, CZE have been developed. Table 6.6.1 presents some of these methods. 

Spectrophotometric resolution for the discrimination of individual colorant molecules found in mixtures is lower than that of chromatographic techniques such as TLC or HPTLC and even low-cost paper chromatography. More expensive but more accurate determinations may be made by RP-HPLC, IP-HPLC with UV-Vis, and diode array detection. ... 

Perry, T. L. and Schroeder, W. A., The occurrence of amines in human urine determination by combined ion exchange and paper chromatography,... 

Jalal and Collin19 used paper chromatography and TLC to determine caffeine in both coffee and tea, and theobromine in tea. Their TLC method used cellulose plates that were developed with butanokhydrochloric acid water (I 00 1 1 28) for 4 h. The spots were eluted from the plates with ammonium hydroxide and measured spectrophotometrically against a blank at 272 nm for caffeine and 274 nm for theobromine. 

Sjoeberg, A.M. and Rajama, J., Simple method for the determination of alkaloids in cocoa using paper chromatography and UV spectrometry, J. Chromatography, 295,291,1984. 

Paper chromatography was applied formerly to the determination of bromocriptine. 

Paper chromatography using Whatman No. 1 paper was once used to determine the homogeneity of halcinonide. 60 Twenty percent formamide in methanol comprises one stationary phase and methylisobutyl ketone-formamide (20 1) is the mobile phase. A second solvent system uses 25% propylene glycol-chloroform as the stationary phase and toluene saturated with propylene glycol as the mobile phase. 

To determine the relative amounts of neomycins B, C and neamine by a radio chemical method, Kaiser 2 separated the -labelled N-acetyl derivatives by paper chromatography and quantitated the chromatograms by liquid scintillation counting. A coefficient of variation of 3.6% was obtained. 

A better method has been described by Schwarz (S6). Washed red blood cells are lysed, precipitated with trichloroacetic acid below 0°C and the supernatant quickly neutralized. Speed and low temperatures are necessary to prevent hydrolysis of galactose-l-phosphate which is very sensitive to acid. Barium acetate and ethanol are added, and the precipitated barium salt of galactose-l-phosphate washed with 80% ethanol. The barium salt is then hydrolyzed by heating with dilute hydrochloric acid, acid and salts removed with mixed ion-exchange resins, and the galactose estimated by paper chromatography as described above. It is probably better to avoid the Amberlite MB-1 resin used by Schwarz and, instead, to use a weak base resin mixture, such as Amberlite MB-4. Recovery of added galactose-l-phosphate should be determined simultaneously. 

The column chromatography technique using Dowex 50 ion-exchange resin, introduced in 1951 (M2) and improved in 1954 (M3) by Moore and Stein, first made possible the precise quantitative analysis of amino acids liberated in the course of acid hydrolysis of urine. Similar results were also obtained by Muting in 1954 (M4), who used paper chromatography methods. In this procedure amino acids were quantitatively determined after staining on the paper and elution of the resulting spots. 

Special attention should be paid to polypeptides isolated from a non-diffusible fraction of normal human urine by Boulanger et al. (BIO). By means of paper chromatography they isolated from this mixture seven single polypeptides and determined their amino acid composition. The results of the analyses are given in Table 3. 

High-voltage electrophoresis and subsequent paper chromatography of the fractions obtained made possible the isolation from the analyzed mixture of twenty-two components giving colored spots with ninhydrin and isatin. Among these, fourteen were identified as peptides and their amino acid composition established (Table 5). In the case of eight peptides, also N- and C-terminal amino acids were determined (Table 6). 

Paper Chromatography.—A ketonic solvent system has been recommended for the rapid determination of nucleotide-inorganic phosphate mixtures,221 and a procedure for the simultaneous determination of metabolites of various chemical classes has been developed.222... 

The anthocyanin profile of the flowers of Vanda (Orchidaceae) was investigated with a similar technique. Flowers (2 kg) were extracted with 101 of methanol-acetic acid-water (9 l 10,v/v) at ambient temperature for 24 h. The extract was purified by column chromatography, paper chromatography, TLC and preparative RP-HPLC. Analytical HPLC was carried out in an ODS column (250 X 4.6 mm, i.d.) at 40°C. Gradient conditions were from 40 per cent to 85 per cent B in 30 min (solvent A 1.5 per cent H3P04 in water solvent B 1.5 per cent H3P04, 20 per cent acetic acid and 25 per cent ACN in water). The flow rate was 1 ml/min and analytes were detected at 530 nm. The chemical structures of acylated anthocyanins present in the flowers are compiled in Table 2.90. The relative concentrations of anthocyanins in the flower extracts are listed in Table 2.91. It can be concluded from the results that the complex separation and identification methods (TLC, HPLC, UV-vis and II NMR spectroscopy, FAB-MS) allow the separation, quantitative determination and identification of anthocyanins in orchid flowers [262],... 

P-Mannanases are generally prepared by conventional chromatographic procedures, which can lead to high yields of enzyme, but in some cases only a poor state of purity. Small quantities of highly purified p-mannanases have been prepared by substrate affinity chromatography of partially purified enzymes on a column of glucomannan immobilised on aminohexane Sepharose 4B (6). The action patterns of enzymes described in this paper were determined using enzymes purified by this latter procedure. 

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