V — Sulfuric Acid Reagent

Another example of applications concerning separation of piperonylic, 5-, 2-, and 6-methoxypiperonylic acids was performed on silica gel layers using ethyl acetate-hexane-acetic acid (50 50 0.5, v/v) as the mobile phase. Detection was performed by spraying with a chromotropic-sulfuric acid reagent. With these conditions the following hRf values of 64, 54, 47, and 37 were obtained, respectively. 

Fig. 25. Solubility values for styrene and methyl methacrylate in aqueous emulsions. A water in styrene, determined with either Karl-Fischer titration or cloud point (261) A strene in water, determined with formaldehyde-sulfuric acid reagent (261) V styrene in...

A solution of 1-2 % (v/v) concentrated sulfuric acid in methanol transesterifies lipids in the same manner and at much the same rate as methanolic hydrogen chloride. It is very easy to prepare, and it is thus the author s preferred reagent for esterification of free fatty acids, but utilised at a temperature below reflux. If the reagent is used carelessly, some decomposition of polyunsaturated fatty acids may occur. 

Note The reagent can be employed on sihca gel, kieselguhr. Si 50 000 and RP layers. Hydrochloric or sulfuric acid can be employed in place of phosphoric acid (q.v.). The detection limits for steroids and digitahs glycosides are several nanograms per chromatogram zone. 

Note The individual components of the reagent can also be applied separately one after the other [12, IS], e.g. the chromatogram is first immersed in an 8% methanolic 4-(dimethylamino)-benzaldehyde solution and then, after intermediate drying, sprayed v th 25% sulfuric acid [12]. 4-(Dimethylamino)-benzaldehyde can be replaced in the reagent with 4-(dimethylamino)-ciimamaldehyde [1]. 

Note Like sulfuric acid (q.v.) ort/io-phosphoric add is a universal reagent, with which almost all classes of substance can be detected at high temperatures (150-180 °C) by charring e.g. high molecular weight hydrocarbons (mineral oils) [20]. The colors and fluorescences produced at lower temperatures (<120°C) and their intensities are very dependent on the temperature and period of heating. It is not possible to use meta- or pyrophosphoric acid in place of or/Ao-phosphoric add, since, for instance, amanita toxins react well with alcoholic phosphoric add only weakly with aqueous phosphoric add and not at all with meta- or pyrophosphoric acid [17]. 

Schnepfe [83] has described yet another procedure for the determination of iodate and total iodine in seawater. To determine total iodine 1 ml of 1% aqueous sulfamic acid is added to 10 ml seawater which, if necessary, is filtered and then adjusted to a pH of less than 2.0. After 15 min, 1 ml sodium hydroxide (0.1 M) and 0.5 ml potassium permanganate (0.1M) are added and the mixture heated on a steam bath for one hour. The cooled solution is filtered and the residue washed. The filtrate and washings are diluted to 16 ml and 1ml of a phosphate solution (0.25 M) added (containing 0.3 xg iodine as iodate per ml) at 0 °C. Then 0.7 ml ferrous chloride (0.1 M) in 0.2% v/v sulfuric acid, 5 ml aqueous sulfuric acid (10%) - phosphoric acid (1 1) are added at 0 °C followed by 2 ml starch-cadmium iodide reagent. The solution is diluted to 25 ml and after 10-15 min the extinction of the starch-iodine complex is measured in a -5 cm cell. To determine iodate the same procedure is followed as is described previously except that the oxidation stage with sodium hydroxide - potassium permanganate is omitted and only 0.2 ml ferrous chloride solution is added. A potassium iodate standard was used in both methods. 

In the malachite green procedure, 10 ml of the sample solution containing up to 0.7 xg phosphorus as orthophosphate was transferred into a 25 ml test tube. To this solution was added 1 ml each of 4.5 M sulfuric acid and the reagent solution. The solution was shaken with 5 ml of a 1 3 v/v mixture of toluene and 4-methylpentan-2-one for 5 min. After phase separation, the absorbance of the organic phase was measured at 630 nm against a reagent blank in 1 cm cells. 

Arsenic (V) causes large positive errors - arsenic (V) at a concentration of 10 pg/1 produces an absorbance of 0.07, but can be masked with tartaric acid (added in the reagent solution). When arsenic (V) was present at concentrations of 50 pg/1 it was masked with 0.1 ml of 1 x 10 4 M sodium thiosulfate added after the sulfuric acid. 

The consumption of the oxime can be checked by thin-layer chromatography on silica gel G with the solvent system chloro-form/methanol (95/5 v/v) and a spray reagent consisting of 5% potassium dichromate in 40% sulfuric acid. The oxime appears as an immediate dark spot and the aziridine as a yellow spot. The checkers observed identical mobilities (Rf 0.8) for both compounds. 

Smooth polycrystalline Au, Pt and Ir thin-layer electrodes were utilized (10-11). Electrodes were cleaned between trials by sequential electrochemical oxidation above 1.2 V [Ag/AgCl (1 M Cl-) reference] and reduction below -0.2 V in 1 M H2SO4 surface cleanliness was verified with the aid of cyclic voltammetry in the same molar sulfuric acid solution. Experiments were carried out in 1 M H2SO4, 1 M NaC104 buffered at pH 7 and 10, and in 1 M NaOH solutions were prepared with pyrolytically triply distilled water (12). Surface reagents employed were iodide, hydroquinone (HQ), 2,5-dihydroxythiophenol [DHT (13)1. and 3,6-dihydroxypyridazine (DHPz). 

The first studies on the sulfation of organic compounds, amino acids, and proteins have shown that pyridine/sulfur trioxide complex (pyridine/S03 or pyridine/Cl S03H),168-721 concentrated sulfuric acid,173,74 sulfuric acid//V,A -dicyclohexylcarbodiimide,175,761 and chloro-sulfonic acid177 are the most efficient reagents for the sulfation of tyrosine. More recently, alternative methods based on dimethylformamide/sulfur trioxide complex (DMF/S03),152,781 trimethylamine/sulfur trioxide (Me3N/S03),1152,1531 pyridinium acetylsulfate,137,791 and pyr-idinium trifluoroacetylsulfate1801 have been proposed to minimize side reactions which are difficult to control for the chemical sulfation of tyrosine peptides. 

The syntheses of iron isonitrile complexes and the reactions of these complexes are reviewed. Nucleophilic reagents polymerize iron isonitrile complexes, displace the isonitrile ligand from the complex, or are alkylated by the complexes. Nitration, sulfonation, alkylation, and bromina-tion of the aromatic rings in a benzyl isonitrile complex are very rapid and the substituent is introduced mainly in the para position. The cyano group in cyanopentakis(benzyl isonitrile)-iron(ll) bromide exhibits a weak "trans" effect-With formaldehyde in sulfuric acid, benzyl isonitrile complexes yield polymeric compositions. One such composition contains an ethane linkage, suggesting dimerization of the transitory benzyl radicals. Measurements of the conductivities of benzyl isonitrile iron complexes indicate a wide range of A f (1.26 e.v.) and o-o (1023 ohm-1 cm.—1) but no definite relationship between the reactivities of these complexes and their conductivities. 

In a further method, sample digests are prepared according to method 1(c) of the Analytical Methods Committee [ 100] using precautions described subsequently [101]. The resulting 100 ml of digest, which is in normally 5% v/v sulfuric acid, should not be colourless and should contain any suspended solids. At the same time, prepare two reagent blanks from the volume of acid used in sample oxidation. 

It is best to purify the methyl esters by thin-layer chromatography of the sample on another silica gel G (250 p,m) plate using this solvent system, but without any spray reagent being used. A comparable plate is run and the methyl ester band is detected by the sulfuric acid/char reaction. Then the unsprayed plate is scraped at the methyl ester area and the silica gel is extracted with petroleum-diethyl ether (80 20, v/v) or with chloroform-methanol-water (1 2 0.8, v/v). It is always prudent to spray the latter plate (after removing the desired area by scraping) with sulfuric acid and then charring. This will validate whether the apparent removal of all of the methyl esters has been accomplished. 

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