Colorimetric procedures, measurement

Bond and Willis 2") have determined ammonia in mitrochondria samples after separation by distillation. The enhancement of zirconium absorption by the ammonia is measured. This is more rapid than colorimetric procedures. 

The precision of absorption measurements depends upon the degree of sophistication of the instrumentation and on the chemical species involved, both of which can affect the apparent validity of the Beer-Lambert law. Where a single absorbing species exhibiting a broad flat maximum is to be determined, adequate results can often be achieved with a simple filter-photometer. In the visible region, this technique is known as colorimetry. The inherent disadvantage of colorimetric procedures using simple filter instruments with a broad bandpass lies in the invalidation of the Beer-Lambert law and the lack of compatibility between results from different... 

GLC, atomic absorption and mass spectrophotometry, enzymatic, and specific colorimetric procedures seem to be the likely candidates for routine use in the future. Automation will certainly be common. GLC is now used to detect imitation muscat wines (127). Characteristic flavor byproducts of yeasts may be detected and measured. Multiple correlation of the amounts of the more influential major and minor constituents with wine quality is the goal of such research. A simple apparatus for the simultaneous determination of the redox potential (two platinum electrodes), pH, specific conductivity, oxygen, and carbon dioxide (ion-specific electrode) has been devised (128). Molecular oxygen in wines has been determined by several procedures—polarography (129) and GLC being the latest. 

Because of their speed and reasonable instrumental requirements, there is continuing interest in colorimetric procedures for lactose determinations. Most procedures are based on the reducing properties of lactose samples that contain only one sugar are easily measured colori-metrically, but samples with three or four sugars may require several different colorimetric assays to determine the composition accurately. 

When identical lipase reaction subsamples are measured for fatty acid levels by both titrimetry (Basic Protocol 1) and colorimetry (Basic Protocol 2), estimates by the colorimetric procedure are only 60% of those obtained by titrimetry. However, the nature of this rela-... 

The other major types of assay are all spectrophotometric (Alternate Protocols 1 and 2) and usually involve measuring the production of the highly colored end-product of the DPO reaction. They are used widely because they are rapid, can be carried out with the simplest of colorimeters/spectrophotometers, and can be adapted to use microtiter plates (Alternate Protocol 3). Thus they are ideal for simple routine comparative or monitoring studies. A major weakness of these colorimetric procedures is that they measure the end-product of a complex sequence of reactions, and different substrates yield different... 

This method is based on the fact that only esters, but not free carboxylic acids, react with hydroxylamine to form hydroxamic acids, which can be determined colorimetrically as complex with ferric chloride (8). The method—in contrast to most other procedures—measures the concentration of remaining substrate instead of products of hydrolysis. It also requires purified enzymes because of the interference of colored contaminants in the colorimetric measurements. 

For instance, suppose one measures the optical absorption of a solution in a colorimetric procedure. This procedure specifies that the measurement should be carried out at pH 2.5 and room temperature. On verification, it is found that a clearly different result is obtained at pH 2.4 and that differences are also observed when the temperature is 20°C and 25°C. The method will display higher variability between laboratories and between days than expected, because it is not rugged. 

If a water sample contains both soluble and insoluble manganese (Mn) compounds and ions, and it is filtered to separate the dissolved and insoluble fractions, and the filtrate and insoluble residue are analyzed separately, the results can be expressed as total dissolved Mn and total suspended or insoluble Mn. Phosphorus (P) can be determined colorimet-rically as the ortho-phosphate ion, P043, in aqueous samples after a reaction that forms an intensely blue-colored derivative. However polyphosphate ions and other ions and compounds containing P do not form this derivative. Total P in a sample can be determined with the same colorimetric procedure after acid hydrolysis and oxidation of all ions and compounds containing P to P04 3. In some elemental analyses the sample is treated with reagents designed to make available for measurement some fraction of an element or elements but not the total amount. For example, a soil sample may be treated with water at pH 3 to simulate the leaching process of acid rain. A total elemental analysis of the filtrate provides information about just those elements solubilized by the mild acid treatment. This can be called the determination of total mild acid leachable elements. 

The activity of alkaline phosphatase in the plasma was measured by a colorimetric procedure. 

Ninhydrin Assay for Adsorbed Proteins. Measurements were made by a colorimetric procedure based on the reaction of ninhydrin with amino acids (25). The films were hydrolyzed in 5 ml of 2.5N NaOH for 2 hrs in capped plastic tubes in a boiling water bath. Then 1.5 ml of glacial acetic acid was added and mixed next I ml of ninhydrin reagent was added and mixed. [The reagent was three times more concentrated in ninhydrin, SnCb, and citrate than prescribed by Moore and Stein (25)]. The tubes were capped and boiled 20 mins more. The solution was clarified by centrifugation, and the absorbance read immediately at 570 nm on a Beckman DB spectrophotometer. If necessary, the sample was diluted with 50-50 2-propanol-water. Calibration curves (absorbance vs. fig of protein) were constructed in the 0-30 and 0-100 fig range with known amounts of each type of protein subjected to this same analysis procedure. 

A title The Analysis of Potassium, ought to mean a procedure for determining the minor constituents in potassium metal, and not a method for the determination of potassium. A title, A Micro Method for Copper, may refer to the use of a very small sample containing copper, or to some method for measuring trace amounts of copper. Used in the latter sense, the speaker has learned to assume that any such title may refer to a colorimetric procedure. It is annoying, then, to find that the summary does not clarify the uncertainty, and consequently to have to read the article, or much of it, in order to find out what was done. A Trace Method for Copper is just as unsatisfactory for a title, as any one of a half-dozen methods applicable to trace amounts might be described. 

Several investigators have reported that ascorbic acid can be analyzed by gas-liquid chromatography following conversion of the parent compound to its trimethylsilyl ether (61-66). The procedures have been found to be reliable and to produce results comparable with those obtained by colorimetric procedures. In most cases, however, measurement of only the reduced form of the vitamin is possible (67). One method is suitable for microanalytical work and has the advantage that several other carbohydrates and carbohydrate derivatives can be measured simultaneously in the same extract (67). 

A new colorimetric procedure has been developed for the quantitative measurement of atmospheric ozone. The specific, sensitive, and reproducible nonaqueous method utilizes the rate of color produced on ozonization of o-dichlorobenzene solutions of N-phenyl-2-naphthylamine. The color reaction conforms to Beer s law and has a practical sensitivity of 1.8 X 10 mg. of ozone. Oxygen and oxides of nitrogen, in concentrations greater than those normally occurring in the atmosphere, do not interfere with the chromogenic reaction. 

In the colorimetric determination of pH we always make use of a comparison solution of known acidity. The accuracy of such a comparison method evidently is limited by the reliability of the comparison solution. As has been mentioned in the preceding chapter, the pH of the comparison or buffer solution is measured by means of the hydrogen electrode. Hence the colorimetric procedure is based ultimately upon the potentiometric method. It is the hydrogen ion activity rather than the con-centration which is determined by the latter method. Thus when it is stated that an unknown and a buffer solution have the same pH on the basis of equal color intensities, it is really the hydrogen ion activity which is implied. 

Quite generally, when the colorimetric procedure is being applied to solutions containing substances of which the influence on the color of the indicator is unknown (for example, colloids, organic compounds, etc.), the results must always be checked electrometrically. The hydrogen ion measurement with the hydrogen electrode must always be considered as exact. 

Cardiac muscle hydroxyproline, measured by a simple colorimetric procedure with inadequate specificity in this situation, increased slightly beyond age 35-40, while a similar rise was seen in the hydroxyproline of aortic tissue (C9). Other studies using identical methods showed no change with age in aortic and pulmonary artery hydroxyproline (Bll, K2). The influence of both age and cardiac hypertrophy on cardiac... 

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