Internal standards, spectrophotometric

A sixth spectrophotometric method for the quantitative determination of Pb + levels in blood uses CQ+ as an internal standard. A standard containing 1.75 ppb Pb + and 2.25 ppb CQ+ yields a ratio of Sa/Sis of 2.37. A sample of blood is spiked with the same concentration of Cu +, giving a signal ratio of 1.80. Determine the concentration of Pb + in the sample of blood. 

A seventh spectrophotometric method for the quantitative determination of Pb + levels in blood gives a linear internal standards calibration curve for which... 

The accuracy of the inductively-coupled plasma procedure was assessed by analysing waters of known sulfate composition, and by comparing measured sulfate values for a wide range of samples with those obtained for the same waters by an automated spectrophotometric procedure. Good agreement is obtained between the derived sulfate measurements and the normal values for International Standard Sea Water and an EPA Quality Control Standard. 

After two decades of quantitative l.c. analyses, it has been established that, when proper precautions are taken, these methods can provide accurate and reproducible results.Quantitative l.c. measurements are usually as accurate as, and often more precise than, those obtained by spectrophotometric, " paper-chromatographic, and gas-liquid-chro-matographic methods. Both external and internal standardization have been used to translate peak height or areas into quantitative, solute-concentration values. Because peak heights are easy to measure, many methods use this parameter, and, when slightly overlapping peaks or unsteady baselines are encountered, it is the method of choice. With introduction of... 

International Standard Organization. 1982. Water quality. Determination of total arsenic. Silver diethyl-dithiocarbamate spectrophotometric method. ISO 6595. International Organization for Standardization, Case Postale 56, CH-1211, Geneva 20 Switzerland. 

Copper in serum and natural waters was determined with bathocuproinedisulfonic acid using bromophenol blue as internal standard [8]. Extraction (CHCI3) with Aliquat 336 preceded the spectrophotometric measurement. The ratio of the absorbances at 471 nm (copper complex) and 602 nm (internal standard) was taken as the analytical parameter. Detection limit was 0.4 g f. ... 

The quantum yield of fluorescence from p-hydroxybenzoic acid is low (10). It was found that this fluorescence was subject to strong quenching in irradiated solutions. In the fluorescence assay, therefore, p-hydroxybenzoic acid was added as an internal standard. The G-values so obtained were in agreement with those obtained by spectrophotometric assay at an isobestic point. 

Stepanov AV, Nikitina SA, Karasev VT, Belyaev BN, Stepanov DA, Pevtsova EV, Bogoroditskij AB (2002) Precise spectrophotometric method for the determination of U, Pu, Nd, and Rh using the principle of internal standardization. Radiokhimiya 44(2) 165-169... 

Surugaya N, Taguchi S, Sato S, Watahiki M, Hiyama T (2008) Spectrophotometric determination of plutonium in highly radioactive liquid waste using an internal standardization technique with Neodymium(III). Anal Sci 24(3) 377... 

Since, In practice, the efficiency frequently varies, it is necesseuy to determine the counting efficiency before one can compare different samples. A radioactive standard of acciirately known activity is essential for the determination of the efficiency counting. The use of radioactive standards is in principle similar to that of standards in colorimetric or spectrophotometric assays. There are two types of standards (i) internal standard (this is usually a p-emitter of accurately known activity which, when dissolved in unquenched scintillation mixture, provides a reference standard), and (ii) external standard (ay-emitter incorporated in most instruments). 

Figure 6.7. Separation of the methyl ester derivatives of fatty acids from the phospholipids of mouse brain by reversed-phase HPLC with spectrophotometric detection at 192 nm [67]. In essence, the column of Zorbax ODS phase, maintained at 35 C, was eluted stepwise with acetonitrile-water (7 3, v/v) then with acetonitrile alone. Methyl elaldate was added as an internal standard. (Reproduced by kind permission of the authors and of the Journal of Lipid Research, and redrawn from the original paper).

Uslu, B. Ozkan, S.A. Determination of lamivudine and zidovudine in binary mixtures using first derivative spectrophotometric, first derivative of the ratio-spectra and high-performance liquid chromatography-UV methods, Anal.Chim.Acta, 2002, 466, 175-185. [LOQ 25 ng/mL for lamivudine finasteride is internal standard]... 

Tatar, S. Saglik, S. Comparison of UV- and second derivative-spectrophotometric and LG methods for the determination of valsartan in pharmaceutical formulation, J.Pharm.Biomed.Anal., 2002,30, 371-375. [capsules losartan is internal standard LOD 200 ng/mL LOQ 1 xg/mL]... 

Spectrophotometric method To overcome the limitations of visual methods, a more scientific approach to color quantification has been developed. The International Commission on Illumination (Commission Internationale de TEclairage, or CIE) introduced the element of standardization of sources and observer. The CIE also introduced methodology to derive numbers that provide a measure of a color seen under a standard source of illumination by a standard observer. The CIE system has become the international standard for color measurement. 

The expert Committee on Biol( cal Standardization of the World Health Organization is proceeding with the establishment of an international standard for vitamin Bu. Until this standard is available Crystalline B 2 Merck is recommended as a standard. The U. S. P. identification and spectrophotometric assay for crystalline B12 are as follows (48) ... 

Prieto et al. (1999) reported another spectrophotometric method for the quantitative determination of antioxidant capacity based on the reduction of Mo(VI) to Mo(V) by vitamin E in acidic conditions with incubation at 95°C for 90 min. The subsequent green phosphate/Mo(V) complex, after cooling at room temperature was monitored at 695 nm with a calibration range of 0.2-2 x 10 M (r = 0.997) and a detection limit of 0.135 pmol vitamin E. The method was applied for measurement of total antioxidant capacity of plant extracts and to determine vitamin E in a variety of grains and seeds, including corn and soybean. The recovery of vitamin E from seeds was determined by supplementing the samples with the different vitamin E isomers or a-tocopherol acetate as internal standard and applying both the proposed method and a standard HPEC assay (Huo et al., 1996) and yielded a recovery of 93%-97% tocopherols. 

Objective Evaluation of Color. In recent years a method has been devised and internationally adopted (International Commission on Illumination, I.C.I.) that makes possible objective specification of color in terms of equivalent stimuli. It provides a common language for description of the color of an object illuminated by a standard illuminant and viewed by a standard observer (H). Reflectance spectro-photometric curves, such as those described above, provide the necessary data. The results are expressed in one of two systems the tristimulus system in which the equivalent stimulus is a mixture of three standard primaries, or the heterogeneous-homogeneous system in which the equivalent stimulus is a mixture of light from a standard heterogeneous illuminant and a pure spectrum color (dominant wave-length-purity system). These systems provide a means of expressing the objective time-constant spectrophotometric results in numerical form, more suitable for tabulation and correlation studies. In the application to food work, the necessary experimental data have been obtained with spectrophotometers or certain photoelectric colorimeters. 

Quantitative data can be obtained by means of the use of an internal or external standard, whose exact concentration can be checked by means of spectrophotometric detector, which measures spe-cihc absorbance or molar absorbance. Spiked authentic samples can be used to measure the limit of detection (LOD) and the limit of quantihcation (LOQ). 

One milligram of microsomal protein is added to 0.1 M potassium phosphate buffer (pH 7.4) containing 50 mM NaF, 10 mM dithiothreitol, 1 mM EDTA, 20% glycerol (v/v), 150 iM 5-cholestene-3/3, 7a-diol, and 0.915% CHAPS. The reaction is initiated by 1 mM NAD+ to give a final reaction volume of 1.0 mL. After incubation at 37°C for 5 minutes, the reaction is terminated by adding 2 mL of 95% ethanol. An internal recovery standard, 4-cholesten-3-one (3 fig in methanol) is also added. The steroid products are extracted into 5 mL of petroleum ether (repeated twice). After the ether has been removed at 40°C under a stream of nitrogen, the products are dissolved in 100 fxL of mobile phase and 20 ju.L is injected into the column. The amount of product formed is linear with protein (to 1.5 mg) and with time (up to 10 min, 1 mg protein). The assay is much more sensitive than the direct spectrophotometric assay, and it avoids the use of thin-layer chromatography and radioisotopes described in other methods. 

Once a reflectance curve is obtained spectrophotometrically, the measured information is mathematically transformed according to standard conventions, into the numbers used to describe the color of the sample. In practice, a standard observer and standard illuminant are selected based on test methods defined by the Commission International de I Eclairage (CIE). Calculation procedures involve the numerical integration of the product of the spectral power distribution S(l) of the light source and the reflectance factor R(l). Reflectance factors represent the percentage of light reflected by the sample at each wavelength. 

OP), and carbamate pesticides are available as standard methods by the AO AC covering both nonfatty and fatty foods. These methods are based on solvent extraction and a variety of column chromatographic cleanup procedures with determination by GC using selective detectors. The International Dairy Federation has approved similar methods for OC and OP pesticide residues in milk and milk products. The AOAC have standard methods for specific pesticides or groups of pesticides that are approved for certain foodstuffs these methods involve colorimetric, spectrophotometric, or gas chromatographic determination. In the case of me-thylcarbamate pesticides, a liquid chromatographic method is approved. 

Five important treatises on color have appeared in recent years. Handbook of Colorimetry (Hardy, 1936) is a direct outcome of the eighth session of the International Commission on Illumination in 1931. It enables us to compute psychophysical values for brightness (later termed luminous refiectance and now, lightness) and chromaticity (dominant wave length and purity) from the raw spectrophotometric data. In these psychophysical terms, a colored sample may be uniquely defined with respect to its color, provided that we follow a standard procedure for determining the spectral refiectance or transmittance of the sample. The perceived color, however, is not defined, and we enter here into psychological considerations as distinct from those of psychophysics. The C.I.E. quantities x and y (the trichromatic coefficients which... 

Table 6.1 displays recommended specifications for wavelength accuracy, photometric linearity, and spectrophotometric noise levels for pharmaceutical applications. The first step in validating any NIR method is to test the suitability of these specifications for a given application. Wavelength accuracy tests conducted using appropriate external standards will prevent potential problems that could occur with proprietary internal calibration protocols. The exact nature of any calibration standard must be noted in a validation protocol. Rare earth oxides and glass standards are candidates for such calibration. 

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