Other Analytical Methods

Two sensitive chemiluminescence methods for determining D-glucose have appeared. One uses immobilized glucose oxidase to produce hydrogen peroxide, then immobilized peroxidase to catalyse the 

Quantitative determinations of L-ascorbic acid, D-galactose, D-glucose, cellobiose, lactose and sucrose were made with enzyme-thermistors , in which the heat produced by reaction of a sugar with an immobilized enzyme (e.g , Invertase for sucrose) in a small column is monitored by a thermistor placed in the column eluent the procedure is suitable for discrete samples or continuous monitoring.  

2-Deoxy-glucose could be determined in the presence of 2-deoxy-galactose by treatment with galactose oxidase followed by indole-hydrochloric acid, and vice versa using glucose oxidase alternatively the hydrogen peroxide released enzymically could be deter- 

An iodometric analysis of aldoses amino-sugars in solution following reaction with fluorescamine have been described. 

The amperometric detection of simple sugars has been accomplished 

Many other analytical methods have been used or are potentially usable (for a complete listing, see Roedder, 1972 and 

Part of the H2O-CO2 system (Takenouchi and Kennedy, I965) Cq Critical curve Ci and C2 Intersection of the surface of immisci-hility hy isocompositional planes (as in 2 and U). 

Projection of the critical curve and of the intersections of the immiscihility surface hy planes of various Xc02 on the P-T plane. 

Section through the 300 C isotherm (l), illustrating the retrograde boiling phenomenon. 3 = maximum composition, Pm = maximum pressure for immiscihility to occur. C section of the critical curve. 

B H2O + CO2 homogenization (for a given XCO2) trend of deviation for the real isochores (from Ypma, 1963). All XCO2 mole %, 

The direct detection of a variety of simple carbohydrate compounds by u.v. methods is mentioned in reference 7 above. Spectrophotometry has also been used to determine ketoses (as furan derivatives and glucose by its reduction of 3,5-dinitrosalicylic acid to the 3-amino-5-nitro analogue.  

A potentiometric method for the determination of D-glucose, D-mannose, and D-fructose in a mixture of hexoses and pentoses employs fermentation by Streptococcus mutans. The response is based on the selective fermentation of these three carbohydrates by the organism, followed by measurement of hydronium ion. The response time is 4 min and other hexoses and pentoses do not interfere.  

A modification of the uronic acid — imidazole reaction (Cariotti reaction) has been described that eliminates interference by DNA.  

The instruments discussed earlier are the primary ones used in toxicant analysis, but an enormous number of analytical techniques are used in the field. Many of the instruments are expensive (e.g., Raman spectrometers, X-ray emission spectrometers) and few laboratories possess them. Many other instruments are available, however, such as the specific-ion electrode, which is both sensitive and portable. Specific-ion electrodes have many other advantages in that sample color, suspended matter, turbidity, and viscosity do not interfere with analysis therefore many of the sample preparation steps are not required. Some of the species that can be detected at ppb levels are ammonia, 

Principles and Methods of Toxicology. London Taylor and Francis, 2000. Morel, F. M. M., and J. G. Heirig. Principles and Applications of Aquatic Chemistry. New York Wiley-Interscience, 1993. 

Nelson, A. H. Organic Chemicals in the Aquatic Environment. Boca Raton, FL CRC Press, 1994. 

Pickering, W. F. Pollution Evaluation The Quantitative Aspects. New York Dekker 1977. Rand, G. M., and S. N. Petrocelli. Fundamentals of Aquatic Toxicology. Washington, DC Hemisphere, 1985. 

Rouessae, F., A. Rouessae, and M. B. Waldron. Chemical Analysis Modern Instrumentation, 

Ilnuma, Y. Hiraga, T. Kinoshita, and M. Watanabe, Chem. Pharm. Bull., 1979, 27, 1268. 

Ikenaka, and Y. Matsushima, Biochem. Biophys. Res. Commun., 1978, 85, 257. 

An electrochemical method involving electrocatalytic oxidation with TEMPO was suitable for the determination of glucose in aqueous solution. The cyanine dye 2, which carries two phenylboronic acid moieties displays increased fluorescence on binding to monosaccharides and provides a novel detection method. Compound 3 also contains a phenylboronic acid residue and binds D-glucose 6- 

Fujishige, S. Uchino and N. Oguri, Kobunshi Ronbunshu, 1996,53,201 Chem. Abstr., 1996,124, 343 864). 

Toyoda, T. Toida, I. Koshiishi and T. Imanari, Chem. Pharm. Bull, 1996,44,1017. 

The mechanism of the Molisch reaction, the very sensitive colour reaction between aldoses and 1-naphthol in concentrated sulphuric acid, has been elucidated following the isolation of the crystalline 

3-aminopropyltrlethoxysilane, gave a peak current linearly related 

A tltrlmetrlc determination of 2-deoxy-D-ribose has been described. Treatment with excess acidic aqueous periodate gave 1 equlv. of CH2(CH0)2 and 2 equiv. of iodate. Excess periodate 

Total ascorbic and dehydroascorbic acid in human serum has been determined by Iodine oxidation followed by reaction of dehydroascorbic acid with l,2-diamino-4,5-dimethoxy-benzene the Intensely fluorescent product permitted detection down to 5 ng of ascorbic acid with samples as small as 4 yl of serum. 

Bauer and W.Voelter, High Perform. Lig. Chromatogr. Biochem., 1985, 393 (Chem. Abstr., 1985, 1, 210 070). 

Removal of boric acid from solutions of carbohydrates with a boron- 

Yamada, T.Ohki, amd A.Saito, J. Chromatogr., 1986, 377, 15. 

Shalaby, euid P.Rocek, LP Spec. Chromatogr., Spektrosk., 1986, 64-8, 73 (Chem. Abstr., 1986, IPS, 126 421). 

A modification, utilizing mutarotase, of an enzymic colorimetric system for determining D-glucose with D-glucose oxidase, peroxidase, and 2,2 -diazobis-(3- 

Oxidation of nanomole amounts of O-substituted 2-acetamido-2-deoxy-D-galactitol and -glucitol, reduction of the resulting aldehydes with sodium borotri-tiide, and hydrolysis with acid afforded 2-amino-2-deoxy[ct)- H]alditols, which can be separated by t.l.c. and detected by autoradiography. Identification of the 2-amino-2-deoxy[ft - H]alditols determined the positions of substitution of the 2-acetamido-2-deoxyhexitols. A simple, rapid, and sensitive method for the simultaneous assay of free and ketosidically bonded sialic acids is noted in Chapter 8. 

Geller, Arch. Hydrobiol, SuppL, 1975,47, 295 Chem. Abs., 1977,86, 145 477w). 

Bagdasarov, Z P. Nebovidova, and V. D. Ponomarev, Fiz.-khim. Metody Anal. Kontrolya Proizvod., 1975, % (Chem. Abs., 1977,87.126 664j). 

Nanba and Y. Matsuo, Hiroshima Daigaku Kogakubu Kenkyu Hokoku, 1977, 25,47 Chem. Abs., 1977,87,184 808v). 

A specific, colorimetric determination of 2-acetamido-2-deoxy-D-galactose is based on its reaction with pentane-2,4-dione under anhydrous basic conditions to give a product identified as (458). Mild alkaline hydrolysis of (458) yields 2-methylpyrrole, which can be determined by the chromophore (Amax 548 nm) 

Periodate ion has been determined by the formation of a chromophore with 2,2 -azinobis(3-ethylbenzothiazoline-6-sulphonic acid) and by means of an iodide-selective electrode. The 2,4-dinitrophenylhydrazones derived from aldehydic fragments produced on periodate oxidation of carbohydrates have been separated by t.l.c., after which the individual hydrazones could be determined spectrophotometrically.  

New reagents have been recommended for the detection on chromatograms of non-reducing disaccharides (sodium periodate-silver nitrate in aqueous 

Kashimura, K. Yoshida, and K. Onodera, Agric. and Biol. Chem. (Japan), 1974, 38, 1725. 

Mahuzier, B. S. Kirkacharian, and C. Harfouche-Obeika, Analyt. Chim. Acta, 1975, 76, 

Aldoses reacted with 4-amino-3-hydrazino-5-mercapto-l,2,4-triazole in alkaline solution to give, after oxidation with hydrogen peroxide, stable 3-monosubsti-tuted mercaptotetrazine derivatives that are violet, and which absorb at 

The reactivity of 5-acetamido-3,5-dideoxy-L-flra6/7io-2-heptulopyranosonic acid (a seven-carbon analogue of iV-acetylneuraminic acid) in the resorcinol and thiobarbituric acid procedures - which are commonly used for the determination of sialic acids - has been evaluated. The chemistry involved in the formation of the characteristic chromophores in these assays was also discussed. 

A fluorometric determination of adenosine, AMP, ATP, and adenosine 3, 5 -phosphate, following their separation by high-pressure liquid chromatography, relies on their conversion into fluorescent l,lV -dietheno derivatives (519) on heating with monochloroacetaldehyde.  

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Although many quantitative applications of acid-base titrimetry have been replaced by other analytical methods, there are several important applications that continue to be listed as standard methods. In this section we review the general application of acid-base titrimetry to the analysis of inorganic and organic compounds, with an emphasis on selected applications in environmental and clinical analysis. First, however, we discuss the selection and standardization of acidic and basic titrants. 

Clinical Applications Perhaps the area in which ion-selective electrodes receive the widest use is in clinical analysis, where their selectivity for the analyte in a complex matrix provides a significant advantage over many other analytical methods. The most common analytes are electrolytes, such as Na+, K+, Ca +, H+, and Ch, and dissolved gases, such as CO2. For extracellular fluids, such as blood and urine, the analysis can be made in vitro with conventional electrodes, provided that sufficient sample is available. Some clinical analyzers place a series of ion-selective electrodes in a flow... 

Environmental Applications Although ion-selective electrodes find use in environmental analysis, their application is not as widespread as in clinical analysis. Standard methods have been developed for the analysis of CN , F , NH3, and in water and wastewater. Except for F , however, other analytical methods are considered superior. By incorporating the ion-selective electrode into a flow cell, the continuous monitoring of wastewater streams and other flow systems is possible. Such applications are limited, however, by the electrode s response to the analyte s activity, rather than its concentration. Considerable interest has been shown in the development of biosensors for the field screening and monitoring of environmental samples for a number of priority pollutants. 

Chemical kinetic methods are particularly useful for reactions that are too slow for a convenient analysis by other analytical methods. In addition, chemical kinetic methods are often easily adapted to an automated analysis. For reactions with fast kinetics, automation allows hundreds (or more) of samples to be analyzed per hour. Another important application of chemical kinetic... 

J)Other analytical methods which include, among many, a thermometric method (104), a high frequency titration (105), and a colored indicator method (106). 

A definitive method for stmctural deterrnination is x-ray crystallography. Extensive x-ray crystal stmcture deterrninations have been done on a wide variety of steroids and these have been collected and Hsted (270). In addition, other analytical methods for steroid quantification or stmcture determination include, mass spectrometry (271), polarography, fluorimetry, radioimmunoassay (264), and various chromatographic techniques (272). 

Infrared Spectroscopy (ir). Infrared curves are used to identify the chemical functionality of waxes. Petroleum waxes with only hydrocarbon functionality show slight differences based on crystallinity, while vegetable and insect waxes contain hydrocarbons, carboxyflc acids, alcohols, and esters. The ir curves are typically used in combination with other analytical methods such as dsc or gc/gpc to characterize waxes. 

The use of agarose as an electrophoretic method is widespread (32—35). An example of its use is in the evaluation and typing of DNA both in forensics (see Forensic chemistry) and to study heritable diseases (36). Agarose electrophoresis is combined with other analytical tools such as Southern blotting, polymerase chain reaction, and fluorescence. The advantages of agarose electrophoresis are that it requires no additives or cross-linkers for polymerization, it is not hazardous, low concentration gels are relatively sturdy, it is inexpensive, and it can be combined with many other analytical methods. 

The other analytical methods necessary to control the typical specification given in Table 5 are, for the most part, common quality-control procedures. When a chemical analysis for purity is desired, acetylation or phthalation procedures are commonly employed. In these cases, the alcohol reacts with a measured volume of either acetic or phthalic anhydride in pyridine solution. The loss in titratable acidity in the anhydride solution is a direct measure of the hydroxyl groups reacting in the sample. These procedures are generally free from interference by other functional groups, but both are affected adversely by the presence of excessive water, as this depletes the anhydride reagent strength to a level below that necessary to ensure complete reaction with the alcohol. Both procedures can be adapted to a semimicro- or even microscale deterrnination. 

A major advantage of static SIMS over many other analytical methods is that usually no sample preparation is required. A solid sample is loaded directly into the instrument with the condition that it be compatible with an ultrahigh vacuum (10" —10 torr) environment. Other than this, the only constraint is one of sample size, which naturally varies from system to system. Most SIMS instruments can handle samples up to 1-2 inches in diameter. 

For those suppliers not conforming to Ford, General Motors, and Daimler Ghrylser requirements, other analytical methods and acceptance criteria may be used if approved by the customer. If you lack any documented methods, the MSA Reference Manual is recommended. 

In as far as other analytical methods are concerned, many specific reactions have been elaborated for the quantitative determination of 2-deoxy aldoses. 2-Deoxy-D-ribose (2-deoxy-D-erythro-pentose), a compound which was recognized early as playing an important role in biological systems, has been of particular interest. Overend and Stacey (43) have given a critical review of the methods available until 1952 for the estimation of 2-deoxy pentoses. A recent summary of specific methods for the identification and quantitative estimation of the different classes of deoxy sugars has been prepared by Dische (13). 

X-ray emission spectrography, in common with other analytical methods, is subject to errors of different kinds. Lacking better information, we shall usually assume these errors to be independent and random. (Drift caused by changes in the electronic system is definitely not random.) Before we consider errors in general, we shall examine one that is not only important and unavoidable, but that also sets x-ray... 

X-ray emission spectrography is no different from other analytical methods in having its reliability threatened by errors of different kinds. 

The refinement of other analytical methods, such as electrophoresis [34,36], the various techniques of optical spectroscopy [103-105], and nuclear magnetic resonance [201], is supplemented by the recent advances in real-time affinity measurements [152,202], contributing to the understanding of biomolecular reactivity. Taken together, the improvement of analytical methods will eventually allow a comprehensive characterization of the structure, topology, and properties of the nucleic acid-based supramolecular components under consideration for distinctive applications in nanobiotechnology. 

Within collaborative work on element concentrations in a number of biological reference materials using solid sampling and other analytical methods, calibration of Cd, Cu, Pb and Zn in BCR CRM 185 Bovine liver with solid CRMs was performed for each element with a reference material of the same matrix, NIST SRM 1577... 

Schauenburg H, Weigert P (1991) Determination of element concentrations in biological reference materials by solid samphng and other analytical methods. Fresenius Anal Chem 342 950-956. 

Consistent with other analytical methods, immunoassays must be validated to ensure that assay results are accurate. Initial validation involves an evaluation of the sensitivity and specificity of the immunoassay, while later validation includes comparison with a reference method. Because a goal of immunoassays is to minimize sample preparation, validation also includes testing the effects of sample matrices and(or) sample cleanup methods on results. The final steps in validation involve testing a limited number of samples containing incurred residues to determine if the method provides reliable data. 

There are a few other analytical methods in which electrochemistry plays an essential role, such as (paper) electrophoresis, isotachophoresis, electrography and electrochromatography (according to Fujinaga) as they belong to analytical separation techniques, they are beyond the scope of this book. 

On the basis of the above, the advantages of polarographic analysis over other analytical methods can be summarized as follows ... 

Flow techniques have become of considerable importance, not only in routine titrations but also in other analytical methods as automated analytical processes they all need to be under the control of a detector, often called a sensor and sometimes a biosensor. We can divide the techniques into the following ... 

The various methods that have been outlined in the previous sections are not exclusive and other analytical methods have been used for the determination of methylxanthines in food systems. One of the most widely used methods for food analysis is flow injection analysis (FIA). In a study by Numata,43 a flow injection analysis method for the determination of hypoxanthine in meat was described. 

Results obtained by CPAA for composition and partial mass thickness have been shown to be consistent with the results obtained via other analytical methods. The main advantage of the use of CPAA as a surface characterization method are its purely instrumental character, requiring no sample preparation, its high accuracy, and its low detection limits. 

This expression contains four quantities n, D, v, and Cx. Since n is normally known for a given electrode reaction, and v can be experimentally determined with a viscometer, the slope permits one to determine the concentration of the diffusing ion, CrJ0, if its diffusivity, D is known. Conversely, one may use the slope to determine the diffusivity, D, if the bulk concentration, Cx can be measured by the other analytical methods. 

Capillary electrophoresis has also been combined with other analytical methods like mass spectrometry, NMR, Raman, and infrared spectroscopy in order to combine the separation speed, high resolving power and minimum sample consumption of capillary electrophoresis with the selectivity and structural information provided by the other techniques [6]. 

In contrast to other analytical methods, ion-selective electrodes respond to an ion activity, not concentration, which makes them especially attractive for clinical applications as health disorders are usually correlated to ion activity. While most ISEs are used in vitro, the possibility to perform measurements in vivo and continuously with implanted sensors could arm a physician with a valuable diagnostic tool. In-vivo detection is still a challenge, as sensors must meet two strict requirements first, minimally perturb the in-vivo environment, which could be problematic due to injuries and inflammation often created by an implanted sensor and also due to leaching of sensing materials second, the sensor must not be susceptible to this environment, and effects of protein adsorption, cell adhesion, and extraction of lipophilic species on a sensor response must be diminished [13], Nevertheless, direct electrolyte measurements in situ in rabbit muscles and in a porcine beating heart were successfully performed with microfabricated sensor arrays [18],... 

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