HPLC method

The major point of these HPLC methods is that they separate the aluminium fluoride species from the other inorganic monomeric aluminium species (classic Driscoll methods do not). 

NMR techniques have been utilized by Bertsch et al. [180] for the determination of Al concentrations and for other Al(OH) species. Problems with maintaining identical instrumental conditions, differences between acidified and unacidified solutions and low sensitivity are the main drawbacks. However, good agreement with values obtained by the HPLC technique for the uncoupled A1(H20)6 species were obtained [172]. The authors concluded that their method may be particularly useful for aluminium speciation in 

Summary of some published intermethod comparison exercises for aluminium speciatiorf 

Computational F vs. Oxine/dialysis At pH 5.5, if = 0.82 for inorganic monomeric aluminium 

Dialysis vs. Driscoll/PCV For inorganic monomeric aluminium = 0.99. Dialysis underestimates PCV values by 6%. 

HPLC log P techniques, first described by Mirrlees et al. [374] and Unger et al., [375], are probably the most frequently used methods for determining log/1. The directly measured retention parameters are hydrophobicity indices, and need to be converted to a log P scale through the use of standards. The newest variants, breadths of scope, and limitations have been described in the literature [292-298]. A commercial automated HPLC system based on an extension of the approach described by Slater et al. [150] has just introduced by Sirius (www. sirius-analytical. com). 

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Remcho, V. T. McNair, H. M. Rasmussen, H. T. HPLC Method Development with the Photodiode Array Detector, /. Chem. Educ. 1992, 69, A117-A119. 

Bohman and colleagues described a reverse-phase HPLC method for the quantitative analysis of vitamin A in food using the method of standard additions. In a typical example, a 10.067-g sample of cereal is placed in a 250-mL Erlenmeyer flask along with 1 g of sodium ascorbate,... 

Snyder, L. R. Glajch, J. L. Kirkland, J. J. Practical HPLC Method Development. Wiley-Interscience New York, 1988. 

Monobasic acids are determined by gas chromatographic analysis of the free acids dibasic acids usually are derivatized by one of several methods prior to chromatographing (176,177). Methyl esters are prepared by treatment of the sample with BF.—methanol, H2SO4—methanol, or tetramethylammonium hydroxide. Gas chromatographic analysis of silylation products also has been used extensively. Liquid chromatographic analysis of free acids or of derivatives also has been used (178). More sophisticated hplc methods have been developed recentiy to meet the needs for trace analyses ia the environment, ia biological fluids, and other sources (179,180). Mass spectral identification of both dibasic and monobasic acids usually is done on gas chromatographicaHy resolved derivatives. 

Chromatographic Method. Progress in the development of chromatographic techniques (55), especially, in high performance Hquid chromatography, or hplc, is remarkable (56). Today, chiral separations are mainly carried out by three hplc methods chiral hplc columns, achiral hplc columns together with chiral mobile phases, and derivatization with optical reagents and separation on achiral columns. All three methods are usehil but none provides universal appHcation. 

High Performance Liquid Chromatography (hpic). Hplc is currently the fastest growing analytical method and is now available in many laboratories. DL-Analysis by hplc has already been described and hplc methods have been reviewed (122). 

Body fluids are analyzed for T and T by a variety of radioimmunoassay procedures (31) (see Immunoassays). The important clinical parameter for estimating thyroid function, the protein-bound iodine (PBI), is measured as described in treatises of clinical chemistry. High performance Hquid chromatographic (hplc) methods have replaced dc (32,33). 

Determination. Various classical techniques are used for the analysis of vanillin, including colorimetric, gravimetric, spectrophotometric, and chromatographic (tic, gc, and hplc) methods. The Food Chemical s Codex (FCC) prescribes infrared spectrophotometry for identifying and testing vanillin. However, more vanillin analyses are made by either gc or hplc. 

For more specific analysis, chromatographic methods have been developed. Using reverse-phase columns and uv detection, hplc methods have been appHed to the analysis of nicotinic acid and nicotinamide in biological fluids such as blood and urine and in foods such as coffee and meat. Derivatization techniques have also been employed to improve sensitivity (55). For example, the reaction of nicotinic amide with DCCI (AT-dicyclohexyl-0-methoxycoumarin-4-yl)methyl isourea to yield the fluorescent coumarin ester has been reported (56). After separation on a reversed-phase column, detection limits of 10 pmol for nicotinic acid have been reported (57). 

Various aspects of the chromatography of vitamin B 2 and related corrinoids have been reviewed (59). A high performance Hquid chromatographic (hplc) method is reported to require a sample containing 20—100 p.g cyanocobalamin and is suitable for premixes, raw material, and pharmaceutical products (60). 

The successful separation of xanthate-related compounds by high performance Hquid chromatography (hplc) methods has been reported (91—93). The thin-layer chromatography procedure has been used to determine the nature of the alcohols in a xanthate mixture. A short mn of 3 cm at a development time of 25 min gives a complete separation of C —alkanol xanthates (94). 

Therefore, hplc methods seem more effective. By usiag a combiaed uv and electrochemical detection technique (52), the gem-chlotinated cyclohexadienones, the chlorophenols, and the phenoxyphenols present ia the chlorination mixtures can be determined with great accuracy. 

L. R. Snyder,. L. Glajsch, and. J. Kirkland, Practical HPLC Method Depe/opmen( Wiley-Interscience, John Wiley Sons, Inc., New York, 1988. 

A high performance Hquid chromotography (hplc) method to determine citric acid and other organic acids has been developed (46). The method is an isocratic system using sulfuric acid to elute organic acids onto a specific hplc column. The method is sensitive for citric acid down to ppm levels and is capable of quantifying citric acid in clear aqueous systems. 

The comparison of analytical characteristics HPLC methods of determination of phenols with application amperometric and photometric detectors was caiiy out in this work. Experiment was executed with use liquid chromatograph Zvet-Yauza and 100 mm-3mm 150mm-3mm column with Silasorb C18 (5 10 p.m). With amperometric detector phenols were detected in oxidizing regime on glass-cai bon electrodes. With photometric detector phenols were detected at 254 nm. 

HPLC method with amperometric detection was applied for detenuination of phenols in sea sediment and some dmg preparation. Peaks of phenol, guaiacol, cresols, hydroquinon and resorcinol were identified on chromatogram of birch tai. The HPLC method with electrochemical detectors was used for detenuination of some drug prepai ation of aminophenol derivate. So p-acetaminophenol (paracetamol) was determined in some drug. 

There have been compared the methods of mycotoxin control in food products with aflatoxin as an example, using both HPLC method with fluorescent detecting on the apparatus Thermo FL 3000 with a column BDS Hypersil C 2.1x150, as well as a chromatodensitometry method on the apparatus CAM AG TLS Scanner 3. 

There have been also found the quantitative characteristics of the methods. They are as follows for HPLC method the linearity is 0.1 ng to 2 ng the detecting limit is 0.1 ng the limit of the quantitative estimation makes up 0.0004 mg/kg a coefficient of variation is 2.74% for the chromatodensitometry method the linearity is 2 ng to 10 ng the detecting limit is 0.6 ng the coefficient of variation is 2.37%. The data obtained have been treated using a regressive analysis. 

The comparison of the obtained quantitative parameters of the methods evidences that HPLC method is better by its perceptibility. However, the chromatodensitometry method is more efficient by the indices of expressity, as far as in a routine analysis it makes it possible to conduct a greater amount of tests during the same period of time, as well as by the criterion cost -efficiency. 

We proposed using MLC for assay of azithromycin in tablets and capsules. As alternative conventional reversed-phase HPLC method MLC was used for analysis of Biseptol (sulfamethoxazole and trimethoprim) tablets and injection. The MLC was proposed to assay of triprolydine hydrochloride and pseudoephedrine hydrochloride in tablets as alternative normal-phase HPLC method described in USP phamiacopoeia. 

A selective, sensitive and stability indicating reversed phase-HPLC method was developed for the determination of clarithromycin antibiotic in human plasma. 

L.R. Snyder, J. Kirkland and J. Glaich, Practical HPLC Method Development, 2nd Edn., Wiley-Interscience, New York, 1997. ISBN 047100703X. 

In the development of a SE-HPLC method the variables that may be manipulated and optimized are the column (matrix type, particle and pore size, and physical dimension), buffer system (type and ionic strength), pH, and solubility additives (e.g., organic solvents, detergents). Once a column and mobile phase system have been selected the system parameters of protein load (amount of material and volume) and flow rate should also be optimized. A beneficial approach to the development of a SE-HPLC method is to optimize the multiple variables by the use of statistical experimental design. Also, information about the physical and chemical properties such as pH or ionic strength, solubility, and especially conditions that promote aggregation can be applied to the development of a SE-HPLC assay. Typical problems encountered during the development of a SE-HPLC assay are protein insolubility and column stationary phase... 

The specification development process is a data-driven activity that requires a validated analytical method. The levels of data needed include assay precision, replicate process results (process precision), and real-time stability profiles. A statistical analysis of these data is critical in setting a realistic specification. Most often, aggregation and fragmentation degradation mechanisms are common to protein and peptide therapeutics. Therefore, the SE-HPLC method provides a critical quality parameter that would need to be controlled by a specification limit. 

Stability constants of metal complexes of 9-hydroxy-4//-pyrido[l,2-n]pyrimidin-4-one [Ni(II), Co(II), Zn(II), and Cd(II)] were determined by potentiometric and polarographic investigations (93JCC283). The distribution coefficient of risperidone (11) in H20- -octanol at pH 7.4 (log D — 2.04) was determined by an RP-HPLC method (01JMC2490). 

A related substance of quinapril, 3-methyl-a-(2-phenylethyl)-l,4-dioxo-1,3,4,6,11,1 lfl-hexahydro-2//-pyrazino[l,2-A]isoquinoline-2-acetic acid, was determined by a HPLC method in drug substance (00MI55). 

Enantiomeric separations have become increasingly important, especially in the pharmaceutical and agricultural industries as optical isomers often possess different biological properties. The analysis and preparation of a pure enantiomer usually involves its resolution from the antipode. Among all the chiral separation techniques, HPLC has proven to be the most convenient, reproducible and widely applicable method. Most of the HPLC methods employ a chiral selector as the chiral stationary phase (CSP). 

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