Detection limit related substances

Owing to poor volatihty, derivatization of nicotinic acid and nicotinamide are important techniques in the gc analysis of these substances. For example, a gc procedure has been reported for nicotinamide using a flame ionisation detector at detection limits of - 0.2 fig (58). The nonvolatile amide was converted to the nitrile by reaction with heptafluorobutryic anhydride (56). For a related molecule, quinolinic acid, fmol detection limits were claimed for a gc procedure using either packed or capillary columns after derivatization to its hexafluoroisopropyl ester (58). 

The application range of cSFC-DFI-MS (Table 7.41) appears to be restricted either to the analysis of low-MW substances or to problems related to high-MW samples where low detection limits are not needed [124,444,445], The analysis of surfactants [446] by SFC-MS is frequently performed to demonstrate the feasibility of newly developed interface technology for practical applications. A rugged cSFC-MS method has been developed for the analysis of ethoxylated alcohols (AEs), which are non-ionic surfactants incorporated into a wide variety of industrial and consumer products [447]. cSFC-DFT-DFS was used for the analysis of low-MW, thermally unstable peroxides, and the higher-MW surfactants Triton X-100 and... 

The use of spiked batches (with levels approaching the specification limit) is recommended for the transfer of impurity or related substances methodology. In parallel, the receiving site should also assess the LOD/LOQ (Limit of Detection/Limit of Quantitation) of the method for each of the specified impurities or related substances. This can help to identify whether specificity and sensitivity are issues at the receiving laboratory. 

The method detection limit (MDL) is the smallest quantity or concentration of a substance that a particular instrument can measure (Patnaik, 2004). It is related to the instrument detection limit (IDL), which depends on the type of instrument and its sensitivity, and on the physical and chemical properties of the test substance. 

Limits for detection and quantitation are usually applied to related substances in drug substances or drug products. The detection limit is the... 

Range. Ideally, linearity should be established from 50% of the ICH reporting limit to the nominal concentration of drug substance in the sample solution (for area percent method). If the linearity does not support such a wide range of concentration, determine the linearity from 50% of the ICH reporting level to 150% of the proposed shelf life specifications of the related substance (for the high-low and external standard methods) as a minimum. This will ensure a linear response for related substances at all concentration levels to be detected during stability. 

Riboflavin (vitamin I 2) is an important chemical species that occurs in many complex biologically related samples. The substance is chiral and strongly fluorescent and the coupling of these molecular properties into the FDCD-HPLC detection system greatly enhances both the specificity and the limit of detection capabilities of the method. In the specific case of riboflavin an HPLC system using FDCD detection where excitation is done at 325nm with a HeCd-laser a detection limit of 170 pg of riboflavin [30] is attainable. This level is about 10 times better than what is possible with a laser excitation -transmission CD-HPLC system [29]. 

Some pharmacopoeial specifications for pethidine hydrochloride(14)now include a tic limit test for related substances as a result of the detection by glc of homologs formed by exchange of alkyl groups between the nitrogen atom and ester function of pethidine base(15) some samples contained up to 9% impurity even though their melting points lay within the range for the pure salt. 1-Benzylnorpethidine has also been identified as a contaminant of pethidine pharmaceuticals.(16)... 

Probably no technique in analytical chemistry has been more quickly and widely adopted than gas chromatography. It is ideally suited to the separation and analysis of nonpolar volatile materials such as hydrocarbons. Factors that have contributed to this rapid development are (/) the success of the method in performing quantitative separations of closely related substances, (2) the ease with which the operation can be made automatic, (5) the speed, predsion, and accuracy with which quantitative determinations can be made, (4) the small quantities of simple required, and (5) the rapid development of sensitive detection devices. The prindpal limitation is that the method is restricted to materials that exert vapor pressures of at least 10 mm Hg at the temperature of the column (ranging up to 300°C or somewhat higher). 

The preponderance of analytical (and preparative) separations that precede mass spectrometric analyses are carried out using HPLC. Despite its potential for extremely high efficiencies, capillary electrophoresis (CE) remains something of a specialized application, particularly with mass spectrometric detection. The technique is hampered by its reliance on small sample volumes (typically nL injections) in order to avoid loss of separation efficiency. The consequence of these small volumes is a concentration detection limit that is substantially higher than that of techniques such as HPLC. Despite the exquisite sensitivity of mass spectrometers, characterization of related substances present at low femtomole levels (assuming a hypothetical related substance compound with molecular weight 500 daltons present at 0.1% w/w in an equally hypothetical drug substance sample... 

Following the establishment of specificity, the method(s) should be validated to allow for use in release and stability testing. Such validation is typically less stringent than for final methods (sec Chapter 12), but should demonstrate specificity, linearity, range, accuracy, and analysis repeatability for the API. For related substances, specificity should be demonstrated and the limit of detection (LOD) and limit of quantitation (LOQ) should be established for the API to serve as surrogate values for the LOD and LOQ of impurities for which authentic substances are not available. To achieve a sufficient LOD and simultaneously keep the API in the linear dynamic range of the detector, it may be necessary to use different sample concentrations for the analyses of the API and related substances. It is additionally beneficial to repeat the separation on new columns from different batches to ascertain that the separation obtained can be maintained column to column. 

In addition to choosing separation conditions, both the injection volume and the sample concentration must be chosen during method development. These must be considered with respect to the detection used. For the simultaneous detection of API and related substances, the detector should minimally be able to quantify compounds from 0.1% (limit of quantitation) to 130% of the concentration of the API for compliance with ICH guidelines.This requires the detector to have a minimum linear dynamic range (LDR) of 1300, assuming equal responses of the API and related substances. For samples where... 

The specific radioactivity of the ligand and the concentration of the ligand added to an RRA are intimately related in determining the sensitivity of a particular assay Given a twofold increase in the specific radioactivity of the tracer, it is possible within limits to reduce the concentration of the tracer in the assay by one-half with little or no loss in detectable radioactivity. Because the sensitivity of an RRA to detect receptor-active substances IS a function of the final concentration of the ligand in the incubation, the use of tracers of the highest specific radioactivity IS warranted... 

Among the related substances in many antibiotics are various structurally related components in the drug substance, the composite mixture of which is obtained in the synthetic or semisynthetic scheme and which gives rise to the drug efficacy. Control of the relative content of components is therefore necessary for these drugs. Test specification limits for the components are normally stated in terms of area percent of each, as maximum, minimum or a range of values for each or for the sums of several components. The types of components seen in these antibiotics were studied as impurities in the semisynthetic antibiotic clarithromycin, where detection limits of 0.1% w/w were found. Normalization factors were determined for each of 15 known related substances using ratios of the slope of linear calibrations for each substance to that for the reference impurity. 

Complications of drug analysis were successfully resolved by the combination of TLC with MALDI-MS. TLC combined with MALDI-MS was used for analysis of psychotropic drugs (3,4-methylenedioxy methamphetamine, 4-hydroxy-3-methoxy methamphetamine, 3,4-methylenedioxy amphetamine, methamphetamine, p-hydroxy methamphetamine, amphetamine, ketamine, caffeine, chlorpromazine, triazolam, and morphine) in biological samples [42]. This technique was able to analyze 3,4-methylenedioxy methamphetamine (MDMA) and its metabolites in urine samples without sample dilution, and the detection limit of the MDMA spot was 0.05 ng/ spot. Crecelius and coworkers described the use of TLC with MALDI-MS/MS for the structural analysis of small drug molecules [43]. This method was successfully applied to analyze two representatives of nonsteroidal antiinflammatory drugs (tenoxi-cam and piroxicam), and pharmaceutically active compound UK-137,457 and one of its related substances UK-124,912. The feasibility of UTLC-atmospheric pressure (AP)-MALDI-MS was described for the analysis of small molecules (triazole, midazolam, verapamil, and metaprolol) [44]. The authors compared the selectivity and sensitivity between UTLC- and HPTLC-AP-MALDI-MS. It was observed that UTLC plates provided 10-100 times better sensitivity in MALDI analysis than the conventional... 

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