Impurity reference standards

While compendial standards are available for some monographed article impurities, it may be difficult at times to obtain pure standards of impurities. Manufacturers of pharmaceuticals function as a potential source for obtaining reference standards of impurities, which may be synthesis precursors, process intermediates, or degradation products. The characterization and evaluation of these impurities reference standards should be constant with their intended use. In many cases, analytical procedures are developed and validated, where the response of an impurity is compared to that of the new drug substance itself. Response factor evaluation of impurities at the chosen detection wavelength is necessary to determine if a correction factor is needed (when the responses differ). Potentiometric detection, fluorescence/ chemiluminescence detection, and refractive index detection are some examples of detection modes available for compounds that may not be suitable for UV detection. 

When impurity reference standards are available only in limited quantities, relative response factors (RRFs) to the active ingredient can be used to quantitate impurity concentrations. RRFs can be determined spectrophotometrically by comparing the molar absorptivity of the impurity to that of the active component. However, in our experience, RRFs determined by HPLC by comparing peak area responses of the impurity to those of the active ingredient have been more accurate than those determined by spectrophotometric method. 

Area normalization against the API External standardization using well-qualified impurity reference standards... 

Analytical and Laboratory Operations. Sulfamic acid has been recommended as a reference standard in acidimetry (55). It can be purified by recrystaUization to give a stable product that is 99.95 wt % pure. The reaction with nitrite as used in the sulfamic acid analytical method has also been adapted for determination of nitrites with the acid as the reagent. This reaction is used commercially in other systems for removal of nitrous acid impurities, eg, in sulfuric and hydrochloric acid purification operations. 

Schirmer has succinctly summarized the strengths and limitations of phase solubility analysis [40]. The principal advantages are that (1) a reference standard known purity is not required, (2) the number and types of impurities in the sample need not be known, (3) all required solubility information is obtained from the analysis, (4) the technique can be applied to the analysis of any solute that can be dissolved in some solvent, (5) the deduced results are both precise and... 

CE is applied to two major categories of quality release testing identity and impurity testing. Identification assays are intended to ensure the unique identity of an analyte in the sample. This is normally achieved by comparison of a property of the sample (e.g., spectrum, chromatographic behavior, chemical reactivity, etc.) to that of a reference standard. As shown in Figure 9, CZE can be used to determine identity for monoclonal antibodies and proteins based on their unique electrophoretic profile. 

The most common technique for monitoring impurities is HPLC with UV detection. Quantification of impurities is achieved by reference standard, when available, or by area percent or height percent relative to the parent compound. Figure 1.4 shows a typical chromatogram enlarged to show the various impurities. One of the impurities is the USP Compound B that is added to the furosemide drug substance. 

More than 220 producers of CRMs throughout the world produce today 12,000 20,000 materials with dif ferent matrixes, analytes and properties [4]. However, many testing (analytical) laboratories cannot find suitable CRMs in the market and develop in-house reference materials (IHRMs) themselves. Often IHRMs are developed in a laboratory to conserve the corresponding expensive CRMs. For example, a pharmaceutical company Chemagis Ltd. produces 30 active pharmaceutical ingredients steroids, benzodiazepines, antihistamines, hipolipidaemics, blood flow reactants, etc. Only for a few of them Mo-metasone Furoate, Fluticasone Propionate and Dobutamine Hydrochloride are of fi-cial reference standards for assay supplied by US, British and European Pharmacopoeias with prices of about 180 per unit (50 200 mg). Thus, to support its customers Chemagis is forced to develop IHRMs for assay as well as for impurities and related substances of each produced compound. Therefore, certification of such IHRMs that leads to traceable values is very important. 

A listing of reference standards, system suitability standards, impurities, and degradants, where applicable. List the purity and source, allowing for equivalency where applicable. 

An internal standard may contain trace amount of the analyte of interest, especially a SIL internal standard due to similar synthesizing routes. In this case, a maximum IS concentration can be obtained based on its impurity (e.g., n % of the analyte in the IS reference standard) and the 20 % acceptance criterion for bias at the LLOQ level [13,16-17], In this regard, the concentration of an IS should be therefore kept as low as possible to reduce its contribution to analyte concentration. 

On the other hand, the analyte or analyte reference standard may contribute to the response of an internal standard either due to impurity or because of natural abundance of stable isotopes (say m %). Therefore, the concentration of the internal standard must not be too low. Otherwise, the contribution from the analyte or its reference standard would be significant, e.g., more than 5 % of IS concentration or response [16], and linearity and accuracy could be greatly affected. Accordingly, a minimum IS concentration could be obtained based on the ULOQ for the analyte. For this reason, a high IS concentration is preferred. 

Once an internal standard is selected, its concentration must be properly determined based on the particular situation, such as impurities of reference standards, concentration range, and detection sensitivity towards the analyte and its internal standard, and finally experimentally verified to ensure adequate linearity, accuracy, and precision. 

The primary reference standard is normally prepared on a laboratory scale using pure starting materials, reagents, and solvents and should be of the highest purity that reasonably can be obtained. The synthetic procedure used to make it and the method(s) used for its purification, also should be provided. (If applicable, the method of manufacture section can be referenced.) The purification procedure is normally performed until little or no change is observed through two consecutive cycles in assay purity and levels of impurities. 

Reference Standard. Characterization and structure elucidation data are typically derived from tests conducted on the primary reference standard. Its suitability must be documented by information much more extensive than prescribed in the specifications. In addition to the prescribed analyses—especially levels of impurities—other tests normally conducted include elemental analysis and ultraviolet, infrared, nuclear magnetic resonance, and mass spectrometry, along with reviews of each providing assignments of the important features supporting the structure(s) of the drug substance. Other tests such as... 

Identify all samples being set aside for FDA validation. The samples should include drug substance, drug product, major impurities, and degradation products being controlled for, references standard, and internal standard (the latter is not required if commercially available but is recommended to facilitate FDA laboratory work). If appropriate, blanks and any other materials not commercially available but specified in the analytical procedures should be provided. The samples are to be maintained by the sponsor until the FDA s reviewing chemist provides instructions as to where they should be forwarded. The total quantities and the manner of their subdivision (e.g., 400 tablets, 4 x 100 tablets/ bottle) should be indicated. The amounts provided should be adequate to permit at least three separate determinations, excluding sterility, by two different laboratories. 

Impurity Standard Preparation Transfer about 25 mg of USP 5-Benzyl-3,6-dioxo-2-piperazineacetic Acid Reference Standard, accurately weighed, into a 100-mL volumetric flask. Add 10 mL of methanol, and dissolve. Dilute to volume with water, and mix. Pipet 15 mL of this solution into a 50-mL volumetric flask, dilute to volume with Diluting Solvent, and mix. Use a freshly prepared solution. 

However, these titration methods can be used in early development when a reference standard is not available. Also, the spectrometric-based assay methods such as ultraviolet (UV) may be nonspecific because most of the drug substance impurities contain a similar chromophore as the parent molecule. If UV is used, UV absorption is measured at one or more wavelengths and the absorbance value is recorded for a particular concentration. Sandor Gorog has critically evaluated the difference between specific and nonspecific assay methods in the European and US Pharmacopoeias [23]. The difference between the mean and the accepted true value with a defined confidence interval should be reported in the acceptance criteria. 

Low-Level Linearity Versus High-Level Linearity When Do You Use a Dilute Standard to Calculate Impurity Levels There are two ways to determine the related substances one in terms of mass percent using a reference standard (at 100% level of API or dilute reference standard) and the... 

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