Preservative assays

Where antioxidants or antimicrobial preservatives are used, the finished product release specification will need to include identification tests and assays for these two types of excipient. The shelf life specification should also include a specification for assay for antimicrobial preservatives. Stability data will be required for both antioxidants and antimicrobial preservatives in the finished product, and in addition the preservative efficacy of the formulated product should be examined over its shelf life and by means of appropriate in-use stability tests. Preservative efficacy data should also be presented at the lower limit of the preservative assay. 

Justify using a stability-indicating preservative assay only at all time intervals as a substitute for the USP Antimicrobial Effectiveness Test by confirming preservative efficacy at 50, 75, and 100% of label claim. 

Compendial (e.g., sterility, endotoxins, microbial limits, volume and container, particulates, content uniformity, preservative assay, moisture)... 

Quantitative preservative assay limits based on analytical capability and levels required for antimicrobial preservative efficacy... 

T Assay/Inspection, Potency, Degradation Products, Preservative Assay, pH, Dissolution, Redispersibility (suspension only), Mean size and Distribution of Particles (as appropriate)... 

Urine may be collected for assays of enzyme activities following cleansing of the genitalia with mild antiseptic soap followed by rinsing with water. The urine is collected in a chemically clean container with no preservative. As the activity of urinary enzymes is a function of the volume of the specimen it is important to time the collection accurately. A collection period of 8 hours is quite adequate, and the use of longer periods is not desirable because enzyme activities can rapidly decrease in the relatively hostile medium of the urine. The urine should be refrigerated and transferred promptly to the laboratory, where it should also be processed promptly. 

Screening the molecular heterogeneity of receptor expression in endothelial cell surfaces is required for the development of vascular-targeted therapies. First, as opposed to targeting purified proteins as discussed above, membrane-bound receptors are more likely to preserve their functional conformation, which can be lost upon purification and immobilization outside the context of intact cells. Moreover, many cell surface receptors require the cell membrane microenvironment to function so that protein-protein interaction may occur. Finally, combinatorial approaches may allow the selection of cell membrane ligands in a functional assay and without any bias about the cellular surface receptor. Therefore, even as yet unidentified receptors may be targeted. 

Xu Y, Sato K, Mawatari K et al (2010) A microfluidic hydrogel capable of cell preservation without perfusion culture under cell-based assay conditions. Adv Mater 22 3017-3021... 

A further complication for immunolocalization studies is that the fixatives used to preserve the tissue chemically modify the antigens that are the target of the antibodies. The aldehyde fixatives attack the amino side chains of proteins that eliminate many epitopes. Osmium postfixation is far more destructive of antigenicity and, as a general rule, osmium most often permanently destroys the possibility of immunocytochemical assay. There are many examples of immunocytochemical assay being conducted on osmicated tissue (2) and many others in which the osmium is chemically removed from the sections by oxidation with periodate restoring antigenicity that had previously been masked by the bound... 

Citrated blood is diluted 1 10 with enzyme buffer solution, and preservative is added (H19). The buffer is prepared by dissolving 0.2 g of Clarase (Fisher Scientific Co., New York) in 100 ml citrate buffer (5 g potassium citrate monohydrate and 1 g citric acid monohydrate in 1000 ml distilled water, pH 5.6). The solution is incubated for 3 days at 37°. After incubation, it is autoclaved 15 minutes to stop enzymatic action and coagulate proteins. It is filtered, and 1.0, 1.5, and 2.0 ml of the supernatant is added to individual flasks and assayed. Control flasks are included to estimate pantothenic acid contamination of the enzyme. 

Enzymatic assay techniques have been developed for several additives by Merck. BIOQUANT kits are available for aspartame (intense sweetener) and nitrate (preservative). Gromes et al. (1995) applied the Bioquant kit to determination of aspartame in yoghurt, quark and confectionery. For low concentrations of aspartame a blank correction procedure was necessary. Recoveries of aspartame were in the range 93-102%. 

In plasma, Lp(a) moves with a pre-beta-1 mobility (B8, B9) and has been referred to as the sinking pre-beta fraction, as it shares the mobility of VLDL yet does not float at the same density in the ultracentrifuge. Serum electrophoresis followed by a lipid staining with Oil Red O or Fat Red should be a convenient method for detection of elevated Lp(a) levels. Sample collection and preservation are, however, critical parameters in this assay (B26, K9). 

Type 1 assay and identification of the main component(s) (active, preservatives and key excipients). 

Methods for the quantitation of major components of bulk drug substances or APIs, including preservatives, in finished drug products are classified in Category I. Assay and content uniformity methods fall into this category. These methods demonstrate that the label claim of the drug... 

Hence, a generic or universal HPLC method interfaced with a UV detector for BHT can be used on any drug as long as the acceptance criteria on accuracy, precision, robustness, and other necessary requirements have been met. Similar to appearance, drug release, assay, and impurities, preservative testing is also required if a certain degree of preservative has to be included in the drug product to ensure an adequate shelf life. 

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