Protein stability tests

First, laboratory testing is conducted to ascertain the stability of the wine. Like tests for protein stability, tests for determining stability and method for correcting instability vary from winery to winery. Berg (34) suggested that a wine stored at — 4° C for four days, without a bitartrate crystalline deposit, may be considered stable. The wines usually are allowed to warm to room temperature before test results are read. Absence of crystals indicates stability. A quantitative method, the concentration product (36), also can be used to evaluate tartrate stability. 

Sarmento, M.R., Oliveira, J.C., Slatner, M. and Boulton, R.B. (2000) Influence of intrinsic factors on conventional wine protein stability tests, Food Control, 11, 423-432. 

Table 5.6. Turbidity levels (NTU) after different protein stability tests carried ont on a Sauvignon Blanc wine during barrel aging on the lees (V. Moine-Ledoux, 1997, unpublished results)...

Treating juice with bentonite is recommended for wines which are to be clarified shortly after the completion of alcoholic fermentation. Additional handling of the wine is avoided in this manner at time when the wine is thought to be more fragile. However, protein stability tests carried out on juice are imprecise and therefore not practical. The same bentonite concentration is generally used on juices from a given winery. In spite of this treatment, white wines are sometimes unstable at bottling and require an additional bentonite treatment. 

Fig. 3.2.7 Left panel Effects of temperature on the luminescence intensity and stability of the protein P from Meganyctiphanes. The initial light intensity was measured with F plus P in 5 ml of 20 mM Tris-HCl/0.15 M NaCl, pH 7.5, at various temperatures. In the stability test, P was kept at the indicated temperature for 10 min, then mixed with 5 ml of 25 mM Tris-HCl/1 M NaCl, pH 7.59, containing F, to measure initial light intensity. Right panel Effect of the concentration of salts on the light intensity of the luminescence of F plus P, in 25 mM Tris-HCl, pH 7.6, at near 0°C. In the case of NaCl, the light intensity decreased to about a half after 10 min. From Shi-momura and Johnson, 1967, with permission from the American Chemical Society.

Quality of biotechnology products, analysis of the expression construct in cells used for production of r-DNA derived protein product Quality of biotechnological products stability testing of biotechnological/Biology products Availability of Draft Guideline on Quality of... 

Finally, the protein assay for the drug product will also be used for realtime and accelerated stability testing if it has been validated to be stability indicating. A stability-indicating protein concentration method usually translates to a method that can reveal how much protein can be recovered from the dosage form. Many protein instabilities result in precipitation of the protein and adsorption to the container. An instability that results in only a modification of the protein structure but not in loss of protein from solution will not be detected by a sequence-independent protein assay such as a colorimetric assay. 

Physical properties of the protein structure should be considered in designing strategies to achieve stable formulations because they can often yield clues about which solution environment would be appropriate for stabilization. For example, the insulin molecule is known to self-associate via a nonspecific hydrophobic mechanism66 Stabilizers tested include phenol derivatives, nonionic and ionic surfactants, polypropylene glycol, glycerol, and carbohydrates. The choice of using stabilizers that are amphiphilic in nature to minimize interactions where protein hydrophobic surfaces instigate the instability is founded upon the hydro-phobic effect.19 It has already been mentioned that hydrophobic surfaces prefer... 

The protein analyzer tests temperature stability and accuracy at three set points over lOmin (e.g., 30—40—50°C). The temperature linearity is collected on the stability data as well. 

The effect of dietary protein on diazinon toxicity was evaluated in a study with male albino Wistar rats. The study concluded that a purified protein test diet (with 26% casein and 59% cornstarch) did not significantly alter the LD50 value (415 mg/kg) for diazinon for this species. However, a low protein purified test diet (3.5% casein, 82% cornstarch), lowered the LD50 to 215 mg/kg. In addition, this study found that diazinon samples that were time-of-manufacture stabilized (to prevent spontaneous degradation to more toxic monothiotetraethyl pyrophosphate) were less toxic (LD50 value = 466 mg/kg) than samples stabilized after manufacture (LD50 value = 271 mg/kg) (Boyd and Carsky 1969). A subsequent study... 

Commercial wines are commonly tested for protein stability. Wine proteins, upon denaturation by heat or cold, may cause cloudiness and unsightly deposits after bottling. In addition, proteins may combine with iron and copper salts to form flocculate material in bottled wines. The reaction and absorption of proteins on bentonite is an effective means of removing protein from wines (109, 110, 111). Therefore, fining wines... 

Unfortunately, in spite of the published literature on wine proteins, we do not know the actual protein levels at which table or dessert wines are stable. The changes in protein content during production and processing of wines are still not known with sufficient accuracy to predict their behavior. The winemaker has to depend on empirical tests if he is to produce protein stable wines. Early separation of new wines from their fermentation yeast greatly improves their chances for protein stability by decreasing the release of yeast autolysis products into the wine. 

Stamper GF, Lambert WJ. Accelerated stability testing of proteins and peptides pH-stability profile of insulinoptropin using traditional Arrhenius and non-linear fitting analysis. Drug Dev Ind Pharm 1995 21 1503-1511. 

Cytochrome c (cyt. c) has become a major protein for testing new approaches and techniques in protein science.1 This is partly due to the venerable position that cytochrome c holds in the field of biochemistry since it was isolated and characterized more than 70 years ago. Cyt. c was one of the first proteins to be sequenced,2 and to have its X-ray structure determined in 1967.3 Cyt. c also has the advantage of stability and a spectroscopically distinct heme group. More than 23,000 articles mentioning cyt. c were published between 1945-2002 (ISI Web of Science). Here, we describe an approach to tetraphenylporphyrin-based protein surface receptors and the characterization of their interactions with the principal target cyt. c. 

Other organic solutes found in thermophilic archaea likely play roles as thermoprotectants, although there have been few in vitro studies in which the effects of these solutes on protein stability have been tested. Among the putative thermoprotectants in hyperthermophilic archaea is di-myo-inositol-1,1 -phosphate (DIP) (see figure 6.2 for the structure of this... 

Complementary to the simple adsorption test, the mass balance of the separation could be investigated. This method is more accurate to determine the loss of free drug, which may occur due to variations in sample preparations methods, non-specific protein binding and to metabolism. The latter may also be tested in separate stability tests in plasma or applied matrix. 

An example for an automated stability test in plasma is described by Linget and du Vignaud (1999). Incubations are performed on a 215 Gilson liquid handler. Incubation was done at substrate concentrations of 50 pM on 96 deep well plates. Each incubation tube contained 375 pL of a 200 pM test compound solution (in 0.1 M Tris buffer with 3% BSA, added to assist dissolution of compounds with poor solubility) and 1125 pL of plasma. Samples are taken after incubation times of 0, 1, 2, 3, 4 and 5 min. At each of these time points an aliquot of the incubation mixture was transferred from the incubation tube into a well in a 96 deep well plate containing an equal volume of acetonitrile for quenching by protein precipitation followed by centrifugation of the plates. Supernatants were analyzed by HPLC for metabolic screening. 

Q5C in Cell Lines used for Production of r-DNA derived Protein Products Quality of Biotechnological Products Stability Testing of Biotechnological/ CPMP/ICH/138/95 Step 5... 

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