Protein cloudiness

Copper. In the presence of sulfur dioxide, copper-protein cloudiness may develop in white wines. Only small amounts of copper (about 0.3 to 0.5 mg/liter) cause cloudiness. Widespread use of stainless steel in modern wineries has reduced copper pickup, but many wineries routinely test their wines for copper. Atomic absorption spectrophotometry is the method of choice (51) although reducing sugars and ethanol interfere, and correction tables must be used (107). To reduce this interference, chelating and extracting with ketone is recommended (108). Lacking this equipment colorimetric procedures can be used, especially with di-ethyldithiocarbamate (3, 4, 6, 9,10, 22,109). Neutron activation analysis has been used for determining copper in musts (110). 

Hauptmann (1952b) recommended a preparation low in pectase for red wines so that the extract and ash content of the wine would not be too high. With the proper enzyme he found the red wines of better color and flavor. For white wine clarification no protease should be present. Better clarification and freedom from protein cloudiness in the bottle was reported for the treated wines. 

Purification of photoprotein. The dialyzed photoprotein solution was centrifuged to remove precipitates, and then subjected to fractional precipitation by ammonium sulfate, taking a fraction precipitated between 30% and 50% saturation. The protein precipitate was dissolved in 50 ml of 10 mM sodium phosphate, pH 6.0, containing 0.1 mM oxine ( pH 6.0 buffer ), dialyzed against the same buffer, and the dialyzed solution was adsorbed on a column of DEAE-cellulose (2.5 x 13 cm) prepared with the pH 6.0 buffer. The elution was done by a stepwise increase of NaCl concentration. The photoprotein was eluted at 0.2-0.25 M NaCl and a cloudy substance (cofactor 1) was eluted at about 0.5 M NaCl. The photoprotein fraction was further purified on a column of Sephadex G-200 or Ultrogel AcA 34 (1.6 x 80 cm) using the pH 6.0 buffer that contained 0.5 M NaCl. 

Add 100 pi of the NHS-iminobiotin solution to each ml of the antibody solution. Mix well to dissolve. Note Some turbidity may be present in the reaction due to incomplete dissolution of the NHS-iminobiotin. The solution may look cloudy or have a microparticulate suspension present. This is normal for many water-insoluble reagents when added to an aqueous solution in an organic solvent. As the reaction takes place, the NHS-iminobiotin will be driven into solution, both by coupling to the protein and by hydrolysis of the NHS ester. 

Under physiological conditions, CSF is clear and colorless. It may be pink or red if many red blood cells (RBCs) are present, and cloudy if more than 400 elements/mm are present, white blood cells (WBCs) are present, or the protein content is especially high. When blood has been present in the CSF for more than four hours, xanthochromia may occur owing to the presence of hemoglobin pigment from lysed red blood cells. Also, high protein levels (>1.5 g L ) may produce a yellowish color that can simulate xanthochromia of red blood cell origin. 

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... 

The cloudy reaction sample is cooled (4°C) for 15 min and then spun at 18,000 g (Eppendorf centrifuge) for 2 min to pellet the precipitated serum proteins. 

This newly fermented wine is cloudy from suspended materials such as yeast, protein, colloids, and fine grape cellular solids. In storage, a natural clarification, or gravity settling out, of these materials takes place. 

One of the major requirements in clarification is to fine wines to remove certain proteins that may cause instability in wines. These unstable proteins, if not removed from the white and rose wines, may precipitate out of the wines after bottling, causing cloudiness and sediment. This precipitation is more likely where the pH of the wine is near to the isoelectic point of... 

Wines are stabilized to prevent cloudiness from a number of causes, such as proteins, metals, colloidal materials, and bitartrates (natural salt of wine). 

If the mixed solutions are cloudy owing to the presence of precipitated protein, centrifuge the tubes for 10 minutes at 4000 rpm in a desk top clinical centrifuge. 

To the dialyzed, cloudy enzyme solution an equal volume of 1 M KPi 7.75) was added to further precipitate the urea-soluble proteins. The suspension was centrifuged and the pellet was collected and redissolved in ca. 130 ml of 6 M urea, 0.03 M NaCl, 16 mM KPi (pH 7.8) buffer prior to being loaded on a Sephacryl S-2(X) column previously equilibrated with 6 M urea, 0.03 M NaCl, 16 mM KPi (pH 7.8). The column was eluted with the same buffer and fractions were collected. TTiose containing enzyme activity or having an enriched protein species of 32 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (16) were pooled and concentrated. The concentrated eluant (20 ml) was applied to a Sephacryl S-100 column previously equilibrated with 6 M urea, 0.03 M NaCl, 16 mM KPi, pH 7.8. The column was eluted with the same buffer, fractions were collected and those containing only 32 kDa (LO) and 24 kDa species were pooled and concentrated. This is referred to as the "two-banded" protein, and was used without further purification (to reduce protein loss) for phenylhydrazine inactivation (see below). 

Haze formation is mostly attributed to proteins, polyphenols, and their interactions. It is also possible that there are also other factors that inbuence haze formation in beer, but their effect has not been yet clearly debned [ 15]. The amount of haze formed depends both on the concentration of proteins and polyphenols, and on their ratio. Polyphenols can combine with proteins to form colloidal suspensions that scatter light, which creates the cloudy appearance of beer. Beer polyphenols originate partly from barley and partly from hops. The beer polyphenols most closely associated with haze formation are the proanthocyanidins, which are dimers and trimers of catechin, epicatechin, and gaUocatechin. These have been shown to interact strongly with haze-active proteins [13,15-17] and their concentration in beer was directly related to the rate of haze formation [18]. Ahrenst-Larsen and Erdal [19] have demonstrated that anthocyanogen-free barley produces beer that is extremely resistant to haze formation, without any stabilizing treatment, provided that hops do not contribute polyphenols either. Not all proteins are equally involved in haze formation. It has been shown that haze-active proteins contain signibcant amounts of proline and that proteins that lack proline form little or no haze in the presence of polyphenols [13,15-17]. In beer, the source of the haze-active protein has been shown to be the barley hordein, an alcohol-soluble protein rich in proUne [16]. 

Intermediate-acting insulin Isophane Cloudy Protein 0.01-0.04 Phosphate 1-2 6-12 18-24... 

After the enzymatic treatment the juice can be clarified. Flocculation aids and fining aids (gelatine, silica sol, etc.) help to coagulate the cloudy substances and facilitate their separation by settling and filtration. Bentonites can be used to eliminate proteins and other cloudy substances. Filtration (Kieselgur precoat filtration with plate and frame filters, rotating vacuum filters, and sheet filters as police filters ) is used to produce a crystal clear juice. 

---