Enzyme washing

The mixture was filtered and the enzyme washed with MTBE until the filtrate turned colorless. The filtrate was then successively washed with 1 M aqueous HCl, saturated NaHCOa and brine and then concentrated and deprotected directly without further purification (typical yield 80 %). 

The mixture was filtered, the enzyme washed with CH2CI2 and the filtrate concentrated. 

Detergent enzyme performance is often reported in the form of such dose-response curves. The performance increases dramatically at the beginning, but reaches a maximum levd at higher enzyme concentrations. The extent to which the enzyme is able to remove stains from the fabric depends on the detergent system, temperature, pH, washing time, wash load, etc. Enzyme wash performance varies between liquid and powder detergents and with the composition of the soiling (Fig. 6). 

Removal of unconjugated enzyme wash on funnel gel filtration chromatography or Concanavalin A-Sepharose chromatography... 

Hydrolysis using chitinolytic enzymes Washing and I drying j... 

Newhouse ML, Tagg B, Pocock SJ (1970) An epidemiological study of workers producing enzyme washing powders. Lancet K689-693... 

In fact, most RIAs and many nonisotopic immunoassays use a competitive binding format (see Fig. 2). In this approach, the analyte in the sample to be measured competes with a known amount of added analyte that has been labeled with an indicator that binds to the immobilized antibody. After reaction, the free analyte—analyte-indicator solution is washed away from the soHd phase. The analyte-indicator on the soHd phase or remaining in the wash solution is then used to quantify the amount of analyte present in the sample as measured against a control assay using only an analyte-indicator. This is done by quantifying the analyte-indicator using the method appropriate for the assay, for example, enzyme activity, fluorescence, radioactivity, etc. 

EIAs can be used per se or with a spectrophotometer. Traditionally, EIAs have been developed in 96-weU microtiter plates which provide the immobilization support for the assay, the reaction vessel, and, when linked to a spectrophotometer-based reader, a rapid means to detect and quantify the color resulting from interaction of a substrate with the antibody—antigen—enzyme complex. Automated immunoassay analyzers targeted primarily for use in the clinical laboratory have taken automation one step further, utilizing robotics to carry out all reagent additions, washings, and final quantification including report preparation. 

Enzyme immunosensors are used in flow injection systems and Hquid chromatography to provide automated on-line analyses (71—73). These systems are capable of continuously executing the steps involved in the immunoassays, including the binding reactions, washing, and the enzyme reaction, in about 10 minutes. 

In the wet method, as practiced in Colombia, freshly picked ripe coffee cherries are fed into a tank for initial washing. Stones and other foreign material are removed. The cherries are then transferred to depulping machines which remove the outer skin and most of the pulp. However, some pulp mucilage clings to the parchment shells that encase the coffee beans. Fermentation tanks, usually containing water, remove the last portions of the pulp. Fermentation may last from twelve hours to several days. Because prolonged fermentation may cause development of undesirable flavors and odors in the beans, some operators use enzymes to accelerate the process. 

Ultrafiltration (qv) (uf) is increasingly used to remove water, salts, and other low molecular-weight impurities (21) water may be added to wash out impurities, ie, diafiltration. Ultrafiltration is rarely used to fractionate the proteins because the capacity and yield are too low when significant protein separation is achieved. Various vacuum evaporators are used to remove water to 20—40% dry matter. Spray drying is used if a powdery intermediate product is desired. Tyophilization (freeze-drying) is only used for heat-sensitive and highly priced enzymes. 

A given enzyme may be assayed by its action on soluble substrates under chemical and physical conditions different from those encountered in a real-life wash. Such experiments indicate the enzyme s performance with respect to pH and temperature variations, or in conjunction with other soluble substances, etc. The analytical data thus obtained are not necessarily representative of the wash performance of the enzyme, and real wash trials are necessary to evaluate wash performance of detergent enzymes. 

Fig. 6. Washing performance on different soilings of a U.S. liquid detergent (B) and a U.S. powder detergent (H in a Terg-o-tometer operating at 20°C for 10 min one enzyme dosage. EMPA 117 (nulk, blood, and ink on polyester/cotton) EMPA 116 (milk, blood, and ink on 100% cotton) grass on (100% cotton) AS 10 (milk, oil, and pigments on 100% cotton) blood soilings (on 100% cotton).

AH detergent proteases are destabilized by linear alkylbenzenesulfonate (LAS), the most common type of anionic surfactant in detergents. The higher the LAS concentration and wash temperature, the greater the inactivation of the enzyme. The presence of nonionic surfactants, however, counteracts the negative effect of LAS. Almost aH detergents contain some nonionic surfactant therefore, the stabHity of proteases in a washing context is not problematic. 

Most ingredients in a detergent formulation contribute to the ionic strength of the wash solution. The effect of ionic strength on protease performance depends on pH and enzyme identity. The pH wash solutions also affects protease performance (Pig. 8). 

Fig. 8. Protease washing performance in a U.S. liquid detergent. Grass soiling in a 10 min wash at 30°C with one enzyme dosage, (a) pH profile of commercial proteases A and B. (b) Effect of increasing ionic strength, adjusted with Na2S04, of commercial protease B at (—°—) pH 8 and (- pH 11.

Bacterial a-amylases used in laundry detergents are fully compatible with detergent proteases, ie, the two enzymes work together in the wash process. During storage in both powder and Hquid detergents, the amylases are very stable in the presence of proteases. 

---