Lipase carboxylester

Soluble substrates an available which are cleaved by carboxylester lipase in the absence of bile salts, such as the not quite specific /wra-nitrophenyUcetate... 

Relevant quality control should not be restricted to the usual triad of activities of pancreas lipase, a-amylase, and trypsin, but should be extended to the content of colipasc, the activities of the two other lipolytic enzymes present in pancreatine (phospholipase Aj and carboxylester lipase), and the dissolution characteristics of enteric-coated preparations as a function of time and pH (Fig. 16). The availability of such information will certainly contribute to a better tailoring of flic management of maldigestion in the individual patient and to a more appropriate correction of the obligate nonphysio logical route of delivery of these enzyme supplements. 

Tsujita T, Okuda H. The synthesis of fatty acid ethyl ester by carboxylester lipase. Eur J Biochem 1994 224 47-62. 

While it was previously thought that the lipase of pancreatic origin, the classical lipase, was responsible for the digestion of most dietary fat, it has become clear in recent years that lipolysis in the gastrointestinal tract is a result of the concerted action of four different types of lipases gastric lipase, classical pancreatic lipase, pancreatic carboxylester lipase (identical to cholesterol esterase), and phospholipase A2 (Table 2). 

Pancreatic carboxylester lipase, secreted by the pancreas as an active enzyme without proteolytic activation, displays broad substrate specificity and has therefore received many names in the literature carboxylesterase, bile salt-stimulated (or activated or dependent) lipase (due to its absolute requirement for bile salts to hydrolyze insoluble substrates), carboxylester lipase or hydrolase, cholesterol esterase, lysophospholipase, nonspecific lipase, and monoglyceride lipase. The IUPAC classification of the enzyme has been either EC.3.1.1.1 (carboxylester hydrolase) or EC.3.1.1.13 (cholesterolester hydrolase) (Table 2). 

Recent studies indicate [38,39] that human pancreas carboxylester lipase and bile salt-stimulated human milk lipase share the same peptide sequence, also displaying a high degree of similarity to acetylcholine esterase. 

Topical pancreatic lipase substrates like tributyrin and triolein emulsions are hydrolyzed by carboxylester lipase in the presence of bile salt but slowly—at a rate lower than 3% and 0.5%, respectively, of that observed with the lipase-colipase complex. On the other hand, the positional specificity is not restricted all three sn positions of triglycerides can be split by the carboxylester lipase. While long-chain phospholipids are resistant, short-chain phospholipids are readily attacked by carboxylester lipase [40]. The low substrate specificity of carboxylester lipase makes possible an essential role for this enzyme in the hydrolysis of triglycerides containing certain esterified polyunsaturated fatty acids, such as eicosapentaenoic, arachidonic, or linoleic acids [41], and which may be resistant to attack by pancreas lipase (see p. 190). 

The various esters of cholesterol, retinoyl esters, esters of vitamin D and E, probably depend solely on the activity of carboxylester lipase for their hydrolysis. They are, before their uptake by the enterocytes, transformed into the corresponding alcohols to form mixed micelles with bile salts. Carboxylester lipase catalyzes not only the cleavage of esters but also their formation. The most studied example is the reversible esterification of cholesterol, which is favored by low bile salt concentrations and pH [42]. Additionally, the laige specificity of carboxylester lipase probably functions as a first-line detoxification mechanism for a broad variety of orally ingested xenobiotics. 

Carboxylester lipase is present in higher amounts, expressed as protein mass, than other lipolytic enzymes in duodenal juice [43]. The enzyme is coded on a polymorphic locus on human chromosome 9 [44,45] and consists of a single chain with a molecular weight varying according to the species for example, 100 kDa for the human enzyme [46], 80 kDa for [47] the pig, and 65 kDa for the rat carboxylester lipase [48]. 

Soluble substrates are available which are cleaved by carboxylester lipase in the absence of bile salts, such as the not quite specific para-nitrophenylacetate [40] or the dilaurate ester of fluorescein [51,52] (Fig. 9). However, bile salts not only protect the enzyme, but interact specifically with the enzyme, affecting approximately 7- to 10-fold the activity on water-soluble substrates like para-nitrophenylacetate. When water-insoluble esters like triolein or cholesterol oleate are selected as the substrate, the enzyme is virtually inactive without bile salts, which gives them the character of essential activators [53]. In contrast to pancreatic lipase, carboxylester lipase needs no cofactor like colipase and is not activated by an oilAvater interface. Purified carboxylester lipase loses the majority of its activity by lyophylization but can be stored frozen at -20°C [46]. 

Figure 9 The pancreolauryl test allows an indirect assessing of exocrine pancreatic function. Orally administered fluorescein dilaurate is hydrolyzed by carboxylester lipase (identical to cholesterol esterase) liberating lauric acid and free, water-soluble fluorescein. The latter is readily absorbed in the small intestine, partly conjugated in the liver, and excreted in urine mainly as fluorescein glucuronide. By measuring the concentration of fluorescein in the urine over a period of, for instance, 10 hours, the total quantity of this dye is determined. (From Ref. 52.)...

It is clear that the interconnection between the activities of the different lipolytic enzymes, where the first enzyme modifies the physicochemical state of the lipid substrate in such a way that it becomes available to another enzyme, is not only of prime importance for lipid digestion, but also results in a broad synergism between gastric lipase, colipase, pancreatic lipase, phospholipase A2, calcium, carboxylester lipase, bile salts, and substrate intermediates [55,62-64],... 

Figure 11 Hydrolysis of an artificial triolein/phospholipid/cholesteiol oleate emulsion with either pancreatic phospholipase A2 (PLA2) and/or pancreatic carboxylester lipase (CEL) in the presence of 10 mM bile salt. Closed squares PLA2 + CEL open squares CEL alone. Hydrolysis is monitored using tritiated tracers. (From Ref. 61.)...

Pancreatin is described in the different pharmacopeia as a slightly brown, amorphous powder with a faint, characteristic odor. It is slowly and incompletely soluble in water it is not recommended to filter the solution as the lipolytic enzymes and part of the proteolytic activity may be retained on the filter. Most pharmacopeia stipulate the minimum units of amylase (12,000 FIP units/g), lipase (15,000 FIP units/g), and protease (1000 FIP units/g) activity, but give no specifications on the content of the other components such as phospholipase A2, carboxylester lipase, colipase, and individual proteolytic enzymes. It... 

T. Tsujita and H. Okuda. Effect of bile salts on the interfacial inactivation of pancreatic carboxylester lipase. J. Lipid Res. 31 831 (1990). 

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