Fig. 1. Effects of pH on the activity (U/mg of enzymes or total proteins) of: A, porcine pancreatic extracts; B, human pancreatic juice; C, rHPLRP2; D, bovine CEH; E, rDGL measured during Gelucire^® 44/14 lipolysis. (●) apparent activity measured by direct titration of the free fatty acids (n = 3); (○) specific activity measured by back-titrating the total free fatty acids at pH 9 (n = 2). Data are means ± SD.

Fig. 2. Effect of pH on the fatty acid ionization extents (expressed as percentages) measured during the lipolysis of Gelucire^® 44/14 by rHPLRP2 (▲), bovine CEH (□), and rDGL (●).

Fig. 3. Specific activities of rHPL, rHPLRP2, bovine CEH, and rDGL on Gelucire^® 44/14, C8-C18 acylglycerol fraction of Gelucire^® 44/14 and PEG-32 ester fraction of Gelucire^® 44/14. Specific activities were measured at 37 °C and pH 6.0 by back-titration of the free fatty acids in 150 mM NaCl, 4 mM NaTDC, 1.4 mM CaCl₂ and 1 mM Tris–HCl. Data are means ± SD (n = 2).

Fig. 4. Specific activities of rHPL, rHPLRP2, bovine CEH, and rDGL on (A) di- and triacylglycerol substrates and (B) monoacylglycerol substrates. Lipase activities were measured at 37 °C and pH 6.0 by back-titration of free fatty acids in 150 mM NaCl, 4 mM NaTDC, 1.4 mM CaCl₂, 1 mM Tris–HCl. Values are means ± SD (n = 2). Values of specific activities on TC8, DC8, and MC8 were those obtained in [25].

Fig. 5. Specific activities of rHPL, rHPLRP2, bovine CEH, and rDGL on (A) PEG-32 monoesters with various acyl chain lengths and (B) PEG-32 diesters with various acyl chain lengths. Specific activities were measured at 37 °C and pH 6.0 by back-titration of free fatty acids in 150 mM NaCl, 4 mM NaTDC, 1.4 mM CaCl₂, 1 mM Tris–HCl. Values are means ± SD (n = 2).

Fig. 6. Effects of PEG length on the specific activity of rHPL, rHPLRP2, bovine CEH, and rDGL on (A) PEG monocaprylates and (B) PEG dicaprylates. Lipase activities were measured at 37 °C and pH 6.0 by back-titration of free fatty acids in 150 mM NaCl, 4 mM NaTDC, 1.4 CaCl₂, 1 mM Tris–HCl. Values are means ± SD (n = 2). Values of specific activities on PEG-6 mono- and dicaprylate were based on [25].

Maximum specific activities of PPE, HPJ, rHPLRP2, bovine CEH, and rDGL on Labrasol^® and Gelucire^® 44/14

Specific activities, as measured by back-titration of free fatty acids, were expressed as unit per milligram of pure enzyme or per milligram of total proteins assayed in the powder in the case of porcine pancreatic extracts and human pancreatic juice. Specific activities on Labrasol^® are based on [25]. Values are means ± SD (n = 3). Values in brackets indicate the optimum pH at which the activity was measured.

Putative and relative contributions of classic pancreatic lipase (PL), pancreatic lipase-related protein 2 (PLRP2), carboxyl ester hydrolase (CEH) and gastric lipase (GL) to the in vivo lipolysis of Labrasol^® and Gelucire^® 44/14