Measurement Of Lipase Activity Research Articles

Optimization of Classical Lipase Activity Assays for Fish Digestive Tract Samples

Fish possess lipases from embryonic development to adulthood. Lipase activity methods vary and significantly differ in terms of the concentration of the substrate used, bile salt, Ca2+, temperature, pH, and type of lipase units, which limits comparative studies. The three most-used substrates are p-nitrophenyl (p-NP), β-naphthyl (β-N) derivates, and emulsified natural oils. These were selected to be redesigned in this study to measure lipase activity under temperature, pH, ion, and bile salt conditions closer to fish physiology, using the appropriate molar absorption coefficient to calculate the lipase units. Cynoscion parvipinnis (CP), Seriola rivoliana (SR), Centropomus viridis (CV), Elop affinis (EA), and Canthidermis maculate (CM) pyloric caeca-intestine extracts were studied. Sodium taurocholate showed the highest activity for intestinal lipases, and the fatty acid length in the substrates changed the lipase hydrolysis rate. The highest lipase activity was obtained with p-NP butyrate and p-NP caprylate in four fish species. Lipase activity was highly activated with Ca2+ (4–7 mM). The β-N absorption spectrum indicates a plateau between 534 and 554 nm for fish lipases. Salmon oil was identified as the most digestible lipid in the four fish species using the in vitro digestibility assay. The lipase zymogram showed an apparent size of 46.3 kDa for CP, 40.2 kDa for SR, 46.2 kDa for CM, 106.6 kDa for EA, and 58.3, 84.6, and 162.1 kDa for CV.

Retrospective analysis of association between hepatopathy and serum DGGR lipase activity in dogs: a pilot study.

1,2-o-dilauryl-rac-glycero-3-glutaric acid-(6′-methylresorufin) ester (DGGR) lipase assays are used to measure lipase activity in the diagnosis of pancreatitis. The effect of hepatic lipases released from damaged hepatocytes on serum DGGR lipase activity has not been reported, to our knowledge. We identified dogs with histologically confirmed liver lesions and concurrent unremarkable pancreatic histology, and dogs with no histologic evidence of hepatic or pancreatic disease. Dogs with relevant comorbidities were excluded. The hepatopathy group (n = 7) included 4 dogs with inflammatory hepatopathies, 2 with hepatic neoplasia, and 1 with unspecified (non-inflammatory, non-neoplastic) hepatopathy. The control group (n = 5) included one dog each with enteritis, subcutaneous hemangiosarcoma, hydrocephalus, myelomalacia, and tetanus. A Mann–Whitney U test compared selected biochemical parameters including serum DGGR lipase, alkaline phosphatase, alanine aminotransferase, and amylase activities, with statistical significance defined as p ≤ 0.05. Data are presented as median and range. Serum DGGR lipase activity (RI: <44 IU/L) was not different between the hepatopathy (52 IU/L; range: 27–85 IU/L) and control (37 IU/L, 25–105 IU/L; p = 0.947) groups. Serum amylase activity (RI: 256–1,610 IU/L) was significantly higher in the hepatopathy group (830 IU/L; 711–1,210 IU/L) than the control group (541 IU/L, 336–695 IU/L; p = 0.028). No association or correlation between serum DGGR lipase activity and hepatic lesions (based on histologic or biochemical findings) was identified, suggesting that clinically relevant changes in serum DGGR lipase activity may not be expected secondary to hepatopathy alone.

The Activation Energies and Optimum Temperatures of Olive Oil Hydrolysis By Lipase Porcine Pancreas

AbstractLipase activity is a perfect indicator for the monitoring of processes of bioremediation of degraded soils. Lipase is also used in the processes of oil hydrolysis in wastewater treatment. To be able to predict and model processes with used lipase in environmental operations, knowledge of the kinetic parameters of the process are required. The paper presents the determined values of activation energies and optimum temperatures for porcine pancreas lipase. The parameters were estimated based on the literature of the activity curvesvs. temperature for hydrolysis of olive oil by lipase. It was noticed that concentration of gum arabic added as an emulsifier during lipase activity measurements influences on the obtained values of determined parameters. A mathematical model describing the effect of temperature on porcine pancreas lipase activity was used. Based on the comparison analysis, the optimum temperatureToptwere obtained in the range from 313.30 ±0.56 to 319.62 ±0.96 K, activation energiesEawere from 51 ±10 to 82.6 ±9.9 kJ/mol, and values of deactivation energiesEdwere in the range from 122.7 ±4.0 to 150.9 ±5.8 kJ/mol.

A simple, rapid, and sensitive fluorescence-based method to assess triacylglycerol hydrolase activity

Lipases constitute an important class of water-soluble enzymes that catalyze the hydrolysis of hydrophobic triacylglycerol (TAG). Their enzymatic activity is typically measured using multistep procedures involving isolation and quantification of the hydrolyzed products. We report here a new fluorescence method to measure lipase activity in real time that does not require the separation of substrates from products. We developed this method using adipose triglyceride lipase (ATGL) and lipoprotein lipase (LpL) as model lipases. We first incubated a source of ATGL or LpL with substrate vesicles containing nitrobenzoxadiazole (NBD)-labeled TAG, then measured increases in NBD fluorescence, and calculated enzyme activities. Incorporation of NBD-TAG into phosphatidylcholine (PC) vesicles resulted in some hydrolysis; however, incorporation of phosphatidylinositol into these NBD-TAG/PC vesicles and increasing the ratio of NBD-TAG to PC greatly enhanced substrate hydrolysis. This assay was also useful in measuring the activity of pancreatic lipase and hormone-sensitive lipase. Next, we tested several small-molecule lipase inhibitors and found that orlistat inhibits all lipases, indicating that it is a pan-lipase inhibitor. In short, we describe a simple, rapid, fluorescence-based triacylglycerol hydrolysis assay to assess four major TAG hydrolases: intracellular ATGL and hormone-sensitive lipase, LpL localized at the extracellular endothelium, and pancreatic lipase present in the intestinal lumen. The major advantages of this method are its speed, simplicity, and elimination of product isolation. This assay is potentially applicable to a wide range of lipases, is amenable to high-throughput screening to discover novel modulators of triacylglycerol hydrolases, and can be used for diagnostic purposes.

Label-Free Fluorescence Quantification of Hydrolytic Enzyme Activity on Native Substrates Reveals How Lipase Function Depends on Membrane Curvature.

Lipases are important hydrolytic enzymes used in a spectrum of technological applications, such as the pharmaceutical and detergent industries. Because of their versatile nature and ability to accept a broad range of substrates, they have been extensively used for biotechnological and industrial applications. Current assays to measure lipase activity primarily rely on low-sensitivity measurements of pH variations or visible changes of material properties, like hydration, and often require high amounts of proteins. Fluorescent readouts, on the other hand, offer high contrast and even single-molecule sensitivity, albeit they are reliant on fluorogenic substrates that structurally resemble the native ones. Here we present a method that combines the highly sensitive readout of fluorescent techniques while reporting enzymatic lipase function on native substrates. The method relies on embedding the environmentally sensitive fluorescent dye pHrodo and native substrates into the bilayer of liposomes. The charged products of the enzymatic hydrolysis alter the local membrane environment and thus the fluorescence intensity of pHrodo. The fluorescence can be accurately quantified and directly assigned to product formation and thus enzymatic activity. We illustrated the capacity of the assay to report the function of diverse lipases and phospholipases both in a microplate setup and at the single-particle level on individual nanoscale liposomes using total internal reflection fluorescence (TIRF). The parallelized sensitive readout of microscopy combined with the inherent polydispersity in sizes of liposomes allowed us to screen the effect of membrane curvature on lipase function and identify how mutations in the lid region control the membrane curvature-dependent activity. We anticipate this methodology to be applicable for sensitive activity readouts for a spectrum of enzymes where the product of the enzymatic reaction is charged.

Analysis of lipase activity in duodenal juice. Comparison of an automated spectrophotometric assay to a fluorometric microplate assay, and factors affecting sample stability

Background: Direct pancreas function testing (DPFT) has been regarded as gold standard for assessment of exocrine pancreas function. One of the outcomes from DPFT is pancreatic lipase activity in duodenal juice, but no standard assay for measuring pancreas lipase activity in duodenal juice exists.Aims: To optimize and evaluate an autoanalyzer assay for measuring lipase activity in duodenal juice.Methods: We used samples of duodenal juice from our biobank, collected through a short endoscopic secretin test in patients with suspected exocrine pancreas insufficiency. Samples were analyzed on a Cobas autoanalyzer (Roche Diagnostics), using a colorimetric, kinetic enzyme activity assay. We compared stability of samples diluted in saline to samples diluted in 3-(N-morpholino) propane sulfonic acid (MOPS) buffer added bovine serum albumin (BSA). Results from the Cobas assay were compared to Confluolip method, a fluorometric, kinetic enzyme assay, modified to fit into a microplate setting.Results: We tested the stability of 54 samples from 21 patients. Diluting samples with MOPS buffer added BSA gave stable results, and was superior to diluting samples in saline. We compared the two assays in 50 samples from 20 patients and found a good correlation between the two assays (r = 0.91, p < .001). There was a significant proportional bias between the two assays, but no significant systematic bias.Conclusion: Pancreatic lipase activity in duodenal juice samples diluted in MOPS buffer added BSA is stable for one hour at room temperature. Quantification of lipase activity in duodenal juice using a standard automated activity assay has comparable accuracy to a manual fluorometric method.

Lipase and Phospholipase Activity Methods for Marine Organisms.

Lipases are very important enzymes having a role in fat digestion and lipid metabolism in marine animals, plants, and microorganisms. The methods for measuring lipase and phospholipase activity have been applied in several studies; however, considering that lipases are water-soluble molecules and their substrates are generally water-insoluble molecules, several steps are requiredfor measuring their digestion products. After a general review of the main type of methods used in marine lipase studies, and experimental procedures, a proposal of new or improved methods is described in order to facilitate the lipase activity measurements in marine organisms.

An improved method to measure lipase activity in aqueous media

An improved method based on the p-nitrophenyl long chain esters method is proposed for measuring lipase hydrolytic activity in aqueous media. Using ethylene glycol as co-solvent for hydrophobic p-nitrophenyl substrates in aqueous buffer, lipase activity is measured by following the release of p-nitrophenol.This fast and easy to handle method improves the solubility of both substrate and product, and also the stability of the substrate. It avoids the use of solvents such as ethanol or propanol, permits the comparison of all the p-nitrophenol acyl ester substrates and allows the influence of ions like Ca+2 to be studied, while avoiding turbidity in the reaction medium.

Steeping of Whole Dried Maize Seeds in Buffers Altered Fatty Acid Ionization State, Composition, and Lipase Activity

Steeping is a simple model of studying the activation and modulation of the physiological pathways involved in seed germination. In this study, steeping of grains of the ‘obatanpa’ maize variety in buffers at different pH was monitored through the measurements of lipase activity, oil yield, fatty acid component and unsaturation, and germination capacity. Lipase activity of grains steeped for four days decreased in the order: pH 3 > pH 5 > pH 7 > pH 9 > pH 11. Decreasing lipase activity was corroborated with decreasing free fatty acid components, protein concentrations and oil yields. The unsaturation components of the oil fractions only marginally increased with increasing steeping media pH. Three major components were detected by TLC in all oil fractions. The unique components were confirmed by their uniform UV-absorption spectra converging at an isosbestic point of 290 nm. Germination capacity was much reduced for seeds steeped in buffered media for 24 hours compared with seeds steeped in portable water though the pattern of germination, which was monitored for five days, did not change. This study has demonstrated the use of pH changes of steeping medium to modulate physicochemical properties and germination of seeds. The physicochemical changes were observed after seeds have been submerged under steeping buffer for four days. Practical application: With proliferation of specialty maize hybrids, the study provides an insight into the development of experimental protocols for the selection of types of maize grain for preparation of foods and beverages in terms of general characterization and lipolytic activity, which have implications for flavor, taste and odor of the final products. The imminence of this in some traditional ways of preparing malted and fermented maize foods and beverages, which go through days of steeping, cannot be overemphasized. This study therefore provides another dimension to the manipulation of the steeping stage to develop varieties of maize-based product.

A Sensitive Microplate Assay for Lipase Activity Measurement Using Olive Oil Emulsion Substrate: Modification of the Copper Soap Colorimetric Method.

The present work involves a sensitive high-throughput microtiter plate based colorimetric assay for estimating lipase activity using cupric acetate pyridine reagent (CAPR). In the first approach, three factors two levels factorial design methodology was used to evaluate the interactive effect of different parameters on the sensitivity of the assay method. The optimization study revealed that the optimum CAPR concentration was 7.5% w/v, the optimum solvent was heptane and the optimum CAPR pH was 6. In the second approach, the optimized colorimetric microplate assay was used to measure lipase activity based on enzymatic hydrolysis of olive oil emulsion substrate at 37°C and 150 rpm. The emulsion substrates were formulated by using olive oil, triton X-100 (10% v/v in pH 8) and sodium phosphate buffer of pH 8 in ratio of 1:1:1 in the case of Candida sp. lipase. While in the case of immobilized lipozyme RMIM, The emulsion substrates were formulated by using olive oil, triton X-100 (1% v/v in pH 8) and sodium phosphate buffer of pH 8 in ratio of 2:1:1. Absorbance was measured at 655 nm. The stability of this assay (in terms of colored heptane phase absorbance readings) retained more than 92.5% after 24 h at 4°C compared to the absorbance readings measured at zero time. In comparison with other lipase assay methods, beside the developed sensitivity, the reproducibility and the lower limit of detection (LOD) of the proposed method, it permits analyzing of 96 samples at one time in a 96-well microplate. Furthermore, it consumes small quantities of chemicals and unit operations.

Radiation-induced injury of the exocrine pancreas after chemoradiotherapy for gastric cancer

Background and purposeThe pancreas is located almost entirely within the treatment area for radiotherapy of gastric cancer. The aim of this study was to analyze radiation-induced injury of the exocrine pancreas. Material and methodsThe study included 127 gastric cancer patients, who underwent preoperative or postoperative chemoradiotherapy. A total dose of 45Gy was given in 25 fractions. Concurrent chemotherapy was 5-fluorouracil-based. Lipase and α-amylase were assayed before, during and after treatment. ResultsLipase and α-amylase deficiencies were found in 48.2% and 19.7% of patients, respectively. In the univariant analysis, age and pretreatment α-amylase and lipase activities influenced on risk of injury of the exocrine pancreas (p<0.05). Younger patients (<65years) had a lower risk of hypoamylasemia than older patients. The probability of insufficiency was lower than 0.2 for patients with pretreatment α-amylase and lipase activities above 50U/L and 55U/L, respectively. The multivariate analyses of the time to hypolipasemia showed that only pretreatment lipase activity was significant. ConclusionsGastric cancer patients have an increased risk of exocrine pancreatic insufficiency after chemoradiotherapy. Thus, the pancreas should be regarded as an OAR. Measuring lipase activity should be the standard for assessing radiation-induced pancreatic injury.

Yarrowia lipolytica Lipase 2 Is Stable and Highly Active in Test Meals and Increases Fat Absorption in an Animal Model of Pancreatic Exocrine Insufficiency

Pancreatic exocrine insufficiency (PEI) reduces pancreatic secretion of digestive enzymes, including lipases. Oral pancreatic enzyme replacement therapy (PERT) with pancreatin produces unsatisfactory results. The lipase 2 produced by the yeast Yarrowia lipolytica (YLLIP2; GenBank: AJ012632) might be used in PERT. We investigated its ability to digest triglycerides in a test meal and its efficacy in reducing fecal fat in an animal model of PEI. YLLIP2 was produced by genetically engineered Y lipolytica and purified from culture media. YLLIP2 or other gastric (LIPF) and pancreatic (PNLIPD) lipases were added to a meal paste containing dietary triglycerides, at a range of pH values (pH 2-7), with and without pepsin or human bile and incubated at 37°C. We collected samples at various time points and measured lipase activities and stabilities. To create an animal model of PEI, steatorrhea was induced by embolization of the exocrine pancreas gland and pancreatic duct ligation in minipigs. The animals were given YLLIP2 (1, 4, 8, 40, or 80 mg/d) or pancreatin (100,000 US Pharmacopeia lipase units/d, controls) for 9 days. We then collected stool samples, measured fat levels, and calculated coefficient of fat absorption (CFA) values. YLLIP2 was highly stable and poorly degraded by pepsin, and had the highest activity of all lipases tested on meal triglyceride at pH 4-7 (pH 6 with bile: 94 ± 34 U/mg; pH 4 without bile: 43 ± 13 U/mg). Only gastric lipase was active and stable at pH 3, whereas YLLIP2 was sensitive to pepsin hydrolysis after pH inactivation. From invitro test meal experiments, the lipase activity of YLLIP2 (10 mg) was estimated to be equivalent to that of pancreatin (1200 mg; 100,000 US Pharmacopeia units) at pH 6. In PEI minipigs, CFA values increased from 60.1% ± 9.3% before surgery to 90.5% ± 3.2% after administration of 1200 mg pancreatin (P < .05); CFA values increased to a range of 84.6% ± 3.0% to 90.0% ± 3.8% after administration of 4-80 mg YLLIP2 (P < .05). The yeast lipase YLLIP2 is stable and has high levels of activity against test meal triglycerides in a large pH range, with and without bile. Oral administration of milligram amounts of YLLIP2 significantly increased CFA values, similar to that of 1.2 g pancreatin, in a minipig model of PEI.

Test system for direct measurement of lipase activities in milk samples

Test system for direct measurement of lipase activities in milk samples

A Sensitive and Robust Method for Direct Determination of Lipolytic Activity in Natural Milk Environment

Microbial lipases may be produced during milk storage and processing. This can lead to organoleptic changes in the corresponding dairy products. Thus, monitoring of lipase activity in milk is desirable. Turbidity of milk prevents a direct photometric measurement of lipase activity using chromophore- or fluorophore-based assays. Laborious pretreatments or alternative analytical methods normally have to be used. With the method for the determination of lipolytic activity (MeDeLi) proposed here, it is possible to measure lipase activity directly in the natural milk utilizing tailored fluorometric substrates. Only a defatting step is carried out initially for the MeDeLi. Then, the conversion of added lipase substrate is carried out in the unmodified milk without addition of any solutions or any enzyme extraction procedure which may influence the enzyme activity. Thereafter, the milk sample is treated with two solutions to remove the turbidity of milk by dissolution. A valid and sensitive fluorometric measurement is then possible. The applicability of the MeDeLi was demonstrated in comparison with tests published previously: The limit of detection for lipolytic activity measured by MeDeLi was the lowest, with 41 pkat/mL. Raw milk, milk products, and spoiled milk samples were also investigated with the MeDeLi.

A convenient test for lipase activity in aqueous-based solutions

We proposed a convenient and accurate method for the measurement of lipase activity in a uniform aqueous-based substrate solution. In this work, lipase from Candida rugosa was used as the model lipase to test its catalytic ability toward p-nitrophenyl palmitate (p-NPP), which was suspended in a mixture of p-NPP ethanol solution and buffer. An ultraviolet–visible spectrophotometer was used to efficiently measure the liberated p-nitrophenol without extraction or centrifugation. Several factors that affected lipase activity were investigated, such as the ratio of p-NPP ethanol solution to buffer, the concentrations of p-NPP and lipase, as well as the temperature, reaction time, pH and agitation rate. Additionally, enzyme catalytic parameters such as Km, Vm and “activation energy” were also assessed. We determined the optimal conditions for lipase in this homogeneous system and demonstrated lipase's catalytic performance in this condition followed Michealis–Menten kinetics.

Measurement of feline lipase activity using a dry-chemistry assay with a triolein substrate and comparison with pancreas-specific lipase (Spec fPL(TM)).

Pancreatic lipase immunoreactivity (Spec fPL) is currently considered to be the most accurate blood test for the diagnosis of feline pancreatitis. In this study, we measured lipase activity in cats using a newer catalytic lipase assay of dry-chemistry system (FDC-v-LIP) to determine the reference range and compared the results with those for Spec fPL. Based on the results of healthy cats, the reference range of FDC-v-LIP was determined to be less than 30 U/l. FDC-v-lip did not show a strong correlation with Spec fPL in cats with various diseases, which resulted in the low sensitivity and positive predictive value. However, the relatively high (>90%) specificity and negative predictive value indicated that FDC-v-LIP could be a useful patient-side screening test for the exclusion of feline pancreatitis.

Altering the activation mechanism in Thermomyces lanuginosus lipase.

It is shown by rational site-directed mutagenesis of the lid region in Thermomyces lanuginosus lipase that it is possible to generate lipase variants with attractive features, e.g., high lipase activity, fast activation at the lipid interface, ability to act on water-soluble substrates, and enhanced calcium independence. The rational design was based on the lid residue composition in Aspergillus niger ferulic acid esterase (FAEA). Five constructs included lipase variants containing the full FAEA lid, a FAEA-like lid, an intermediate lid of FAEA and TlL character, and the entire lid region from Aspergillus terreus lipase (AtL). To investigate an altered activation mechanism for each variant compared to that of TlL, a combination of activity- and spectroscopic-based measurements were applied. The engineered variant with a lid from AtL displayed interfacial activation comparable to that of TlL, whereas variants with FAEA lid character showed interfacial activation independence with pronounced activity toward pNP-acetate and pNP-butyrate below the critical micelle concentration. For variants with lipase and esterase character, lipase activity measurements further indicated a faster activation at the lipid interface. Relative to their activity toward pNP-ester substrates in calcium-rich buffer, all lid variants retained between 15 and 100% activity in buffer containing 5 mM EDTA whereas TlL activity was reduced to less than 2%, demonstrating the lid's central role in governing calcium dependency. For FAEA-like lid variants, accessible hydrophobic surface area measurements showed an approximate 10-fold increase in the level of binding of extrinsic fluorophores to the protein surface relative to that of TlL accompanied by a blue shift in emission indicative of an open lid in aqueous solution. Together, these studies report on the successful alteration of the activation mechanism in TlL by rational design creating novel lipases with new, intriguing functionalities.

Development of a high-throughput assay for measuring lipase activity using natural triacylglycerols coated on microtiter plates

We have designed a convenient, specific, sensitive and continuous lipase assay based on the use of natural triacylglycerols (TAGs) from the Aleurites fordii seed oil which contains α-eleostearic acid (9,11,13,cis,trans,trans-octadecatrienoic acid) and which was coated in the wells of microtiter plates. The coated TAG film cannot be desorbed by the various buffers used during the lipase assay. Upon lipase action, α-eleostearic acid is liberated and desorbed from the interface and then solubilized into the micellar phase. Consequently, the UV absorbance of the α-eleostearic acid is considerably enhanced due to the transformation from an adsorbed to a water soluble state. The lipase activity can be measured continuously by recording the variations with time of the UV absorption spectra. The rate of lipolysis was monitored by measuring the increase of OD at 272 nm, which was found to be linear with time and directly proportional to the amount of added lipase. This microtiter plate lipase assay, based on coated TAGs, presents various advantages as compared to the classical systems: (i) coated TAGs on the microtiter plates could be stored for a long-time at 4 °C, (ii) higher sensitivity in lipase detection, (iii) good reproducibility, and (iv) increase of signal to noise ratio due to high UV absorption after transfer of α-eleostearic acid from an adsorbed to a soluble state. Low concentrations, down to 1 pg mL(-1) of pure Thermomyces lanuginosus or human pancreatic lipase, could be detected under standard assay conditions. The detection sensitivity of this coated method is around 1000 times higher as compared to those obtained with the classical emulsified systems. This continuous high throughput lipase assay could be used to screen new lipases and/or lipase inhibitors present in various biological samples.

Intraduktal papillär muzinöse Neoplasie: Welche Befunde sprechen für Beobachtung?

On abdominal CT scans asymptomatic cystic lesions of the pancreas are accidentally detected in 1-2% of patients. Congenital cysts and pancreatic pseudocysts account for two thirds of these lesions. Pancreatic pseudocysts are a frequent complication of acute and chronic pancreatitis. Among resected cystic neoplasms serous cystic adenoma accounts for 30%, mucinous cystic neoplasms for 45% and intraductal papillary mucinous neoplasms for 25%. The diagnosis of a cystic pancreatic lesion is usually made by diagnostic imaging. Symptomatic lesions require definitive therapeutic treatment after appropriate diagnostic work-up. In the diagnosis of asymptomatic cystic lesions several factors are important, among them whether the cyst is connected to the pancreatic duct (as in IPMN and pseudocysts), the size of lesion (for treatment indications) and whether nodules form in the wall of the cyst (a sign of potential malignancy). EUS-guided fine needle aspiration of the cyst fluid adds to the discrimination between benign, premalignant and malignant cystic lesions. Measuring lipase activity, CEA, viscosity and mucin as well as cytology can help in differentiating cystic lesions. An algorithm is discussed for the differential diagnosis and for selection of the appropriate treatment for pancreatic cystic lesions, most of which never require surgery.

Kinetic properties of mouse pancreatic lipase-related protein-2 suggest the mouse may not model human fat digestion

Genetically engineered mice have been employed to understand the role of lipases in dietary fat digestion with the expectation that the results can be extrapolated to humans. However, little is known about the properties of mouse pancreatic triglyceride lipase (mPTL) and pancreatic lipase-related protein-2 (mPLRP2). In this study, both lipases were expressed in Pichia Pastoris GS115, purified to near homogeneity, and their properties were characterized. Mouse PTL displayed the kinetics typical of PTL from other species. Like mPTL, mPLRP2 exhibited strong activity against various triglycerides. In contrast to mPTL, mPLRP2 was not inhibited by increasing bile salt concentration. Colipase stimulated mPLRP2 activity 2- to 4-fold. Additionally, mPTL absolutely required colipase for absorption to a lipid interface, whereas mPLRP2 absorbed fully without colipase. mPLRP2 had full activity in the presence of BSA, whereas BSA completely inhibited mPTL unless colipase was present. All of these properties of mPLRP2 differ from the properties of human PLRP2 (hPLRP2). Furthermore, mPLRP2 appears capable of compensating for mPTL deficiency. These findings suggest that the molecular mechanisms of dietary fat digestion may be different in humans and mice. Thus, extrapolation of dietary fat digestion in mice to humans should be done with care.