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Concept: Pancreatic lipase

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Plumbagin is a naphthoquinone found in the roots of Plumbago zeylanica. Here, we report an investigation to evaluate its antiobesity activity. The preliminary binding affinity of plumbagin to human pancreatic lipase (PL) was determined using molecular docking simulation. The in vitro PL inhibitory potential and the kinetics of inhibition were studied to validate and confirm the results obtained from molecular docking. The IC50 for PL was found to be 82.08 ± 9.47 μM, and the kinetics of inhibition was found to be of the mixed type. Further, the in vivo evaluation revealed that rats treated with plumbagin 1 mg/kg showed significant decrease in serum triglycerides (TG) and area under the curve of serum TG when compared with vehicle-treated rats. It was also seen that plumbagin possessed significant antiadipogenic effect as demonstrated by reduced oil red O staining and decreased TG contents. Thus, we conclude that plumbagin is a promising molecule to combat obesity and further optimization of plumbagin to yield plumbagin analogues will result in its improved activity profile.

Concepts: Pancreas, Atom, In vivo, Computational chemistry, Enzyme inhibitor, In vitro, Lipase, Pancreatic lipase

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The interfacial activity of pancreatic lipases (PL) depends on the presence of colipase and bile salt. The activity of PL is inhibited by micellar concentrations of bile salt which can be restored by the addition of colipase. Though the formation of 1:1:1 tertiary complex by lipase-colipase-bile salt micelle is well accepted, the residue-level interactions between lipase-colipase and bile salt are yet to be clearly understood. Molecular dynamic simulations of lipase-colipase complex, lipase and colipase were performed in the presence of a model bile salt, sodium taurocholate (NaTC), at its near-CMC and supra-micellar concentrations. From the interactions obtained from the molecular dynamic simulations, the ternary complex was modelled and compared with earlier reports. The analysis suggested that a micelle of NaTC consisting of nine monomers was formed at the concave groove between lipase and colipase chain and it mainly interacted with the fourth finger of colipase. This complex was mainly stabilized by van der Waals interactions. Interestingly, the C-terminal domain of lipase which holds the colipase did not show any significant role in formation or stabilization of NaTC micelle.

Concepts: Chemistry, Van der Waals force, Emergence, Bile, Bile acid, Taurocholic acid, Cholic acid, Pancreatic lipase

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Two new dihydrophenanthrenes, dendroinfundin A (1) and dendroinfundin B (2) were isolated from the whole plant of Dendrobium infundibulum, together with 7 known compounds (3-9). The structures of the new compounds (1 and 2) were established on the basis of their spectroscopic data. All of the isolates were evaluated for their α-glucosidase and pancreatic lipase inhibitory activities. Batatasin III (5) and dendrosinen B (9) showed strong α-glucosidase inhibitory activity. Dendrosinen B (9) also exhibited appreciable pancreatic lipase inhibitory effect.

Concepts: Activity, Lipase, Pancreatic lipase

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Pancreatic lipase inhibitors, such as tetrahydrolipstatin (orlistat), are used in anti-obesity treatments. Orlistat is the only anti-obesity drug approved by the European Medicines Agency (EMA). The drug is synthesized by saturation of lipstatin, a β-lactone compound, isolated from Streptomyces toxytricini and S. virginiae. To identify producers of novel pancreatic lipase inhibitors or microbial strains with improved lipstatin production and higher chemical purity remains still a priority. In this study, a high-throughput screening method to identify Streptomyces strains producing potent pancreatic lipase inhibitors was established. The assay was optimized and validated using S. toxytricini NRRL 15443 and its mutants. Strains grew in 24-well titer plates. Lipstatin levels were assessed directly in culture medium at the end of cultivation by monitoring lipolytic activity in the presence of a chromogenic substrate, 1,2-Di-O-lauryl-rac-glycero-3-glutaric acid 6-methylresorufin ester (DGGR). The lipase activity decreased in response to lipstatin production, and this was demonstrated by accumulation of red-purple methylresorufin, a product of DGGR digestion. The sensitivity of the assay was achieved by adding a lipase of high lipolytic activity and sensitivity to lipstatin to the reaction mixture. In the assay, the fungal lipase from Mucor javanicus was used as an alternative to the human pancreatic lipase. Many fungal lipases preserve high lipolytic activity in extreme conditions and are not colipase dependent. The assay proved to be reliable in differentiation of strains with high and low lipstatin productivity.

Concepts: Bacteria, Enzyme, Pancreas, Lipase, Pancreatic lipase, Orlistat, Streptomyces toxytricini, Lipstatin

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A polyphenol-rich extract (PRE) from the edible seaweed, Ascophyllum nodosum, inhibited pancreatic lipase activity in an oil-based turbidimetric assay with an IC50 of 200 μg gallic acid equivalents (GAE) perassay) [∼230 μg DW] whereas the known inhibitor, Orlistat, gave an IC50 at 0.4 μg per assay. A phlorotannin-enriched fraction (TRF) purified from the PRE was more potent with an IC50 = 60 μg GAE per assay (∼65 μg DW). When the assay was started by the addition of lipase, both Orlistat and TRF were much less effective which suggests that pre-incubation of enzyme and inhibitor improved inhibition. Based on phenol content, water extracts from Ascophyllum were more potent lipase inhibitors than PRE (IC50 ∼ 150 μg GAE per assay). However, this was equivalent to ∼580 μg DW and these extracts contained polysaccharides (e.g. alginate content = 110 μg mL-1) which may also contribute to inhibition. Indeed, a polysaccharide-enriched fraction obtained by ethanol precipitation gave an IC50 of 1000 μg DW which was equivalent to 130 μg GAE and 420 μg alginate per assay. Therefore a >3 fold increase in alginate content did not markedly improve inhibition. Re-precipitation increased alginate content and reduced polyphenol content but lipase inhibition was markedly reduced (i.e. IC50 at ∼1100 μg DW per assay, 700 μg alginate and 25 μg GAE). Purifying the polysaccharide fraction by ion exchange removed all phenolics but the IC50 increased to >2500 μg DW, equivalent to >1970 μg alginate per assay. In conclusion, polysaccharides and phlorotannins may inhibit lipase in an additive fashion, with phlorotannins apparently more effective in vitro. However, interactions between these components may be important when food products containing this edible seaweed are consumed.

Concepts: Cell wall, Enzyme inhibitor, Inhibitor, Xanthine oxidase inhibitor, Lipase, Gallic acid, Pancreatic lipase, Ascophyllum nodosum

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Roots of Boesenbergia rotunda (L.) Mansf. are prominent ingredients in the cuisine of several Asian countries, including Thailand, Malaysia, Indonesia, India, and China. An extract prepared from the roots of this plant showed strong inhibitory activity against enzymes α-glucosidase and pancreatic lipase and was subjected to chromatographic separation to identify the active components. Three new biflavonoids of the flavanone-chalcone type (9, 12, and 13) were isolated, along with 12 known compounds. Among the 15 isolates, the three new compounds showed stronger inhibitory activity against α-glucosidase than the drug acarbose but displayed lower pancreatic lipase inhibitory effect than the drug orlistat. The results indicated the potential of B. rotunda roots as a functional food for controlling after-meal blood glucose levels.

Concepts: Protein, Enzyme, Pancreas, Blood sugar, Chromatography, Lipase, Enzymes, Pancreatic lipase

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In this work, poly(carboxybetaine methacrylate) as an extremely hydrophilic polymer was modified on superparamagnetic Fe3O4 nanoparticles (pCB-Fe3O4), which were employed to immobilize porcine pancreatic lipase. The properties of immobilized lipase were investigated in comparison with the free enzyme counterpart. Enzymatic stability, reusability, and activity of the immobilized lipase were found significantly superior to that of the free lipase. In particular, at an elevated temperature of 60°C, the immobilized lipase retained 50% of its initial activity after 150min, while the free enzyme displayed only one-third activity of the immobilized enzyme. Besides, the immobilized lipase retained >60% of its initial activity after 7 cycles. Furthermore, the value of Kcat/Km indicated the catalytic efficiency of the immobilized lipase was increased by 50% compared to that of the free one. The pCB-Fe3O4 particles displayed non-cytotoxicity, while the naked Fe3O4 particles caused only 50% viability of NIH 3T3 cells. These results showed that pCB-Fe3O4 composite particles had higher efficiency and improved stability for lipase immobilization, which are more promising for industrial scale up of biocatalytic systems with excellent biocompatibility.

Concepts: Enzyme, Pancreas, Magnetism, Lipase, Enzymes, Immobilized enzyme, 3T3 cells, Pancreatic lipase

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The present study investigates the relationship between the high-order structure and hypolipidemic activity of four well-defined sulfated fucans from sea cucumber. The chain conformation, determined by a combination of AFM and SEC-MALLS-RI, indicate that fucosylated chondroitin sulfate (fCS) from Pearsonothuria graeffei (fCS-Pg) and Isostichopus badionotus (fCS-Ib), and fucoidan from P.graeffei (fuc-Pg) were assigned as a random coil conformation with polysaccharide chain outstretched, while I. badionotus (fuc-Ib) was assigned as a spherical conformation and exhibited high viscosity. Fuc-Pg and fuc-Ib with higher molecular weights had a greater impact in inhibiting pancreatic lipase activity in vitro. However, fCS-Pg, fCS-Ib and fuc-Pg with random linear conformation exhibited excellent hypolipidemic activity in Sprague-Dawley rats (SD rats) fed on high-fat diet (HFD), whereas fuc-Ib showed only a modest effect. Our results indicate that structural characteristics, including side branch and sulfation pattern can affect the chain conformation of polysaccharides, which determine their physicochemical properties and hypolipidemic activity.

Concepts: Effect, Polymer, Affect, Physical chemistry, Lipase, Pancreatic lipase, Polysaccharides, Xylose

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A series of novel diaryl substituted pyrazolyl 2,4-thiazolidinediones were synthesized via reaction of appropriate pyrazolecarboxaldehydes with 2,4-thiazolidinedione (TZD) and nitrobenzyl substituted 2,4-thiazolidinedione. The resulting compounds were screened in vitro for pancreatic lipase (PL) inhibitory activity. Two assay protocols were performed viz., methods A and B using p-nitrophenyl butyrate and tributyrin as substrates, respectively. Compound 11e exhibited potent PL inhibitory activity (IC50=4.81µM and Xi50=10.01, respectively in method A and B), comparable to that of the standard drug, orlistat (IC50=0.99µM and Xi50=3.72). Presence of nitrobenzyl group at N-3 position of TZD and nature of substituent at para position of phenyl ring at C-3 position of pyrazole ring notably affected the PL inhibitory activity of the tested compounds. Enzyme inhibition kinetics of 11e revealed its reversible competitive inhibition, similar to that of orlistat. Molecular docking studies validated the rationale of pharmacophoric design and are in accordance to the in vitro results. Compound 11e exhibited a potential MolDock score of -153.349kcal/mol. Further, the diaryl pyrazolyl wing exhibited hydrophobic interactions with the amino acids of the hydrophobic lid domain. Moreover, the carbonyl group at 2(nd) position of the TZD ring existed adjacent to Ser 152 (≈3Å) similar to that of orlistat. A 10ns molecular dynamics simulation of 11e-PL complex revealed a stable binding conformation of 11e in the active site of PL (Maximum RMSD≈3Å). The present study identified novel thiazolidinedione based leads with promising PL inhibitory activity. Further development of the leads might result in potent PL inhibitors.

Concepts: Enzyme, Molecular dynamics, Computational chemistry, Enzyme inhibitor, Thiazolidinedione, Docking, Molecular mechanics, Pancreatic lipase

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As per the recent statistical reports of World Health Organisation (WHO), 13% of total global population is obese. Orlistat remains to be the only drug approved for the long term treatment of obesity. Recent findings highlighted severe adverse effects of orlistat that included hepatotoxicity, gall stones, kidney stones and acute pancreatitis. Therefore, search for new drug is required. The investigations based on endophytic natural products would prove pivotal in the global fight against this health issue.

Concepts: Fungus, Obesity, Adverse drug reaction, Pancreatic cancer, World population, Endophyte, Pancreatic lipase, Orlistat