K562 cells were divided into five groups and treated with 0 µM, 20 µM, 40 µM, 60 µM, and 80 µM of Orlistat (Sigma) and Rosuvastatin (Sigma) or with a mixture of Orlistat and Rosuvastatin for 24 h, 48 h, and 72 h. After IC50 analyzing, for downstream measurements, cells were co-cultured with Orlistat and Rosuvastatin over a period of 72 h. Compounds were dissolved in 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (DMSO, Merck) solution.
Orlistat
Manufactured by Merck Group
Sourced in United States, Germany, Spain, United Kingdom, China
Orlistat is a synthetic drug that functions as a lipase inhibitor. It is designed to inhibit the absorption of dietary fats by the human body.
Automatically generated - may contain errors
Lab products found in correlation
Acarbose, by Merck Group (35 mentions)
Folin ciocalteu reagent, by Merck Group (18 mentions)
α amylase, by Merck Group (17 mentions)
2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (17 mentions)
Porcine pancreatic lipase, by Merck Group (16 mentions)
Quercetin, by Merck Group (16 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (13 mentions)
Gallic acid, by Merck Group (13 mentions)
α glucosidase, by Merck Group (13 mentions)
Lipase, by Merck Group (11 mentions)
Linoleic acid, by Merck Group (11 mentions)
Ascorbic acid, by Merck Group (10 mentions)
Sodium carbonate, by Merck Group (10 mentions)
Methanol, by Merck Group (9 mentions)
P nitrophenyl α d glucopyranoside, by Merck Group (8 mentions)
Catechin, by Merck Group (7 mentions)
Caffeic acid, by Merck Group (7 mentions)
Bovine serum albumin (bsa), by Merck Group (7 mentions)
Pancreatic lipase, by Merck Group (6 mentions)
Trolox, by Merck Group (6 mentions)
131 protocols using orlistat
1
Cytotoxicity Evaluation of Orlistat and Rosuvastatin
2
Inhibition of FASN in Colorectal Cancer
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For the application of the FASN synthetase inhibitor Orlistat (O4139-25MG, Merck, Darmstadt, Germany) in vitro, the inhibitor was prepared by referencing the procedure of Browne et al. [17 (link)], and 200 µmol/L Orlistat was adopted to treat CRC cells for 48 h when necessary. Another FASN inhibitor, C75 (25–200 µmol/L) (HY-12364, MedChemExpress, Monmouth Junction, NJ, USA), was used to treat CRC cells for 48 h as per a report from Chen et al. [11 (link)]. Based on a previous study [18 (link)], 800 µmol/L of palmitoleic acid (HY-N1966, MedChemExpress) was applied for 72 h for in vitro experiments.
3
Inhibition of CRC Cell Growth
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For the application of the FA synthetase inhibitor orlistat (O4139‐25MG, Merck, Darmstadt, Germany) in vitro, it was prepared by referencing the procedure of Browne et al. [25 (link)], and 50 μmol/L orlistat was adopted to treat CRC cells for 48 h when necessary. As for in vivo experiments, it was prepared as described by Kridel et al. [26 (link)], and it was intraperitoneally injected into nude mice with a concentration of 240 mg/kg. The phosphoInositide‐3 kinase (PI3K) inhibitor LY294002 (20 μmol/L) (HY‐10108, MedChemExpress, Monmouth Junction, NJ, USA) was used to treat CRC cells for 24 h as per a report from Chen et al. [27 (link)]. Based on a previous study [28 (link)], 40 μmol/L mitogen‐activated protein kinase (MAPK) inhibitor (E)‐osmundacetone (HY‐N1966, MedChemExpres) was applied for 24 h for in vitro experiments.
4
Cytotoxic Effects of Orlistat on HEK293T/17 Cells
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Cytotoxicity of orlistat (Merck KGaA) was evaluated in HEK293T/17 cells at 36 h with pre-, post-, or pre- and post-combined treatments using a Trypan Blue viability testing protocol. Dilutions of cell suspensions were mixed 1:1 with 0.4% Trypan Blue prior to counting live cells using a hemocytometer under a microscope. Negative controls (only DMEM and vehicle DMSO control of each concentration) and positive controls (milli-Q water) were undertaken in parallel. In addition, cell morphology was evaluated under an inverted light microscope.
5
Preparation of Baicalein and Orlistat Solutions
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Baicalein (Supplemental Fig. 1 ; Merck KGaA, Darmstadt, Germany) was dissolved in 100% DMSO (Merck KGaA, Darmstadt, Germany) to give a stock solution concentration of 100 mM, which was stored at -30 ºC until used. Working solutions were prepared by diluting the stock solution in complete cell culture medium to the desired concentration. Orlistat (Merck KGaA, Darmstadt, Germany) was dissolved in 100% DMSO to give a stock concentration of 38.3 mM and was also stored at -30oC until used.
6
Antibody Reagents and Protocols
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Antibodies used in this study are shown in Supplementary Table 2 . The following reagents were purchased from the manufacturers as noted: DMXAA (14617, Cayman), anti-FLAG M2 Affinity Gel (A2220, Sigma), LysoTracker Deep Red (L12492 , Thermo Fisher Scientific), E64d (4321, Peptide Institute), pepstatin A (4397, Peptide Institute), orlistat (O4139, MERCK) and HT-DNA (D6898, Sigma).
7
Optimizing C. pyrenoidosa Protein Extraction
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C. pyrenoidosa powder (62.4% of total protein contents) was provided by Professor Zhang Daojing, East China University of Technology, Shanghai, China. BCA Protein Assay Kit and Triglyceride Assay Kit were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). Pepsin (300,000 U/g), papain (800,000 U/g), and trypsin (1:250 U/g) were obtained from Guangzhou Qiyun Biotech Co., Ltd., Guangzhou, China. Alcalase 2.4 L (P4860), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), pancreatic lipase, Orlistat, and simvastatin were obtained from Sigma, San Jose, CA, USA. Primary and secondary antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA). Gel filtration chromatography Sephadex G-25 was from Whatman Liluo Science & Technology Instrument Co., Guangzhou, China. Other reagents were of analytical grade and were commercially available.
8
Lipase and Pepsin Activity Assay
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Sodium hydroxide (NaOH), hydrochloric acid (HCl), trichloroacetic acid (TCA) and acetic acid were from Fisher Scientific (Dublin, Ireland). Kjeldahl tablets, sulfuric acid (>98%, H2SO4), boric acid, trifluoroacetic acid (TFA), Trizma® base, dimethyl sulfoxide (DMSO), 2-mercaptoethanol, methanol, protein markers (6.5–200 kDa), Sudan III, 2,4,6-trinitrobenzenesulfonic acid (TNBS), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), porcine pepsin (2500 U/mg), porcine pancreatic lipase (50 mU/mL), 4-methylumbelliferyl oleate (4-MUO) and Orlistat® were from Sigma-Aldrich (Dublin, Ireland). Hexane, MS-grade water and acetonitrile (ACN) were from Honeywell International Inc. (Dublin, Ireland). Comassie R, Laemmli buffer and Mini-Protean TGX 4–20% pre-cast polyacrylamide gels were from Bio-Rad Laboratories Inc. (Hercules, CA, USA). Sodium dodecyl sulfate (SDS) was from National Diagnostics (Atlanta, GA, USA). Corolase PP® (4.4 U/mg) was from AB Enzymes GmbH (Darmstadt, Germany). Cellulose acetate filters were from Millipore (Carrigtwohill, Ireland) and polytetrafluoroethylene syringe filters were from VWR (Dublin, Ireland).
9
Antioxidant Capacity Evaluation
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1,1-diphenyl-2 picrylhydrazyl (DPPH), 2,2′-Azinobis (3-ethyl benzthiazoline-6-sulphonic acid) liquid substrate (ABTS), trolox, ascorbic acid (AA), hydrogen peroxide (H2O2), Folin-Ciocalteu phenol reagent, nitroblue tetrazolium (NBT), superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), catechin, quercetin, orlistat were obtained from Sigma-Aldrich Chemicals (St Louis, MO, USA). Other chemicals of analytical grade were purchased from Merck Limited (Mumbai, India).
10
Orlistat Inhibits Triglyceride Hydrolysis
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Orlistat (Sigma-Aldrich Co.) was solubilized in 33% ethanol and 66% PEG 400 (Hampton Research, Aliso Viejo, CA, USA). The C57BL/6 mice fed a standard chow diet received Orlistat (250 mg kg−1 body weight) via injection into the intraperitoneal space26 (link), followed 90 min later by measurement of plasma TG-hydrolysis activities. Next, serum TG levels were measured and HPLC analyses of lipid profiles were performed.
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