All solvents and chemicals were purchased from Sigma-Aldrich (St. Louis, MO) and used as received. Phosphate-buffered saline without calcium and magnesium was from Thermo Scientific (Logan, UT). 10,000 MWCO dialysis membrane was from Spectrum Laboratories (Rancho Dominguez, CA). Posaconazole (Noxafil, Schering Corporation, Kenilworth, NJ) was purchased from Biosynth Carbosynth® (catalog no. FP27107). The G5 PAMAM dendrimer was from Dendritech (Midland, MI) and purified by dialysis (10,000 MWCO dialysis membrane) against H2O. The number of terminal amino groups on G5 dendrimer is experimentally 114 (Choi et al., 2012 (link); Mukherjee et al., 2015 (link)).
Carbosynth
Manufactured by Biosynth
Sourced in United Kingdom
Carbosynth is a laboratory equipment product that specializes in the synthesis and purification of carbohydrates and related compounds. It provides tools and technologies for researchers and scientists to efficiently produce and analyze various types of carbohydrates.
Automatically generated - may contain errors
Lab products found in correlation
Phosphate buffered saline without calcium and magnesium, by Thermo Fisher Scientific (1 mentions)
Naringenin, by Biosynth (1 mentions)
Rutin, by Biosynth (1 mentions)
Phloretin, by Biosynth (1 mentions)
Ellagic acid, by Merck Group (1 mentions)
Quercetin, by Merck Group (1 mentions)
Phloroglucinol, by Merck Group (1 mentions)
Apigenin, by Biosynth (1 mentions)
Naringin, by Biosynth (1 mentions)
Kinetex xb c18, by Phenomenex (1 mentions)
Melting point b 540, by Büchi (1 mentions)
Mercury plus 400, by Agilent Technologies (1 mentions)
Merck silica gel 60 f254, by Merck Group (1 mentions)
R 114, by Büchi (1 mentions)
Molnupiravir, by Biosynth (1 mentions)
Heparin, by Biosynth (1 mentions)
8 protocols using carbosynth
1
Posaconazole Loaded PAMAM Dendrimer
2
Phenolic Compound Characterization Protocol
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The phenolic standards, rutin (≥ 95% purity, CAS No. 153-18-4), epigallocatechin-3-O -gallate (EGCG) (≥ 95% purity, CAS No. 989-51-5), (+)-naringenin (≥ 98% purity, CAS No. 67604-48-2), naringin (≥ 95% purity, CAS No. 10236-47-2), apigenin (≥ 94% purity, CAS No. 520-36-5), daidzein (≥ 98% purity, CAS No. 486-66-8), phloretin (≥ 98% purity, CAS No. 60-82-2) were purchased from Biosynth® CarboSynth (Bratislava, Slovakia); whereas quercetin (≥ 95% purity, CAS No. 117-39-5), ellagic acid (≥ 96% purity, CAS No. 476-66-4) and phloroglucinol (≥ 99% purity, CAS No. 108-73-6) - were purchased from Sigma-Aldrich (Madrid, Spain). HPLC-grade methanol, water, and acetonitrile were supplied by Fisher Scientific Chemicals. The enzymes α-amylase (A1031), pepsin from porcine stomach mucosa (P7012), pancreatin from porcine pancreas (P7545), and bile salts (B863) were purchased from Sigma-Aldrich (Taufkirchen, Germany). The rabbit gastric extract (RGE15) with gastric lipase activity was purchased from Lipolytech (Marseille, France). The ethanol was purchased from Merck (Algés, Portugal). A dialysis membrane with a molecular pore size of 3 kDa was purchased from Spectra/Pro (Spectrum Lab, Breda, Netherlands).
3
Analytical Characterization of Organic Compounds
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The reagents were purchased from Merck Life Science S.r.l. and Biosynth Carbosynth®. Melting points were evaluated on a Buchi Melting Point B-540 instrument. They are uncorrected and were evaluated on chromatographically purified or recrystallized compounds. The reactions were controlled by thin layer chromatography (TLC), using Merck silica gel 60 F254 plates preloaded with fluorescent indicator and visualized with UV light (254 nm). Silica gel column (Kieselgel 60) was employed for preparative chromatographic purifications. Na2SO4 was used to dry all the solutions which were concentrated with a Buchi R-114 rotary evaporator at low pressure. Homogeneity of the products was evaluated by analytical reversed-phase HPLC (RP-HPLC) on a Shimadzu-10 ADsp HPLC system by means of a Phenomenex Kinetex XB-C18 column, 5 μm, 4.6 × 250 mm and applying the gradient of acetonitrile (CH3CN) in 0.1% aqueous trifluoroacetic acid (TFA), from 0% to 100% in 25 min, at a flow rate of 1 mL/min. Analytical RP-HPLC indicated a purity > 98%. 1H and 13C NMR spectra were recorded on Varian Mercury Plus 400 MHz instrument in dimethyl sulfoxide (DMSO‑d6). Chemical shifts are described in ppm, while peak patterns are reported using the abbreviations: m (multiplet), t (triplet), d (doublet), s (singlet). API 2000 Applied Biosystem mass spectrometer was used to determine mass spectra of the final products.
4
Anti-viral Efficacy of Kuwanon G
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Prior to the antiviral tests, the cytotoxicity of kuwanon G was evaluated as described in 4.4.1. The tested concentrations ranged from 0 to 20 µg/mL. The results were calculated after 3 independent experiments, and 4 replicates for each concentration were tested. The cells were then incubated for 24 h at 37 °C. On the day of infection, dilutions of kuwanon G (Biosynth® carbosynth, CAS No 75629-19-5) from 0.1 to 10 µg/mL were prepared in the culture medium, containing 2% of FBS. These concentrations have been chosen according to the kuwanon G cytotoxicity assay. Cascade dilutions of the HCoV 229E virus were prepared for each kuwanon G dilution in order to test the virus at a multiplicity of infections (MOIs) equal at 1, 0.1, and 0.01, and wells were incubated with the virus/kuwanon G mix. Each test was reproduced in 8 identical wells. The plates were incubated for 72 h at 33 °C. The analysis of the wells was performed by microscopic observation of the cytopathogenic effect of HCoV 229E in the presence and absence of kuwanon G and after staining of the cell monolayers with crystal violet, as previously described.
5
Evaluation of Anti-COVID-19 Compounds
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Samples of the target compounds, ChloViD2022, CoViTris2022, and Taroxaz-26, were obtained from our previous work (Purity of each of them: ≥ 95%) [23 (link)]. While the reference anti-COVID-19 drug molnupiravir (EIDD-2801, CAS Registry Number: 2349386-89-4) was purchased from Biosynth Carbosynth (Carbosynth Ltd., Berkshire, U.K.) (Product Code: AE176721, Purity: ≥ 98%). The ultrapure solvent dimethyl sulfoxide (DMSO, CAS Registry Number: 67-68-5) was purchased from a local distributor, El-Gomhouria Company For Drugs (El-Gomhouria Co. For Trading Drugs, Chemicals & Medical Supplies, Mansoura Branch, Egypt) (Purity: ≥ 99.9% “anhydrous”).
6
Enzymatic Deconstruction of Lignocellulosic Biomass
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Methyl ferulate, methyl p-coumarate, ferulic acid, and p-coumaric acid were obtained from Apin Chemicals Ltd. (Abingdon, United Kingdom), Carbosynth (Compton, United Kingdom), FlukaTM (Leicestershire, United Kingdom) and Sigma Aldrich (Darmstadt, Germany), respectively. Sugar beet pectin (SBP, Pectin Betapec RU301) was purchased from Herbstreith & Fox KG (Neuenbürg, Germany). Wheat arabinoxylan insoluble was obtained from (WAX-i, Megazyme, Bray, Ireland) and the corn fiber oligomers (CFoligo)-preparation was obtained as previously described (Appeldoorn et al., 2010 (link)). Corn stover (CS) was obtained from DSM (Delft, The Netherlands) and wheat straw (WS) were kindly provided by CNC Grondstoffen B.V. (Milsbeek, The Netherlands). Prior to incubation, both CS and WS were ball milled as previously described (van Erven et al., 2017 (link)), giving rise to deconstruction of cellulose crystallinity and making the substrates more susceptible to enzymatic pretreatment (Broxterman et al., 2018 (link)). The carbohydrate content and composition of all substrates used was performed as previously described (Englyst and Cummings, 1984 (link)). The analysis details are presented in the supporting information (Supplementary Table S1 ).
7
Synthesis of Heparin-Modified Hierarchical Zeolites
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The process of synthesis of hierarchical zeolites based on commercial types of zeolites such as ZSM-5, β, or Y, modified with heparin, was analogous to the synthesis of materials described in Section 2.1.2 and Section 2.1.3 , only during the addition of 0.84 g of TEOS, 0.05 g of heparin (Biosynth Carbosynth, Staad, Switzerland) was added instead of inulin or hyaluronic acid. The remaining course of synthesis of heparin-modified materials, was analogous to those described above.
8
Kinase Inhibition Screening Protocol
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All predicted inhibitors for testing (except compound 3 ) were purchased from Vitas-M Laboratory, with purity ≥ 90%. Compound 3 was purchased from Biosynth Carbosynth with purity of 95%. Binding assay experiments against Homo sapiens GSK-3β were performed using a specialist service from the MRC Protein Phosphorylation & Ubiquitylation Unit at the University of Dundee (http://www.kinase-screen.mrc.ac.uk/ (accessed on 1 April 2023)). Initial assays underwent single concentration screening at 50 µM concentrations to determine those compounds showing the best inhibitory potential. Inhibitory activities were calculated based on maximal activities measured in the absence of an inhibitor. For selected candidates, the IC50 values were determined, defined as the concentration of a compound that reduces the enzymatic activity by 50% with respect to activity without inhibitors. In the kinase selectivity panel screening for 14 and 19 , the compounds were assayed at single concentration (50 μM) against CDK2, CDK5, CDK9, ERK1, ERK2, PKA, PKBβ, PKCα, PKCγ, GSK-3α, and GSK-3β. All binding assay experiments were performed in duplicate.
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