Pterostilbene was purchased from Sigma-Aldrich. The stock solution of Pterostilbene was prepared in dimethyl sulfoxide (DMSO) (Sigma-Aldrich) and further diluted in a sterile culture medium to the desired concentrations immediately before use. The final DMSO concentration in the working solutions was 0.1%.
Pterostilbene
Manufactured by Merck Group
Sourced in United States, Germany, Canada
Pterostilbene is a laboratory reagent used as a standard compound for analytical and research purposes. It is a naturally occurring stilbenoid compound structurally similar to resveratrol. Pterostilbene serves as a reference standard for identification, quantification, and purity analysis of samples containing this compound.
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Lab products found in correlation
Resveratrol, by Merck Group (13 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (9 mentions)
Piceatannol, by Merck Group (4 mentions)
Trans resveratrol, by Merck Group (3 mentions)
β actin antibody, by Merck Group (2 mentions)
Sybr green pcr master mix, by Thermo Fisher Scientific (2 mentions)
T blunt vector, by Solgent (2 mentions)
Pinostilbene, by Tokyo Chemical Industry (2 mentions)
Cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
β actin, by Cell Signaling Technology (2 mentions)
Erythromycin, by Meilun (1 mentions)
Streptomycin sulfate, by Meilun (1 mentions)
Nf κb p65, by Santa Cruz Biotechnology (1 mentions)
α tubulin, by Santa Cruz Biotechnology (1 mentions)
Horseradish peroxidase hrp conjugated secondary antibody, by Merck Group (1 mentions)
Proliferating cell nuclear antigen (pcna), by Santa Cruz Biotechnology (1 mentions)
Phospho iκb, by Santa Cruz Biotechnology (1 mentions)
Phospho akt, by Santa Cruz Biotechnology (1 mentions)
Caffeic acid, by Merck Group (1 mentions)
38 protocols using pterostilbene
1
Pterostilbene Preparation and Dilution
2
Characterization of mcr-1-producing Isolates
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Human clinical MCR-1 producing isolates K. pneumoniae ZJ02, K. pneumoniae ZJ05 and E. coli ZJ40 were collected in our previous study (Wang et al., 2017 (link)). And the mcr-1 gene was chromosomally located in E. coli ZJ40. K. pneumoniae E8.31, K.pneumoniae 13b5 and K.pneumoniae L18 were collected from food animals. We also used E. coli strain DH5α (pUC19-mcr-1) (Zhou et al., 2018 (link)), which carries a mcr-1 gene originating from K. pneumoniae ZJ05. Polymyxin-resistant mcr-1-negative K. pneumoniae isolate 16ZJJ9-19BC was obtained from a chicken cloacae sample collected in Zhejiang, China. E. coli ATCC25922, K. pneumoniae ATCC700603 and K. pneumoniae K7 were used as quality control strains. Pterostilbene (≥97% HPLC-pure) was purchased from Sigma-Aldrich (St. Louis, MO, USA). Colistin sulfate, polymyxin B sulfate, penicillin, imipenem, gentamicin sulfate, and chloramphenicol were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). Cephalothin sodium, streptomycin sulfate, kanamycin sulfate, erythromycin, and acheomycin were purchased from Dalian Meilun Biotechnology Co. (Dalian, China). Stock solutions of Pterostilbene were prepared in dimethyl sulfoxide (Sigma-Aldrich).
3
Antibacterial Potency of Resveratrol and Pterostilbene
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The antibacterial activity of resveratrol and pterostilbene (Sigma-Aldrich, St. Louis, MO, USA). was evaluated by MIC and MBC determination. A 2-fold broth-dilution method was utilized to assess MIC. The bacterial population was exposed to several dilutions of compounds ranging from 0.018 to 10 mM and incubated at 37°C for 16 h. An ELISA reader was used to detect MIC at 595 nm. MIC was measured as the highest dilution revealing no bacterial growth. For MBC assay, the bacteria were diluted in PBS and positioned on plates. The compounds with different dilutions were incubated with the microorganisms for 16 h. The colony-forming unit (CFU) was counted. The highest dilution, which resulted in a 99.9% reduction of cell numbers, was recognized as MBC.
4
Stilbene Compounds Preparation and Use
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Resveratrol and pterostilbene were purchased from Sigma-Aldrich (St. Louis, MO, USA). Gnetin C was a generous gift from Hosoda SHC Co., Ltd. (Fukui, Japan). All compounds were dissolved in dimethyl sulfoxide (DMSO) such that final concentration was 0.1% DMSO.
5
Isolation and Purification of Stilbenoids from Rheum undulatum
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Stilbenoids were purified from R. undulatum as described previously.[34 (link)] Briefly, the methanolic extract of the dried rhizoma of R. undulatum was suspended in water and partitioned with ethyl acetate. The ethyl acetate soluble fraction was subjected to column chromatography on silica gel eluting with n-hexane-ethyl acetate (20:1→1:1) and CHCl3-MeOH(10:1→1:1) to obtain six fractions. From these fractions, pterostilbene, rhapontigenin, resveratrol, and piceatannol were isolated with column chromatography and the structure of isolated compounds were conformed as described previously. [34 (link)] The pterostilbene purchased from Sigma Aldrich (Saint Louis, MO) were also used.
6
Pterostilbene Signaling Pathway Analysis
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Pterostilbene, β-actin antibodies, dimethyl sulfoxide (DMSO), and other reagents and chemicals used in our experiments were obtained from Sigma-Aldrich Company (St. Louis, MO, USA). All the monoclonal primary antibodies, including NF-kappa (NF-κ) Bp50, NF-κ Bp65, phospho-IκB, phospho-IKK(Ser176), AKT, phospho-AKT, ERK(p44/42), phosphor-ERK(p44/42), Egr-1, secondary antibody (FITC conjugated goat anti-mouse IgG (1: 100 dilution), α-tubulin, and PCNA were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The primers for control, actin, and TNF-α, and 2X SYBR Green PCR Master Mix were obtained from Life Technologies. Horseradish peroxidase (HRP) conjugated secondary antibodies were obtained from Chemicon International (Temecula, CA, USA).
7
Heterologous Expression of Polyphenol Synthases
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The strains and plasmids used in this study are listed in Additional file 1 : Table S1. Antibiotics were added to the medium as required at the following concentrations: ampicillin, 100 mg/L; kanamycin, 50 mg/L. T-blunt vector (Solgent, Korea) was used in the polymerase chain reaction (PCR) cloning. pET-22b(+) and pET-28a(+) were purchased from Novagen (USA). Caffeic acid, ferulic acid, resveratrol, and pterostilbene were purchased from Sigma-Aldrich (USA), pinostilbene was purchased from Tokyo Chemical Industry, Co (Japan) as a standard for compound identification by HPLC.
8
Resveratrol and Pterostilbene Treatment
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Resveratrol (>99 % pure; GC) and pterostilbene (>97 % pure; HPLC) were purchased from Sigma-Aldrich. The compounds were prepared in dimethyl sulfoxide (DMSO), which was obtained from Sigma-Aldrich and stored at a stock concentration of 50 mM at –20 °C. Cells were treated with fresh Resveratrol (Res) and pterostilbene (Ptero) every 24 h (one day) for up to 72 h (three days) after seeding. DMSO (1 μL/1 ml) was used as the vehicle control.
9
Prostate Cancer Cell Culture and Treatment
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Prostate cancer 22Rv1 cells (ATCC, Manassas, VA, USA) were grown and maintained in RPMI-1640 media containing 10% FBS, as described previously [27 (link),62 ]. Cells were validated for mycoplasma-free condition using the Universal Mycoplasma Detection Kit (ATCC, Manassas, VA, USA). Gnetin C was a generous gift from Hosoda SHC Co., Ltd. (Fukui, Japan). Pterostilbene was purchased from Sigma-Aldrich (St. Louis, MO, USA). Compounds were dissolved in pure dimethyl sulfoxide (DMSO, 0.1% final concentration) and stored in the dark at −20 °C until use. At approximately 60% confluency, cells were treated with Pterostilbene and gnetin C at the same concentration of 25 µM for 24 h, after which, protein lysates were isolated for Western blot analysis, as described above. The dilutions for antibodies used in cell lines were the same as those used for tissues.
10
Standardization of Grape Powder Extract
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Grape powder, which is a proportional representation of the different varieties of table grapes grown in California, was obtained from the California Table Grape Commission. The grape powder extract (GPE) was prepared and standardized as described previously [37 ] and was a generous gift from Dr. Richard van Breemen (Linus Pauling Institute, Oregon State University, Corvallis, OR, USA). Resveratrol and pterostilbene were purchased from Sigma-Aldrich (St. Louis, MO, USA) and were dissolved in dimethyl sulfoxide (DMSO) for the in vitro experiments.
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