The methanol extracts were prepared according to the method described by Pongmalai et al. [19 (link)] with some modifications. Ten grams of each sample was mixed with 17.96 mL of pre-heated methanol (70 °C) to obtain a 70:30 v/v methanol: water ratio. The mixture was then blended using an Ultra-Turrax T25 (JANKE and KUNKEL; IKA Labortechnik, Staufen, Germany) for one minute at 13,500 rpm. This was followed by incubation in a recirculating water bath (Thermoline, Wetherill Park, NSW, Australia) maintained at 70 °C for 30 min under continuous stirring at 100 rpm. Samples were then cooled in ice-water, centrifuged (5000× g/15 min/4 °C), and the supernatants were collected. The extraction was repeated twice. The supernatants were combined and the solvent was removed using vacuum spin dryer (SC250EXP, Thermo Fisher Scientific, Waltham, MA, USA) at ambient temperature (~22 °C). The residue was dissolved appropriately in solvent (acetonitrile/water (85:15, v/v) with 30 mM ammonium formate), filtered through a 0.22 µm membrane filter (Merck Millipore, Billerica, MA, USA) and analyzed by a HPLC method.
Sc250exp
Manufactured by Thermo Fisher Scientific
Sourced in United States
The SC250EXP is a compact, high-performance centrifuge designed for a variety of laboratory applications. It features a maximum speed of 6,000 rpm and a maximum RCF of 5,627 x g. The centrifuge accommodates a range of rotor options and can be programmed to run specific protocols.
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Lab products found in correlation
4 protocols using sc250exp
1
Methanol Extraction of Samples
2
Broccoli Polyphenol Extraction Protocol
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The water extracts of broccoli samples were prepared according to the method previously described by Cai et al. [27 (link)] with some modifications. Four grams of frozen broccoli puree was mixed with 5 mL of boiling Milli-Q water and vortexed for 1 min. The mixture was then incubated in a boiling water bath for 5 min. After cooling in ice, samples were centrifuged at 5000× g for 15 min at 10 °C. Supernatants were collected and a second extraction of the residue was conducted with 5 mL of boiling Milli-Q water and the supernatant was collected. Finally, the supernatants were combined and evaporated to dryness with a vacuum spin dryer (SC250EXP, Thermo Fisher Scientific, CA, USA) at ambient temperature (~22 °C). Dried samples were dissolved appropriately in a solvent (acetonitrile/water (85:15, v/v) with 30 mM ammonium formate), filtered through a 0.22 µm membrane filter (Merck Millipore, Billerica, MA, USA) and analyzed by HPLC.
3
Sulforaphane Extraction and Analysis
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The extraction and analysis of sulforaphane was conducted as described in Cai et al. [28 (link)] with some modification. Briefly, 4 g of frozen sample was mixed with 2 mL of Milli-Q water and vortexed for 1 min. Then, 20 mL of ethyl acetate was added and the mixture was sonicated using a sonication bath (IDK technology Pty Ltd., VIC, Australia) for 5 min followed by shaking for 20 min at 4 °C. The mixture was centrifuged at 5000× g for 15 min at 10 °C and the supernatant was collected. A second extraction was carried out by adding 15 mL ethyl acetate to the residue. The supernatants from the two extractions were combined and dried using a vacuum spin dryer (SC250EXP, Thermo Fisher Scientific, Waltham, MA, USA) at room temperature (22 °C). All samples were dissolved appropriately in 30% acetonitrile, filtered through a 0.22 µm membrane filter (Merk Millipore, Billerica, MA, USA) and analyzed by UPLC.
4
Metabolite Extraction from Macro- and Microscale Cultures
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Metabolites in the macroscale samples were extracted by removing 10 cores of 10 mm diameter from the culture and homogenizing them in 0.01% Tween-20. The homogenized suspension was removed to a glass vial and a volume of solvent, 2.449 ml, proportional to the culture surface area of the cores was added. Samples were agitated every 5 min for 30 min after which they were centrifuged at 2,500 r.p.m. (608g) for 10 min. The solvent layer was removed to a fresh vial for evaporation as below. Metabolites were extracted from the microscale cultures by pipetting solvent into the tapered end of the teardrop channel. Solvent volumes used were proportional to the surface area of the culture; for standard 3.5 mm diameter microscale cultures, 30 μl solvent was used. The metabolites were allowed to passively diffuse into the solvent layer for 30 min after which the solvent was collected into glass vials. Samples were evaporated to dryness (1–4 h) in a vacuum concentrator (Thermo Express SC250EXP) without heat, except samples extracted with γ-caprolactone that were heated to 37 °C for the first hour of evaporation.
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