Sevencompact ph ion meter s220

Ageing was undertaken on n = 33 specimens per composition prepared as standard, with a further n = 10 specimens per composition not subjected to ageing as a control group. Individual PEEK/CS specimens were submerged in 10 mL of calcium and magnesium free Dulbecco’s phosphate buffered saline (PBS) solution (Sigma-Aldrich, UK) in screw cap 60 mL capacity clear vessels. For a fixed set of n = 3 specimens, a daily record of PBS supernatant pH value, wet mass, dry mass, wet height, dry height, wet mid-diameter, dry mid-diameter, wet end-diameter and dry end-diameter was recorded. Supernatant pH values were measured using a S220 SevenCompact™ pH/Ion meter (Mettler Toledo, USA) equipped with InLab Expert Pro-ISM pH electrode (Mettler Toledo, USA). PBS supernatant was drained from every specimen vessel and replenished with a further 10 mL. At 7 day intervals, n = 10 specimens per composition were removed from the ageing protocol until day 21.

The anthocyanin extracts were diluted to concentrations of 50 µM in buffers of pH 1–9 ± 0.05. Buffer systems were composed of 0.025 M KCl for pH 1–2, 0.1 M sodium acetate for pH 3–6, 0.25 M TRIS for pH 7–8, and 0.1 M sodium bicarbonate for pH 9. The pH of the buffer systems was adjusted with concentrated HCl or 10% NaOH prior to final volume adjustment. Anthocyanin concentrations of the extracts and isolates were determined by the pH differential methodology [36 ]. Dp-3-rut-5-glu and Pt-3-rut-5-glu isolates were expressed as cyanidin-3-diglucoside-5-glucoside equivalents, while the Dp-3-p-cou-rut-5-glu and Pt-3-p-cou-rut-5-glu isolates were expressed as Cy-3-p-cou-diglu-5-glu equivalents using ε reported by Ahmadiani et al [17 (link)]. After anthocyanins were diluted in buffers, the pH of every sample was verified to be ± 0.05 using a Mettler Toledo Inc. S220 SevenCompact™ pH/Ion meter (Columbus, OH, USA). Samples were equilibrated at room temperature in the dark for 30 min prior to initial analysis. Samples were then sealed and stored in the dark for 72 h at 25 °C to briefly assess the color stability of the pigments. Three replicates were prepared for each treatment.

The measurement of pH transients in the medium using Ricinus cotyledons was similar to that described previously (Komor et al., 1977 ; Hutchings, 1978 (link); Robinson and Beevers, 1981b (link)). The cotyledon still attached to the seedling was incubated in a solution (10 ml) containing 0.25 mM MgCl₂ and 0.5 mM CaCl₂. The solution was stirred continuously and the pH of the solution was monitored every 30 s with a microelectrode and a pH meter (S220 SevenCompact pH/Ion Meter, Mettler Toledo). The pH of the solution bathing the cotyledons decreased to pH 4.6–4.9 after 30 min then stabilized between 4.8 and 5.0. Small aliquots of concentrated solutions of D-GFC or PCMBS (1 mM final concentration) were added at 30 min and small aliquots of concentrated solutions of Suc (20 mM final concentration) were added at 60 min. The pH was monitored continuously over 3 h.