Chemicals/solutions
All solutions were prepared using 99% purity Sigma reagent grade free Gly formula weight (FW) 75.1 g.mol-1 or Glycine hydrochloride Gly.HCl FW 111.5 g.mole-1. Cu2+ solutions were prepared using Copper sulfate penta-hydrate, Cu(SO4).5H2O, formula weight 249.68 g.mol-1. All solutions were prepared by using doubly deionized water. The pH values of all solutions were adjusted using (0.09064 ± 0.00104 mol.L-1) sodium hydroxide (NaOH) solution. The pH values were measured using Orion Membrane pH meter (model 720) with a combination Orion-glass electrode in 0.0 mole.L-1 ionic strength (I).
Preparation of the potentiometric titration solutions
In all free Cu2+, or free Gly, or free Gly.HCl, or Cu2+ -Gly, or Cu2+-Gly.HCl potentiometric titrations in 1:1, and 1:2, and 1:3, and 1:4, and 1:5 ratios, NaOH solution was the titrant. NaOH solutions were prepared from NaOH laboratory grade pellets in carbonate free water. The methods used to prevent the contamination of the titrant with atmospheric CO2 had been described elsewhere.10-15 The NaOH solutions were standardized using primary standard potassium hydrogen phthalate (KHP). Both NaOH and KHP were purchased from Fisher Chemical Co. Before any KHP titration, the KHP was dried at 110˚C for 24 hours and stored in a desiccator. A stock indicator solution of about 0.2% phenolphthalein in about 90% ethanol was prepared from reagent grade phenolphthalein. KHP was titrated to the phenolphthalein end point. Typically, thirteen-fifteen runs were carried out to standardize the NaOH solution. Standard statistical treatments of the data such as the arithmetic mean, standard deviation, T-test, and Q-test were conducted using Excel software.
Potentiometric titrations
The potentiometric titration solutions were contained in a 250 mL beaker equipped with a magnetic stirring bar. The beaker was covered with a custom made Teflon cover. In a typical titration; the Gly or Gly.HCl solutions were added first (in independent experiments) followed by the addition of Cu2+ solution. The mixture was allowed to stand for a minimum of five minutes to reach a state of equilibrium. No other solutions were added to adjust the ionic strength of the solution. The total volume of the final titration solution was 100 mL. The final concentration of the Cu2+ ion titrated was in the range of 2.0 mmoles.L-1. Before each titration, the titration solution mixtures were allowed to stir for an extra 25 minutes for complete equilibrium.
The NaOH titrant was added in segments of 100 mL increments using an Eppendorf micropipette with continuous stirring. The time intervals between the additions of the NaOH solution were set to 5 minutes, which was sufficient to get each of the pH values stabilized and reach complete equilibrium. The start pH-value was in the range of 3-4 (unless otherwise is specified) and the final pH-value was in the range of 10-11. Each titration took about 5 to 6 hours to complete. All titrations were conducted at room temperature.
UV-Vis spectroscopy
We have gathered all UV-Vis spectroscopy measurements on the T60 high-performance spectrophotometer in connection with UVWIN software version 5.0, both purchased from Advanced ChemTech (Louisville, KY). UV-Vis Samples were prepared in D.I. water at 25˚C. The entire UV-Vis spectrum was scanned from 250 to 1000 nm using quartz cuvettes with optical path length of 1 cm. A reference cuvette filled with D.I. water was used with all measurements. The concentration of Cu2+ was = 8.75×10−3 mol.L−1. The UV-Vis spectra were collected at the pH values of 3.00 after 60 minutes equilibrium time and the measurements were repeated after 1440 minutes (24 hours equilibrium time) to ensure complete equilibrium.
IR spectroscopy
All IR spectroscopy measurements were conducted using Nicolet iS10 spectrophotometer in connection with OMNIC software version 8.1, both purchased from Thermo Fisher Scientific (Madison, WI). Samples were prepared in D.I. water at 25˚C. The entire IR spectrum was scanned from 400 to 4000 cm-1 using the provided attenuated total reflectance (ATR) accessory cell compartment equipped with a diamond cell that can accommodate solid samples or aqueous solution samples. The following data parameters were used in collecting the IR spectra: number of sample scans and the number of background scans was set at 32 with resolution of 4.000, and Laser frequency of 15798.7 cm-1. Typical IR spectra were generated in which the X-axis was given as Wave numbers in cm-1 and Y- axis was recorded as % Transmittance.