IC experiment

Ion Chromatography
Lab instructions

The exchange ion mechanism: When the eluent passes through the column, eluent anions bond to the N(+)-R4 and as eluent continueously passes throgh the column the equibrium is reached.

This experiment was conducted with the use of instrumentations of SeQuant
The column is made from N(+)-R4, when the column is continuously washed with NaOH, OH- will bond with N(+) by affinity force. When the sample is injected, for example, Cloride will replace OH- to make a equilibration ions between stationary phase and mobile phase. The more anions have negative charge the more they will be retained in the column.

Compare anions F-, Cl-, Br-, anion Br- is the most retained, then Cl-, and the least retained F-.

Then anion sulfate and phosphate, if the eluent used is NaOH, anion sulphate will give signals first, because it is less negative charge than anion phosphate almost existed in PO4.

if the eluent used is 3.9 mM NaHCO3/ 3.1 mM Na2CO3 ion phosphate(existed HPO4, H2PO4) will give signals first, because it is less negative charge than anion sulphate, so anion sulphate will retain longer than anion phosphate.

General consideration for eluent:

I. Determination of the Number of theoretical plates in the column
Eluent 3.9 mM NaHCO3/ 3.1 mM Na2CO3
@ 1.0 ml/min

The process of separation is described above, the eluent is purged with helium or nitrogen, and filtered with 0.45um pore membrane. The procedure filtration and degassing can be made atomically by connecting to the system or off-line.

Then the sample is injected to the injection port: put the needle in the port on loading manual, and then turn the injection port to ''inject manual''.

Different eluent will give different separation for a sample with the same column. There are two most commonly eluents used for this system.

the first one is NaHCO3/Na2CO3, and the second is NaOH solution

in this experiment, 3.9 mM NaHCO₃/ 3.1 mM Na₂CO3 were used and pumped with flow rate 0.1 ml/min

And this is some factors associated with the instrumentation:

Injection loop: 5um

Regenerant: 18 mM acid sunfuric, pumped with flow rate 3-5 ml/min and suppressor 100 cn croicheted PTFE filament insert.

I. Determination of the number of theoretical plates in the column.

N= 16*(retention time/width peak)2

Inject the solution containing 200 uM sodium chloride and 100uM sulfate ion, the result was observed.

Table 1. The result for ion Chloride and sulfate

Table 2. the result for chloride and sulphate, numbers of theoretical plate

The separation capability compared to other liquid chromatography regards to separation kinetics is fairly good. Although the peaks are fronting, but they were separated well, with ion Chloride appeared first because chloride has less negative charge than ion sulphate so it would be retained in the column in a shorter time.

II. Carbonate Dip

Eluent dips or system peaks and their causes were first described by Gjerde and

Fritz. Stevens et al. described the effect of the system peak in suppressed ion

chromatography. Called a carbonate dip, the system peak was said to be the absent

peak (from the injection) of the carbonic acid that is retained by the unexhausted por-

tion of suppressor column[1].

Prepare standards for

1uM, 200uM, 400uM, 600uM, 800uM, 1000uM Solution containing NaF and NaCl. And then inject each sample we have results below:

Table 3. Results obtained from the sequentially injected samples.

And log(peak areas)/log(concentration) calibration curve with log(peak areas) against concentration:

Calibration curves from floride and Chloride

the slop b(floride) = 1.028

the slop b(cloride) = 0,911

III. Non- linearity due to incomplete dissociation

The eluent: 3.9 mM NaHCO₃/ 3.1 mM Na₂CO3

The sample is sodium acetate

Before injection of the sample, the equilibrium is reached in the column between the the mobile phase and the stationary phase (ion CO₃^2-, HCO₃^-) Mobile phase (ion CO₃^2-, HCO₃^-), the mobile phase then passes through the suppressor, ion Na⁺ is exchanged by suppressor ion H⁺, so the effluent from the suppressor will contain a 7.10^-3 M H₂CO₃, it dissociates following: H₂CO₃ HCO₃⁻ + H⁺ Ka1 = 4,5. 10^-7

HCO₃⁻ CO₃²⁻ + H⁺ Ka2 = 5,6. 10^-11

[H⁺]=559.10^-5, pH = 4.25. This pH is higher than the pH measured = 4.14 because of the process: H2CO₃ CO2 + H₂O

When the sample is injected, the effluent from the suppressor will contain the weak acid CH₃COOH. The charge on ions in solution facilities the conductance of electrical current, the conductivity of a solution is proportional to its ion concentration. The conductivity detector measures conductance of solution containing CH3COO^- that partially forms CH₃COOH, so the conductivity will not proportional to the concentration of acetate and the relationship between log(Sodium acetate uM)&log(peak areas) is non-linear.

CH3COOH CH3COO^- +H⁺ K=1,8. 10^-5

K = [CH3COO^-][ H⁺]/C-[[CH3COO^-]

Degree of dissociation of CH₃COOH with peak total acetate concentration 20uM, α =[CH3COO^-]/C = 0.47

if the peak total acetate concentration equals 20 μM,

H⁺ + F^- -à HF

IV. Influence of pH on the retention Order

Experimental Condition

Eluent: 3.9 mM NaHCO₃/ 3.1 mM Na₂CO3, Inject sequentially a standard containing 0.1mM sulfate ion and another containing 0.4 mM phosphate ion

Change the eluent to 25mM NaOH (Need to wait for equilibration procedure in the column), and then Run separate standards with 0.1mM sulfate ion and 0.4 mM phosphate ion.

Now