Ion Chromatography 101 submit PDF

Preview

Summary

Ion Chromatography ...

Description

Ion Chromatography Student ID Number: Lab Partner: Mark Eivers. Class: CS351-A. Date of Experiment: AIM The main aim of the experiment was to use Ion Chromatography (IC) to determine and identify ions present in different samples. Peak Spiking was the technique used to identify the ions present in a sample mix. Additional aims were to determine the concentration of nitrate in fertiliser by the standard addition method plot and to determine ions present in Tap water and a Leafy Green solution.

INTRODUCTION Ion Chromatography (IC) is a specialised liquid chromatographic technique that has taken over more traditional techniques used to analyse inorganic anions in the last 15 years. Examples of these include titrimetric, gravimetric and spectrophotometric techniques or the use of ion-selective electrodes. IC is commonly used for ionic species in a solution for a large range of different applications which include pharmaceutical development, food analysis, water quality and control testing. (1) IC methods are split into two groups. The use of an ionexchange column which is called the separator column. This separates sample anions and a second column called the suppressor column is used to reduce the conductivity of the eluent. This will lead to conductometric detection. This is known as the first group and can be described as Suppressed IC methods. The second group is described as Non-Suppressed IC methods and it uses one column only. The one column is known as a low-capacity anionexchanger. An interaction with resin is what will separate the ionic species. The difference in separation is down to the size and charge of the analyte. The ions electrostatic charge is how they are separated on the stationary phase. Ions that are larger in size and have a greater charge, will have stronger interactions with the stationary phase. The stronger interactions will determine the retention times and order of elution of compounds. Sodium carbonate and sodium bicarbonate consisted of the mobile phase and the stationary phase was a cation stationary phase column. An electrical conductivity detector was the detector used where the

ions are separated as they are eluted from the column. Special eluents are used to keep a low background conductivity of the eluent. Aromatic acid anions are used as these special eluents examples of these include: phthalate or salicylate and benzoate. The uses of these aromatic acid anions allow conductivity detection to be successfully employed. (2) The standard addition method is used for the matrix effect. This effect is where there are changes in the analytical signal which are caused by components of the sample other than the analyte. This may occur for quantitative analysis of samples. For standard addition known quantities of analyte are added to the unknown sample. The conditions for standard addition are: there must be linearity in the calibration graph and the analyte calibration curve must go through the origin (3) To achieve a standard addition this is done by pipetting equal volumes of unknown sample into several volumetric flasks. Then the solutions from these volumetric flasks are added to each flask by increasing the volumes of standard. Then each flask is diluted to the same final volume. These flasks all contain different concentrations of standard but have identical concentration of unknown. The analyte signal is measured for each flask and then plotted to the concentration of standard added. To determine the concentration of the unknown solution, the trend line is brought back to where it intercepts the y-axis. This is the signal generated by the analyte in the unknown sample due to there being no standard added to that solution. (2) The analytes in this experiment were Bromide, Chloride, Fluoride, Nitrite and Nitrate. Bromide, Chloride and Fluoride ions are derived from Bromine, Chlorine, Fluorine and belong to the Group 17 on the periodic table, which are halogens. These are known as salt formers. They are non-metallic elements and are diatomic molecules. As you go down the periodic table the boiling points increase. The Van der Waals forces increase, size and atomic mass also increase as you go down the group on the periodic table for Fl, Cl then Br. Fl, Cl and Br have very low melting and boiling points and an oxidation number of -1 as they contain seven electrons on their outer shells. Fluorine and Chlorine are gases at room temperature and Bromide is a liquid at room temperature. (4) These are the most reactive of all chemical elements as they are electronegative and will gain electrons fast and they will dissociate into atomic particles with ease. There combination with other elements forms them into compounds and they are not good conductors of electricity and heat. Nitrite and Nitrate are the other analytes of interest. There chemical formulas are NO2- and NO31- .These are polyatomic ions and covalent compounds that were formed from nitrogen and oxygen. Nitrate has three oxygen atoms and Nitrite has two oxygen atoms and one

nitrogen atom. (5) Nitrate and Nitrites are found in water and soil and can be produced by bacteria. Nitrite molecular mass is 46.006 g/mol. Nitrate molecular mass is 62.05 g/mol. (6) When

temperatures are very high Nitrate and Nitrite can become explosive although they are

stable compounds at room temperatures. Nitrates may form NO or/and NO2 but Nitrites on the contrary will decompose NO and N2. Nitrates and Nitrites may become destabilized by chlorides and organic material. Nitrates in the form as a salt are colourless, odourless and are known as hygroscopic. Nitrites in the form as salts are yellow or sometimes will be colourless and are also known as hygroscopic. (7) The analytes interactions with the stationary phase will show the suitability of the mechanism for the analytes. The type of interaction between the stationary phase and the analyte in IC is an ionic interaction. The analytes in this experiment are anions so anion retention will take place on the stationary phase. The stationary phase (SP) as discussed previous is a cation stationary phase column. The SP is where the separation takes place and the SP properties are very important for a good separation. (8) Within the stationary phase the functional groups on the surface of the SP are fixed positive charged species known as M+. The counter ions known as C- are in the mobile phase as will neutralize the fixed positive charged species when equilibrium is achieved. The analytes are known as the anionic samples: A-. The anionic samples enter the column and go between the mobile and stationary phase where the interactions occur. The anionic samples take the position of the counter ions and swap over many times with the mobile phase ions. This can take place in the column as the analytes are negatively charged and the column is the opposite so its positively charged. The competitive nature of the sample components and the anions means that there is a distribution equilibrium. The equation below shows this. M + C- + A- → M + A- + C This ion-exchange reaction won’t occur if the column was not charged and if there was no stationary or mobile phase present. However, this is not the case so this makes this separation method very suitable for the analytes. Considering throughout the ion-exchange process the electro-neutrality has to be constant, this means that the ion exchange is stoichiometric. This is because there is a single monovalent counter ion that is displaced by a single monovalent anion. Cation retention is also another retention mechanism. The stationary phase would have to be changed to have a negative charge and the counter ions would have a positive charge. These are more reasons why this method is suitable for the analytes of interest. (9)

Ion chromatography is a type of separation technique that can be compared to High Performance Liquid Chromatography. Both uses a liquid mixture sample that is dissolved into a mobile phase. In both separation techniques the mobile phase passes through the Stationary Phase and the SP column is both made of a certain type of packing material. The separation of the compounds takes place in the SP and then pass through a detector that will register a signal. The graph’s that are produced contain peaks of different elution times that illustrate the interactions between the analytes and the Stationary phases. Base line resolved peaks with good resolutions is most desirable for both techniques. These graphs show the signal intensity from the detector and the time the compounds elute at. Although these two separation techniques have some similarities they are still different separation techniques. IC analyses samples for cations and anions. HPLC separate different compounds based on their polarity and the term ‘like for like’ is often discussed. This term means that if the analytes has the same polarity as the stationary phase there will be stronger interactions and thus will be retained for longer. In these two separation techniques the quantities of each component can be found if they were to be compared to a standard. This standard can only be compared to if this was analysed under the exact same conditions and operating parameters as the analyte of interest. On comparison on popularity HPLC is more widely used around the world as there are many more operating parameters such as flow rate, column length, column temperature, compositions of mobile phase and detector type. These operating parameters give more options to the lab technician of which can be altered to change certain components of the method such as the order of elution or the retention time of the compounds. Thus making it a more popular separation technique. HPLC differs greatly to IC with regards to its instrumentation. IC instrumentation contains non-metal components. This is so to avoid contamination due to leaching. IC uses specific conductivity detectors and columns. This specialized equipment allows for quantitation at low concentrations. (10)

MATERIALS AND METHOD Reagents: Eluent Preparation: Sodium carbonate, sodium bicarbonate and deionised water. Preparation of anion standards: Fluoride, Chloride, Nitrite, Bromide, Nitrate and deionised water. Identification of ions and the determination of nitrate concentration in commercial plant fertiliser by the standard addition method: Commercial plant fertiliser, bromide solution, nitrate solution and deionised water. Determination of ions present in Tap water and Leafy greens: Tap water, spinach (leafy greens), bromide solution and deionised water. Instruments: Dionex Model ICS 1500 with in-built conductivity detection was the instrument used for the suppressed ion chromatography experiment. The PC controlled instrument utilised Chromeleon software and had an IonPac AS22 4x25 cm analytical separation column, AG 22 Guard Column and an ASRS300 4 mm suppressor column. Procedure: The experiment was carried out in accordance with laboratory manual CS351-A pages 36 to 40. PRE-LAB CALCULATIONS MW of Sodium Bicarbonate: 84.01 g/mol MW of Sodium Carbonate 105.9885 g/mol Sodium Bicarbonate: 1Mm = 0.001M 1.4Mm = 0.0014M Mass = MW x Molarity 84.01 g/mol x 0.0014M = 0.1776 g

Sodium Carbonate Mass = 0.4769 g Preparation of Anion Standards – 100 ml at concentration of 100 mg/L Sodium Fluoride: MW of Sodium Fluoride = 41.99 MW of Fluoride = 18.99 g/mol 100 mg/L = 0.1 g/L = 0.0001 g/ml Concentration / MW of Fluoride = 0.0001 g/ml / 18.99 g/mol = 5.262x10-6 5.262x10-6 x 100 ml = 5.2659x10-3 mole / 100ml 5.2659x10-3 mole / 100 mL x MW of Sodium Fluoride 5.2659x10-3 mole / 100 mL x 41.99 g/mol = 0.02211g

Table 1: Amount needed for 100mL aqueous mixed anion standard solution at100 mg/L. Anion Standard

Amount Required (g):

Actual Amount weighed

Sodium Fluoride Potassium Chloride Sodium Nitrite Potassium Bromide Potassium Nitrate

0.0221 0.0165 0.0149 0.0129 0.0169

(g) 0.022 0.0158 0.0160 0.0121 0.0158

For the concentration of 200 mg/L the same calculations were used, the volume was 50ml. The Nitrite was an exception as 100ml was the volume using a 200 mg/L concentration. Table 2: Amount needed for 50mL individual standard solutions of anions at 200 mg/L concentration, with exception to Nitrate which was prepared at 100ml of 200 mg/L concentration. Anion Standard

Amount Required (g):

Actual Amount weighed (g)

Sodium Fluoride Potassium Chloride Sodium Nitrite Potassium Bromide Potassium Nitrate

0.0221 0.0165 0.0149 0.0129 0.0338

0.024 0.0170 0.0149 0.0125 0.0340

Standard Addition Solutions Solution 2 Conc of nitrate required in final solution = 20 mg/L Conc of Nitrate Standard = 200 mg/L Volume = 25ml 200 mg/L / 20 mg/L = 10 dilution factor 25 ml / 10 = 2.5 ml of nitrate required

Table 3. Volumes of nitrate standard required for the Standard Addition Solutions in 200 mg/L concentration of nitrate. Solution

Vol of

Conc of nitrate

Volume of 200

Final Vol

fertiliser

required in final

mg/L nitrate

(ml)

solution ml

solution (mg/L)

standard to add (mL)

(bromide as 1 2 3 4 5

I.S.) 10 10 10 10 10

0 20 50 80 100

0 2.5 6.25 10 12.5

25 25 25 25 25

DISCUSSION AND RESULTS Ions are stripped of an ion exchange material by other ions, this is known as elution in IC. If ions have a greater affinity for the material on the stationary phase or if their concentration is higher than others, then this will take place. The order of elution allows the order to be predicted and controlled by knowing the analytes chemical properties. The analytes are absorbed onto the adsorbent in the column. The properties of the adsorbent will result in the analytes having different affinity’s in the stationary phase. The affinity the analytes has to the adsorbent results in a thin film on the columns surface. The eluent solvent displaces the analytes when it is run through the column and then the analytes runs completely through and out of the column. This is what is known as the elution process. In this experiment carbonate was used as the eluent, this eluent gives a total ionic strength. To give optimum retention times and sensitivity between monovalent and multivalent sample ions, the monovalent and divalent ions are varied. The low pH of 4 which the eluent possess means it converts the analytes into strong acids to allow for dissociation to take place and it will also have control of the dissociation of weak acids. For this experiment the molecules with the weakest ionic interactions will elute first and the strongest ionic interactions will elute last. Determination of ions in sample mix by ‘peak spiking’ Table 4: Retention times of mixed anions from figure 1A with a eluent flow rate of 1.2 and back pressure of 1900-2300 psi. Anions Fluoride Chloride Nitrite Bromide Nitrate

Retention Time (min ) 3.04 4.22 4.96 5.87 6.49

From the chromatogram of the five peaks Fluoride anions eluted first, then Chloride. This is because fluoride has the highest electronegativity of the analytes with 4.0. When the electronegativity’s are equal between analytes then the analyte who has the lower molecular weight will elute first as discussed previous. Chloride and Nitrite have the same

electronegativity. This was the case with Chloride as it has a lower molecular weight than Nitrite. Nitrite then eluted after Chloride. Bromide eluted then after Nitrite. Nitrogen has a higher electronegativity of 3.0 then compared to Bromide which is 2.8. So Nitrogen should have eluted first but this was not the case as the overall order of elution was Fluoride, Chloride, Nitrite, Bromide and lastly Nitrate. Nitrate in theory should of eluted before Bromide because of its higher electronegativity but because the Nitrate anion has an extra oxygen molecule than compared to nitrite as discussed in the introduction this will decrease then electronegativity. Thus bromide eluted before Nitrate. The elution times perpetrate this as Fluoride has the lowest retention time of 3.04 due to it having the highest electronegativity of the analytes thus it had a lower affinity with the stationary phases so it passes through the column faster and is eluted first. If this was compared to Nitrate which had a retention time of 6.49 min this was because Nitrate had the strongest affinity in the stationary phases thus it was adsorbed for longer on the packing material and then it was eluted last after the rest of the analytes. It can be stated that Bromide then had the second strongest affinity for the stationary phase then followed by Nitrite. Calculations Resolution = 2(t2 – t1)/(W2 + W1) Resolution calculation for fluoride and Chloride. Rs = 2(4.22- 3.04) / (0.35 + 0.40) Rs = 3.14 Retention Factor for Fluoride k’ = (tR – to)/to k’ = (3.04 – 0.39) / 0.39 k’ = 6.76 Separation Efficiency (N) for Fluoride N = 5.54(tR/W1/2)2 N = 5.54 (3.04/0.2)2

N = 1279.96 of theoretical plates

Table 5: This table illustrates the calculated Resolution, retention factor and Separation efficiency of all the peaks, eluent flow rate was1.2 ml/min and back pressure of 1900-2300 psi. Anion’s

Resolution

Anion Peak Retention

Peaks

Factor (k’)

Separatio n Efficiency

Fluoride and

3.14

Fluoride

6.76

(N) 1279.96

Chloride Chloride and

1.78

Chloride

9.82

3221.5

Nitrite Nitrite and

1.86

Nitrite

11.7

2366.19

Bromide Bromide and

1.078

Bromide

14.05

3054.25

Nitrate

15.64

2209.18

Nitrate

From viewing the chromatogram all of the peaks are not base line resolved. The peaks with the greatest resolution is the Fluoride and Chloride peak as they have a resolution of 3.14 which is greater than the standard 1.5 for resolution. Although the highest resolution doesn’t indicate the greatest as the peaks can’t be too spaced apart. This is not the case for these two

peaks as they are well spaced apart but only by less than a minute which keeps the run time low which is what is wanted. Bromide and Nitrate have the worst resolution with 1.078 and this is evident from the chromatogram as the peaks are not base line resolved and are eluting very close to each other as they are almost co-eluting. The retention factors are all very high but this is because the t0 was eluted out of the column very fast at 0.39 minutes which indicates that it had a very low affinity for the stationary phase. The most efficient separation was for the Bromide as it had 3054.25 theoretical plates. The more number of theoretical plates the better as it shows the separation was very efficient. Theoretical plate is when the analytes goes from the mobile phases into the stationary phase and back into the mobile phases again and this would illustrate one theoretical plate. The least efficient separation was Fluoride which had an N value of 1279.96 theoretical plates and this was because it had a very low affinity for the stationary phase because of its high electronegative number which was 4. This means it went from the mobile phase into the stationary phase then back into the mobile phase the least amount of times then compared to any other anion. Identification of ions and the determination of nitrate concentration in commercial plant fertiliser by the standard addition method Table 6: This table illustrates the Conductivity of each run as 0, 20 and 50 mg/L concentration. Concentratio n (mg/L): 0 20 50

Conductivit y Run 1 (µS): 14.1 14.09 17.7

Conductivit y Run 2 (µS): 14.2 15.6 18.9

Average (µS):

Standard Deviation

%RSD

14.15 14.845 18.3

0.070 1.067 0.8...