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Analytical Techniques Lab Report 2: Experiment 4
Chloe Condon
X00147696
DNA & Forensic Analysis
Date:
11th October 2019
Title:
Quantitative analysis of organic waste by capillary GC
Lab partners:
Meagan Dooley, Sandra Kenny, Conor Delargy, Lewis Townsend
Objectives:
The objectives of this experiment were:
To prepare a series of organic solvent samples and an unknown organic waste sample.
To analyze these samples using capillary GC.
To use information from chromatograms produced in order to calculate % composition and response factor of each sample.
To identify what organic solvent is present in the unknown organic waste sample.
To calculate the amount of organic solvent in the samples.
Methods:
As per manual page 10 – 11
Introduction:
47339251252855194310013766802047875809625000-66675148272500459041570389750028575790575000An internal standard (IS) can be defined as a known concentration of substance present in every sample being analyzed. It is used in order to increase precision and accuracy of analysis. Using an internal standard allows the GC conditions to vary during an experiment without inaccurate results being produced. The IS must be structurally similar to the solute of interest.
In this practical, the IS used was ethyl benzene. This compound is structurally similar to all solutions being analyzed because they all contain methyl groups (CH3).
Methanol structure Acetonitrile structure
46291502108201762125582295107632558229538671508909685001152525885825000 Toluene structure
Acetone structure Ethyl Benzene structure (IS)
Response factor is how the detector responds to the sample being analyzed. It can be calculated by the following formula, where F = Response factor:
Conc. of solventConc. of IS =FArea of solventArea of ISA response factor is used in order to represent response differences between the internal standard and the analytes.
In capillary GC, the stationary phase is coated on the inside of the thin column. The compounds separate in the column based on their affinity for the stationary or mobile phase and their polarity. Compounds with a high affinity for the stationary phase will spend some time attached to the column and therefore, will take longer to elute from the system. Compounds that have a high affinity for the mobile phase will elute through the system much faster. Capillary columns are said to provide better efficiency than packed columns that are used in HPLC.
Retention time (Rt) is the amount of time taken for a sample to elute from a chromatography system. Each compound has a different retention time and will separate based on their affinity for the stationary or mobile phase and their polarity as mentioned above. Non-polar compounds will generally have a high affinity for non-polar stationary phases and will have long retention times. The column used in this practical was methyl silicone, which is non-polar. It is expected that other non-polar compounds will have longer retention times than the polar compounds present in the samples being analyzed.
Results/calculations:
Figure 1 showing retention times and peak areas for various samples
Sample Retention time (mins) Peak area (solvent) Peak area
(IS) Total peak area
Methanol 5.13 89802 392441 482243
Acetonitrile 5.17 407984 478263 886247
Toluene 7.17 291888 278925 570813
Acetone 5.26 196428 234240 430668
Figure 2 showing retention times and peak areas for the unknown organic waste sample
Retention time (mins) Peak area
5.15 86070
5.33 110588
7.11 95206
8.61 814560
Sample calculation for calculating solvent concentrations:
Methanol:
Density = 0.792g/ml
Molar mass = 32.04g/mol
Volume used = 1ml
Total volume = 10ml
Mass = density x volume (ml)
Mass = 0.792g/ml x 1ml = 0.792g
Moles= mass (g)molar mass (g/mol)
Moles= 0.792 g32.04g/mol=0.024 molesMolarity= molestotal volume (L)Molarity= 0.024 moles0.01L=2.24 MFigure 3 showing density, molar mass and calculated concentration for various samples
Sample Density
(g/ml) Molar mass
(g/mol) Calculated concentration (M)
Methanol 0.792 32.04 2.4
Acetonitrile 0.786 41.05 1.91
Toluene 0.867 92.14 0.9
Acetone 0.784 58.08 1.3
Ethyl Benzene (IS) 0.866 106.17 0.815
Sample calculation for % composition:
% composition= peak area of solventtotal peak area x 100Methanol: 89802482243 x 100=18.62%Sample calculation for response factor (F):
Conc. of solventConc. of IS =FArea of solventArea of ISMethanol:
2.40.815=F898023924412.94 = F (0.228)
2.940.228=FF = 12.89
Figure 4 showing calculated % composition and Response factor for various samples
Sample Calculated % composition Calculated response factor
Methanol 18.62 12.89
Acetonitrile 46.03 2.74
Toluene 51.13 1.05
Acetone 45.61 1.90
Sample calculation for concentration of methanol in unknown sample.
unknown (x)0.815M=12.8986070392441×0.815M=12.890.219×0.815M=2.822X = (2.822) (0.815M)
X = 2.29M
Figure 5 showing identification of samples, concentration and % composition in the unknown organic waste sample.
Peak number Retention time
(mins) Identified compound Calculated Conc.
(M) Calculated % composition
1 5.15 Methanol 2.29 7.77
3 7.11 Toluene 0.29 8.60
2 5.33 Acetone 0.73 9.99
Discussion:
The results that were obtained in this practical were deemed inconclusive and would not have been accurate to use when calculating the concentration and % composition of the unknown sample. Therefore, a set of results were provided by lab personnel.
left436753000The original results would not have worked due to co-elution in the acetonitrile and methanol samples. There was also no peak observed for acetone in the acetone sample (see images below).
Images showing co-elution in methanol and acetonitrile samples.
3276600425767500
center42862500Image showing no peak for acetone.
Co-elution occurs when compounds elute from the column at the same time, which can make it difficult to identify each separate compound. Separation can be increased by changing the temperature or the column length. Acetone did not give a peak on the chromatogram, only the dichloromethane and the internal standard can be seen above. This may have been due to an error in the making up of the standard. Acetone is sometimes considered to be problematic as it is both a polar and non-polar molecules due to its two methyl groups and the fact it can form hydrogen bonds with water.
The compounds separated based on their affinity for the stationary phase which was a methyl silicone column. The methyl groups in the compound are non-polar, so the compounds in the samples that are also non-polar took the longest to elute.
Methanol is polar, and so did not have a high affinity for the column and eluted the quickest. The internal standard ethylbenzene was the most non-polar among all the samples analyzed and had a high affinity for the column and took the longest to elute in all samples. This also aided as an indication of when the sample had fully ran through the system.
Conclusion:
The unknown organic waste sample was found to contain methanol, toluene and acetone based on the similar retention times to the standards. Peak number 1 came out at a retention time of 5.15 mins, which was similar to that of methanol at 5.13 mins. Peak 2 came out at a time of 7.11 mins, similar to toluene at 7.17 mins. Peak 3 came out at a retention time of 5.33 mins, similar to the time of acetone at 5.26 mins.
