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ABSTRACT

These experiments totally determine the unknown concentration of the mixed sample that was injected onto the head of the chromatographic column from the standard curve method. Besides that, the goal of this experiment is to identify the compounds present in a mixed sample by comparing the retention time of the peaks obtained. The sample of 1% of Octane, Cyclohexane, and 1% of Hexane are prepared by diluting 1 mL of analyte with 99 mL of solvent of acetone into 100 mL of volumetric flask. From the 1% of Hexane, three standard samples with different concentration are prepared to determine the area obtained and hence, the concentration of the unknown mixed sample could be obtained. The standard sample with 5 mL of 1% of Hexane is diluted to a 20 mL of measuring cylinder with Acetone. This is 0.25% of concentration. The standard sample with 10 mL of 1% of Hexane is diluted into 20 mL of measuring cylinder with Acetone. This is 0.5% of concentration. For 0.75% of concentration, 15 mL of 1% of Hexane is diluted into 20 mL of measuring cylinder with solvent Acetone. Lastly, the mixture of all sample is mixed and vigorously stir in the volumetric flask to produce the mixed sample with unknown concentration. Each of the sample are injected manually into the head of chromatography one by one and peaks and retention time are obtained. The compound present from mixed sample are obtained. There are Octane, (Retention Time - 11.898), Cyclohexane, (Retention Time - 12.370), Hexane, (Retention Time - 12.370) and Acetone, (Retention Time - 10.982). From the calibration graph, the unknown concentration of the mixed sample injected to the column is 0.465 %.

OBJECTIVE To determine the unknown concentration of mixed sample by standard curve method. To identify the compound present in a mixed sample by comparing the retention time of the peaks.

THEORY Chromatography is the separation of a mixture of compounds (solutes) into separate components. Gas chromatography or called gas-liquid chromatography is the method of involving a sample being vapourised and injected onto the head of the chromatographic column. The sample is transferred through the column by the flow of inert or gaseous mobile phase. One or more high purity gases are supplied to the gas chromatography. One of the gases called the carrier gas flows into the injector, through the column and then into the detector. A sample is introduced into the injector with a syringe. The injector is usually heated to 150-250C which causes the volatile sample solutes to vaporize. The vaporized solutes are transported into the column by the carrier gas. The carrier gas must be chemically inert. Commonly used gases include nitrogen, helium, argon, and carbon dioxide. The choice of carrier gas is often dependant upon the type of detector which is used. The carrier gas system also contains a molecular sieve to remove water and other impurities. There are many detectors, which can be used in gas chromatography. Different detectors give different types of selectivity. A non-selective detector responds to all compounds except the carrier gas, a selective detector responds to a range of compounds with a common physical or chemical property and a specific detector responds to a single chemical compound.

Schematic diagram of a gas chromatography

METHODOLOGY CHEMICAL AND APPARATUS n-hexane Isooctane Cyclohexane Acetone GC instrument a) Name: Agilent 6860N b) Column: D=530m, L=30m c) Total flowrate: 23.8mL/min d) Mobile phase: Nitrogen, N2 (P = 400-700 kPa) Hydrogen, H2 (P = 4-7 bar) Compressed air (P = 4-7 bar) e) Pressure: 4.76 f) Heater: SP = 250 C g) Oven: SP = 50 C h) Syringe size: 10.0L i) Injection volume: Manually Volumetric flask (100mL) Measuring cylinder (10mL)

Beaker (100mL) Dropper

PROCEDURE Preparation of Standard sample with standard concentration:

I.

1% of Octane 1 mL of Octane is diluted with 99 mL of acetone in 100 ml of volumetric flask.

II.

1 % of Cyclohexane 1 mL of Cyclohexane is diluted with 99 mL of acetone in 100 ml of volumetric flask.

III.

1% of Hexane

1 mL of Hexane is diluted with 99 mL of acetone in 100 ml of volumetric flask.

IV.

For preparation of 0.25 %: 5 mL of sample 1% of Hexane is diluted with 15 mL solvent Acetone in 20 mL of measuring cylinder.

V.

For preparation of 0.5 %: 10 mL of sample 1% of Hexane is diluted with 10 mL of solvent Acetone in 20 mL of measuring cylinder.

VI.

For preparation of 0.75 %: 15 mL of sample 1% of Hexane is diluted with 5 mL of solvent Acetone in 20 mL of measuring cylinder.

VII.

The entire sixth samples are mixed together and was mixed thoroughly to produce one unknown mixed sample with unknown concentration.

VIII.

2L portion of 1% of Octane is manually injected into GC column and the chromatograms are recorded.

IX.

Step VIII is repeated using 1% of Cyclohexane, 1% of Hexane, 0.25 %, 0.5 % and 0.75 of Hexane.

X.

The compounds present in the unknown mixture are identified from a comparison of retention times.

XI.

The area of mixed sample peak is measured. From the calibration curve, the concentration of mixed sample in the unknown mixture is determined and reported.

RESULT Table 1: Reference Data Sample Name Octane Cyclohexane Hexane 11.964 11.916 11.916 Retention Time (min)

Table 2: Retention time of unknown compound present in the mixed sample Sample Name A B C D 11.898 12.370 12.370 10.982 Retention Time (min)

A- Octane B-Cyclohexane C-Hexane

D-Acetone in mixed sample Table 3: Data for calibration curve Amount 0.25 0.5 0.75 Unknown concentration Area 320.27142 761.28558 1059.69385 626.59680

Graph of Area (mAU.min) Versus Concentration (%)

CALCULATION For preparation of 0.25 % M1V1=M2V2 1 mL x V1=0.25 x 20 mL For preparation of 0.5 % V1 = 5 mL M1V1=M2V2 1 mL x V1=0.5 x 20 mL For preparation of 0.75 % V1 = 10 mL M1V1=M2V2 1 mL x V1=0.75 x 20 mL V1 = 15 mL

DISCUSSION

In this experiment, achievely the amount of unknown concentration is 0.465% and the compound presents from mixed sample are obtained by comparing its retention times. There are Octane, (Retention Time - 11.898), Cyclohexane, (Retention Time - 12.370), Hexane, (Retention Time - 12.370) and Acetone, (Retention Time - 10.982). Retention time is the time it takes a solute to travel through the column. The retention time is assigned to the corresponding solute peak. The retention time is a measure of the amount of time a solute spends in a column. It is the sum of the time spent in the stationary phase and the mobile phase. The mobile phase is a carrier gas, usually an inert gas such as helium or an unreactive gas such as nitrogen. The mobile phase is the phase which moves in a definite direction. The mobile phase consists of the sample being separated/ analyzed and the solvent that moves the sample through the column. The stationary phase is the substance which is fixed in place for the chromatography method. It is a solid or an immobilized liquid on which analyte species are partitioned during passage of a mobile phase.

There are many types of detectors that can be used in gas chromatography. Below are the comparisons of these detectors. Detector Thermal conductivity Flame ionization Application General, respond to all substances All organic substances, some oxygenated products respond poorly. Good for hydrocarbons Sulphur compounds (393 nm), phosphorus compounds (526 nm) All nitrogen and phosphorus containing substances Specific for nitrogen and phosphorus containing substances All organic substances: with ultrapure He carrier gas, also for inorganic and permanent gases All substances that have affinity to capture electrons: no response for aliphatic and naphthenic hydrocarbons Nearly all substances. Depends on ionization method Sensitivity range Fair. 5 100 ng. 10 ppm~100% Very good. 10 100 pg. 100 ppb~99% Linearity Good, except thermistors at higher temperatures Excellent

Flame photometric

Very good, 10 pg S. 1 pg P. Excellent. 0.1 10 pg. 100 ppt~0.1% Excellent

Excellent

Flame thermionic

Excellent

Rubidium silicate head Argon ionization

Very good. 0.1 100 ng. 0.1 100 ppm

Good

Electron capture

Excellent for halogen containing substances. 0.05 1 pg. 50 ppt 1ppm Excellent

Poor

Mass spectrometry

Excellent

CONCLUSION As the conclusion from this experiment, can conclude that the retention time of the mixed sample have been identified. There are Octane, (Retention Time - 11.898), Cyclohexane, (Retention Time - 12.370), Hexane, (Retention Time - 12.370) and Acetone, (Retention Time 10.982). From the calibration graph, the unknown concentration of the mixed sample injected to the column is 0.465 %.

REFERENCES http://www.chromatography-online.org/topics/retention/time.html http://www.chem.ucla.edu/~bacher/General/30BL/gc/theory.html http://delloyd.50megs.com/gaschromatography.whats_new.html