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Determination of Ethanol in Beer by Gas Liquid Chromatography (GLC)

Dec 26, 2017 | 0 comments

Dec 26, 2017 | Essays | 0 comments

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Determination of Ethanol in Beer by Gas Liquid Chromatography (GLC)

Abstract

The method of using gas liquid chromatography for the determination of percent of ethanol in beer has been applied in many breweries. Ethanol percentage in beer is determined by liquid gas chromatography because it is thermally stable and is also volatile as the operating temperatures of gas liquid chromatography. The report details the gas liquid chromatography used in determining the percent of alcohol in a beer. Stella Artois beer was degassed by pouring into two beakers. 400 µl (0.4 cm3) of Propane-1-ol was added in all ethanol concentrations from 0 to 6% v/v then made to the mark with distilled water. The beer sample was prepared by pouring 400 µl of Propane-1-ol into a 10 cm3 volumetric flask and made to the mark with degassed beer. Then the volumetric flasks, which were labeled from 0 to 6 and the one-labeled beer, were taken to the GLC machine where 2 µl were injected each time and twice for each solution on to Gas liquid chromatographs. Finally, the results were printed by chromatograms. The results indicated that the percent ethanol concentration in a beer solution being (4.7%, 4.9%) and (5.5% and 5.4%). This was approximately in the range of alcohol percentage allowed in the beer of 4.8%- 5.2%. The error could have been through human error during the injection, air bubbles in the syringe, or may be the syringe was not cleaned properly.

1. 0 Introduction

Traditional methods of ethanol analysis that involves mass determinations and/or distillations are known to have small inaccuracies because of the presence of other beer components and interfering volatiles (Buckee and Mundy 1993). Similarly, the current recommended reference method for determining alcohol in beers majorly relies on distillation then by a specific distillate gravity measurement. However, the procedure cannot be automated, is labour intensive and requires the operator to possess analytical skills. Moreover, the method of distillation is not specific to ethanol unlike gas chromatography, since it is subject to interference from different materials. Therefore, many laboratories has shifted to other alternative methods which has the capability of automation and are rapid such as gas-liquid chromatography. Chromatograph is a technique of separation where component molecules in a mixture of sample are transported over a stationery phase by a mobile phase. The mobile phase may be a liquid or a gas (solvent system) (Buckee and Mundy 1993). Gas liquid chromatography, with column packing that is appropriately chosen is inherently specific and separates volatile compounds on the basis partitioning properties specific to compounds between gas phase and liquid phase. The volatile constituents’ contribution is approximately in the region of 0.02%V/V (Buckee and Mundy 1993). Further advantages of gas chromatography method is its ability to be amenable to the traceability to National Standards.

The aim of the report is to determine, accurately, the ethanol content of a can of beer by gas liquid chromatography using external standardization, with an internal standard. The work is very important because in determining the content of ethanol in beer, the manufacturer must declare the content of alcohol to the consumer and also to pay the customs duty. Ethanol can be determined by GLC since it is volatile and thermally stable at the operating temperature of GLC

2.0 Method

2.1 Principle

  • The sample of beer was de-gassed by pouring into two beakers
  • As an internal standard, the sample was then diluted with a known Propan-1-ol quantity before going for injection.
  • The diluted liquid sample was injected into a gas chromatography column, which was calibrated previously using standard solutions of ethanol
  • A can of propan-1-ol and ethanol has been used in the experiment. The GLC detector response that uses the integrator which was already set to operate for ethanol analysis included packed carbon wax 20mpolar phase column and 800C, nitrogen carrier gas and ionization detector at 1500C in order to carry out the experiment.
  • The ethanol v/v percentage in the sample was calculated from the ethanol peak areas and Propan-1-ol peaks, using a calibration factor previously calculated

2.2 Reagents

The experiment used only recognised reagents of the analytical grade and the distilled water

  • Ethanol
  • Internal standard, Propan-1-ol
  • Distilled water

2.3 Instruments

  • Gas chromatograph that was fitted with a detector for flame ionization and an oven that was capable of isothermally operate at 1150C.
  • 10 µl gas-tight positive displacement syringe/ gas chromatograph auto sampler- for injecting 1µ or 0.5 of liquid sample
  • Automatic pipettes
  • Chart recorder and integrator. Care was observed when interpreting the signals from the integrators. The given peak areas was checked against a made manual calculation my measuring the width and height of peak. For this reason, a trace of peaks was produced in addition to the value of peak area printed
  • Accurate thermometers to ± 0.10C
  • Chromatography columns, 100/100 mesh packed stainless steel or glass with 20M on 15% Chromosorb WAW DMCS
  • Pipettes- Grade A bulk 20ml, 5ml, 3ml, 1 ml
  • Seven Volumetric flasks- (10cm3)
  • Beakers

2.4 Sample preparation

  1. The beer was degassed by pouring it between two beakers
  2. 400 µl (0.4 cm3) of Propane-1-ol was added in all concentrations from 0 to 6% v/v then made to the mark with distilled water
  3. Beer: 400 µl of propane-1-ol + degased beer to the mark (in volumetric flask marked at 10cm3). See table 1 below
Volumetric flask of 10cm3 0 1% 2% 3% 4% 5% 6% Beer
Propane-1-ol concentration must be constant

at 4% v/v

(400 µl)

400 µl 400 µl 400 µl 400 µl 400 µl 400 µl 400 µl 400 µl
Ethanol at different concentrations 0 100 µl 200 µl 300 µl 400 µl 500 µl 600 µl Make to the mark with degassed beer
Distilled water:

Make to the mark (of volumetric flask of 10 cm3) with distilled water

  1. The group took the solutions to M block, second floor where Gas Liquid Chromatograms were placed
  2. Only 2 µl were injected each time and twice for each solution
  3. The results were printed by chromatograms

2.5.2 Calibration

  1. The gas chromatograph was prepared according to the instructions of the manufacturer. The provided conditions from the manufacturer was as follows
  • Injector temperature 1500C
  • Oven temperature 1150C
  • Detector temperature 1500C
  • Flow rate of Carrier gas 45ml/min
  • Carrier gas Nitrogen
  1. Only 2 µl were injected each time and twice for each solution
  2. Determination of the areas of Propan-1-ol and ethanol internal standard peaks was done
  3. The experiment repeated step 2 and step 3 in duplicate for each of the standard solution

2.5.3 Analysis

  1. The degassed beer sample by pouring between two beakers was thoroughly mixed
  2. Only 2 µl were injected each time and twice for each solution in gas chromatographs
  3. The area of the peaks due to the Propan-1-ol internal standard and ethanol was then calculated

2.6 calculation

Percentage of ethanol is the ethanol’s peak height divided by the peak height of Propan-1-ol and the quantity s divided by the gradient/slope of the line gotten from the calibration curve.

%EtOH =

The percentage of ethanol was calculated from the slope/gradient of the line equation gotten from the calibration curve

Solve for X

Y=mx+b

3.0 Results

The results of the experiments are shown in table 2 below:

Table 2: Determination Of Ethanol In Beer By Gas Liquid Chromatography (GLC)
Ratio peak area Peak area of Peak area of Conc. Of ethanol
(A/B) Propane 1-o1 (B) Ethanol % v/v
    (A)  
0 4809760 0 0
0 5044691 0 0
0.096782159 2377339 230084 1
0.099844018 5646177 563737 1
0.3997279 5214262 2084286 2
0.397057124 5365228 2130302 2
17.23659409 181543 3129183 3
17.93714916 183132 3284866 3
0.746444892 5907205 4409403 4
0.718800408 5776566 4152198 4
0.830910722 670007 556716 5
0.789387662 5973952 4715764 5
1.022640164 5799428 5930728 6
1.0136309 5898950 5979358 6
0.835784621 5681843 4748797 Beer
0.808190871 5771279 4664295 Beer

Ethanol calibration graph

  1. Graph of variation in peak area of ethanol verses ethanol concentration

Table 3

Peak Area of ethanol (A) Conc of Ethanol % v/v
0 0
0 0
230084 1
563737 1
2084286 2
2130302 2
4409403 4
4152198 4
556716 5
4715764 5
5930728 6
5979358 6
4748797 Beer
4664295 Beer
Experimental calculation:

The ethanol concentration of beer at a given peak area of ethanol is calculated as follow:

Where peak area of ethanol is (4748797%v/v, 4664295%v/v). As it can be seen from the graph (figure1) the equation of the straight line is given by

Y=875618x-64188

X= (Y+64188) / 875618

Where Y= peak area of ethanol

X=concentration % v/v

Beer concentration according to the equation will be:

X = (474879764188) / 875618 X = (4664295 64188) / 875618

X=5.5%, X=5.4%

  1. Graph of variation of peak area ratio verses ethanol concentration

Table 4

Parent Ethanol Ratio peak area
0 0
0 0
1 0.096782159
1 0.099844018
2 0.3997279
2 0.397057124
4 0.746444892
4 0.718800408
5 0.830910722
5 0.789387662
6 1.022640164
6 1.0136309
Beer 0.835784621
Beer 0.808190871
Experimental calculation:

The ethanol concentration of beer at a given ration peak area is calculated as follow:

Where ration peak area of ethanol is (0.835784621, 0.808190871). As it can be seen from the graph (figure1) the equation of the straight line is given by

Y = 0.1719x -0.006

X= (Y+ 0.006) / 0.1719

Where Y= ratio peak area

X= concentration % v/v

Beer concentration according to the equation will be:

X= (0.835784621+0.006)/ 0.1719 x= (0.808190871+0.006)/ 0.1719

X=4.9% X=4.7%

Results
Beer Variation in Peak Area % Alcohol
Stella Artois 4748797 5.5%
4664295 5.4%
Beer Ratio peak area % Alcohol
Stella Artois 0.835784621 4.9%
0.808190871 4.7%

4.0 Discussion

During the process of experimentation, there was success after multiple troubleshooting trials. The first obstacle was with the gas chromatography. Moreover, it was difficult to adjust the flow rate.

The results of the experiment from the graph showed the peak area of ethanol gave the concentration of ethanol in beer as 5.5% and 5.4%, and the graph for the ratio peak area gave the concentration of ethanol of beer solution to be 4.9% and 4.7%. Therefore, based on the obtained results, it can be concluded that the contained results from ratio peak area is more accurate than the ethanol peak area since it is more close to the expected value, which is 4.8%v/v.

The standards for the experiment was from 1% ethanol to 6% ethanol. The beer sample use used in the experiment was Stella Artois (can of 568ml). The experiment figured that more peak height could be gotten with more percentage of ethanol, but from the results, no pattern seemed to be formulating. Therefore, the peaks were cut out and massed, where the results showed a general incline in weight as the percent of ethanol increased. Comparison was then made between the beer sample and the calibration curve gotten.

The ratio graph is more likely to give more accurate results because of the proximity of the derived results with the standard ethanol concentration in beer. Moreover, it also factored the input of the values of propan-1-ol unlike the peak ethanol graph which exclusively used the results of ethanol concentration. The peak area of propan-1-ol is not constant despite every solution having concentration of 4% v/v. this could probably be from experimental errors when conducting the experiment.

The two graphs tended to produce a linear graph but there were errors in the values gotten hence the appearance of the final graphs. The report omitted the results of 3%v/v ethanol concentration because of high values obtained due to experimental errors. However, after omitting the results of 3%v/v ethanol concentration, the results closely agreed with the label declaration from the manufacturer. However, there were different values from the two graphs

The error in the differences in the results could probably be from human error during the injection, air bubbles in the syringe, or may be the syringe was not cleaned properly (YCP 2015). The glassware is highly accurate hence errors from the glassware have been ruled out. The only problem experienced during the experiment was the limited time offered for the experiment and the rush manner it was conducted. The results of the experiment can be improved in various ways, however, the simplest way is to repeat the experiment several times and take the average reading for drawing the graph. Moreover, adequate time should be provided to avoid rushing the experiment and to minimise errors.

5.0 Conclusion

The alcohol percentage is usually around 5% and therefore an alcohol content of (4.7%, 4.9%) and (5.5% and 5.4%) was in the approximate range in the recommended value. The experiment determined the percent alcohol in Stella Artois beer and found (4.7%, 4.9%) of alcohol from ratio peak calibration graph and (5.5% and 5.4%) of alcohol from peak ethanol calibration graph. This was close to the average percent of alcohol, Becks and Budweiser, the makers of Stella Artois announced in 2012 (Artois 2015). The company announced that they were reducing the content of alcohol in Stella Artois from 5% to 4.8% (Artois 2015). According to the theory, the obtained results from the graph of ratio peak area should produce more accurate concentration of ethanol in beer. However, the results of the experiment showed close similarity with minor errors. The error in the differences in the results could probably be from human error during the injection, air bubbles in the syringe, or may be the syringe was not cleaned properly (YCP 2015). The results of the experiment can be improved in various ways, however, the simplest way is to repeat the experiment several times and take the average reading for drawing the graph.

6.0 References

Artois, S. (2015). Home. [online] Stellaartois.com. Available at: http://www.stellaartois.com/[Accessed 1 May 2015].

Buckee, G. and Mundy, A. (1993). DETERMINATION OF ETHANOL IN BEER BY GAS CHROMATOGRAPHY (DIRECT INJECTION)-COLLABORATIVE TRIAL. Journal of the Institute of Brewing, 99(5), pp.381-384. Accessed from onlinelibrary.wiley.com/doi/10.1002/j.2050…/pdf

York College of Pennsylvania (YCP), (2015).Faculty.ycp.edu. [online] Available at: http://faculty.ycp.edu/~jleake/instrumental/Beer.ppt [Accessed 1 May 2015].

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