05 expiremental approach___affecting retention time
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An Experimental Approach forAn Experimental Approach for
Students to Learn about FactorsStudents to Learn about FactorsAffecting Retention Time in GLCAffecting Retention Time in GLC
Scott DonnellyScott Donnelly
Arizona Western CollegeArizona Western CollegeYuma, AZYuma, AZ
Colorado River
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Presentation TopicsPresentation Topics
Introduction to AWCIntroduction to AWC
Educational Benefits of GC LabsEducational Benefits of GC Labs
Prelab Exercise/QuestionsPrelab Exercise/Questions
Postlab Assessment QuestionsPostlab Assessment Questions
Student CommentsStudent Comments
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AWC
TucsonMEXICOMEXICO
Grand CanyonGrand Canyon
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Public TwoPublic Two--YearYear
CollegeCollege
6,000 FTSE (Full6,000 FTSE (Full--
time Studenttime Student
Equivalents)Equivalents)
Partnerships withPartnerships with
NAU & UANAU & UA
New 40,000ftNew 40,000ft22
AgricultureAgriculture--ScienceScience
bldg., Spring 2007bldg., Spring 2007
Arizona Western CollegeArizona Western College
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Driving Forces for GC LabsDriving Forces for GC Labs
Earlier work with UVEarlier work with UV--Visible SpectroscopyVisible Spectroscopy
Chemistry of ColorChemistry of Color labslabs
Students could mathematically manipulate A =Students could mathematically manipulate A = --log Tlog Tbut had little understanding why red food coloring isbut had little understanding why red food coloring is
redred
Consistent exposure to instrumentation used inConsistent exposure to instrumentation used inmodern scientific inquirymodern scientific inquiry
Why wait until organic to use GC?Why wait until organic to use GC?
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Classes: Organic Chemistry, General Chemistry 2, &Environmental Science
GC Prelab:
Conceptual questions
5-10 minutes in length
Four consecutive weeks
GC Lab: runs simultaneously
with scheduled lab
GC Postlab Assessment Questions:
Embedded in quizzes and exams Take home lab exercise(s)
Chromatography/Visible Spectroscopy Exam
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Educational BenefitsEducational Benefits
Demystify instrumentsDemystify instruments-- notnot magicalmagical
black boxesblack boxes.based on known.based on knownchemical principleschemical principles
Increase breadth of labsIncrease breadth of labs Expand lecture topic coverageExpand lecture topic coverage
Illustrate complementaryIllustrate complementaryrelationship between theory andrelationship between theory and
applicationapplication
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GC Labs:GC Labs:PotentialPotentialFactorsFactors
Affecting Retention Time, RAffecting Retention Time, Rtt
Molecular WeightMolecular Weight
Boiling PointBoiling Point
Oven TemperatureOven Temperature
Injector TemperatureInjector Temperature
Carrier Gas Flow RateCarrier Gas Flow Rate
Sample Volume InjectedSample Volume Injected
Clarus 500 GCClarus 500 GC
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Chromatogram of Paint ThinnerChromatogram of Paint Thinner
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Every peak, regardless ofEvery peak, regardless of
its height, throughout theits height, throughout the
chromatogram representschromatogram represents
at least one compound.at least one compound.
xx--axis:axis: Time (minutes)Time (minutes)
yy--axis:axis: Detector ResponseDetector Response(mV)(mV)
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Clarus 500 Gas ChromatographClarus 500 Gas Chromatograph
Students shown GC and the majorStudents shown GC and the major
hardware componentshardware components-- injector,injector,
oven, column, and detectoroven, column, and detector-- areare
described briefly.described briefly.
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GasGas--liquid chromatography, or GC for short, physically separatesliquid chromatography, or GC for short, physically separates
liquids in a mixture according to a number of factors. Suppose yliquids in a mixture according to a number of factors. Suppose yououcarry out a GC analysis of an unlabeled bottle. Using a smallcarry out a GC analysis of an unlabeled bottle. Using a small
volume syringe you withdraw some sample from the bottle andvolume syringe you withdraw some sample from the bottle and
inject it into the GC. The chromatogram produced is shown below.inject it into the GC. The chromatogram produced is shown below.
Retention timeRetention time (R(Rtt) is the difference in time between when a) is the difference in time between when asubstance was injected and when it exits the GC into the detectosubstance was injected and when it exits the GC into the detector.r.
Answer the questions below.Answer the questions below.
start Time (arbitrary units)
A) Which peak represents the compound that travels through theA) Which peak represents the compound that travels through the
GC instrument in the least amount of time?GC instrument in the least amount of time?
B) Which peak has the shortest RB) Which peak has the shortest Rtt? the longest?? the longest?
Prelab 1: Retention Time, RPrelab 1: Retention Time, Rtt
A B C DA B C D EE
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Prelab 1: Molecular WeightPrelab 1: Molecular Weight
GasGas--liquid chromatography, or GC for short, physicallyliquid chromatography, or GC for short, physicallyseparates liquids in a mixture according to a number ofseparates liquids in a mixture according to a number of
factors, one of which is molecular weight (MW). Suppose afactors, one of which is molecular weight (MW). Suppose a
mixture of three liquid hydrocarbonsmixture of three liquid hydrocarbons-- AA,, BB, and, and CC-- waswasinjected into a GC with constant oven temperature, Tinjected into a GC with constant oven temperature, Tovenoven..
The relative MWs of the three hydrocarbons areThe relative MWs of the three hydrocarbons are BB >> AA >> CC..
Based on this information arrange the three compounds inBased on this information arrange the three compounds in
the order that they elute or exit from the column. Thenthe order that they elute or exit from the column. Then
explain your reasoning.explain your reasoning.
start Time (arbitrary units)
= A = B = C
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Postlab 1 Assessment Questions:Postlab 1 Assessment Questions:
RRttand Molecular Weightand Molecular Weight
1) Suppose there are four colorless alcohols labeled A, B, C & D1) Suppose there are four colorless alcohols labeled A, B, C & D, all of, all ofwhich are liquids at room temperature. With respect to theirwhich are liquids at room temperature. With respect to their
molecular weights B > D > C > A. GC analysis was done at Tmolecular weights B > D > C > A. GC analysis was done at Tovenoven==
115115OOC. Arrange these four alcohols with respect to their retentionC. Arrange these four alcohols with respect to their retention
times.times.2) A mixture of four hydrocarbons was injected into a GC at T2) A mixture of four hydrocarbons was injected into a GC at Tovenoven ==
8585OOC, giving the chromatogram below. Which compound spends theC, giving the chromatogram below. Which compound spends the
most time in the liquid phase?most time in the liquid phase?
3) Of the choices below, which is predicted to have the longest3) Of the choices below, which is predicted to have the longest RRtt??
a. CHa. CH33CHCH33 b. CHb. CH33CHCH22ClCl c. CHc. CH33CHCH22CHCH22CHCH22CHCH22OHOH
d. CHd. CH33CHCH22CHCH22CHCH33 e. CHe. CH33CHOCHO
start Time (arbitrary units)
AA B CB C DD
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Postlab Assessment Question:Postlab Assessment Question:
Molecular WeightMolecular Weight
A mixture of three liquid saturated hydrocarbonsA mixture of three liquid saturated hydrocarbons-- AA,, BB, and, and CC--was injected into a GC under isothermal heating conditions atwas injected into a GC under isothermal heating conditions at
8585OOC. The GC was outfitted with a 30m, 0.250mm i.d. capillaryC. The GC was outfitted with a 30m, 0.250mm i.d. capillary
column. The resulting chromatogram is shown below.column. The resulting chromatogram is shown below.
Suppose the three hydrocarbons wereSuppose the three hydrocarbons were22--methylhexanemethylhexane,,pentanepentane,,
andand2,2,32,2,3--trimethylhexanetrimethylhexane. Of the three which is represented by. Of the three which is represented by
B in the chromatogram above?B in the chromatogram above? Offer an explanation to supportOffer an explanation to support
your choice.your choice.
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4.30 4.40 4.50 4.60 4.70 4.80 4.90 5.00 5.10 5.20 5.30 5.40 5.50 5.60 5.70 5.80 5.90
AA BB CC
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Prelab 2: Boiling PointPrelab 2: Boiling Point
The chromatogram below is for a two component liquidThe chromatogram below is for a two component liquid
mixture, containing the alcohols 1mixture, containing the alcohols 1--propanol and 2propanol and 2--
propanol. Both alcohols have the molecular formula Cpropanol. Both alcohols have the molecular formula C33HH88OO
and hence the same molecular weight (MW). Yet, as shownand hence the same molecular weight (MW). Yet, as shown
the two alcohols have different retention times (Rthe two alcohols have different retention times (Rtt) under) underidentical GC conditions. Since their MWs are the same,identical GC conditions. Since their MWs are the same,
their differences in Rtheir differences in Rtt cannot be explained on account of acannot be explained on account of a
difference in MW. So what then is a plausible explanationdifference in MW. So what then is a plausible explanationfor why they have different retention times?for why they have different retention times?
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2.15 2.20 2.25 2.30 2.35 2.40 2.45 2.50 2.55 2.60 2.65 2.70 2.75 2.80 2.85 2.90 2.95 3.00
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Prelab 2: RPrelab 2: Rtt & Molecular Weight& Molecular Weight
As you learned last week gasAs you learned last week gas--liquid chromatography physically separatesliquid chromatography physically separates
liquids in a mixture according to a number of factors, one of whliquids in a mixture according to a number of factors, one of which isich ismolecular weight. Suppose you carry out a GC analysis of a bottlmolecular weight. Suppose you carry out a GC analysis of a bottle that ise that is
not labeled. Using a small volume syringe you withdraw some sampnot labeled. Using a small volume syringe you withdraw some samplele
from the bottle and inject it into the GC. Below is a hypotheticfrom the bottle and inject it into the GC. Below is a hypotheticalal
chromatogram of the analysis. Answer the questions below.chromatogram of the analysis. Answer the questions below.
Detector
Response
1)1) Which peak represents the compound that travels through the GCWhich peak represents the compound that travels through the GC
instrument in the least amount of time?instrument in the least amount of time?
2)2) Arrange the five peaks with respect to their relative molecularArrange the five peaks with respect to their relative molecular
weights, from heaviest to lightest.weights, from heaviest to lightest.
A B C DA B C D EE
___ > ___ > ___ > ___ > ______ > ___ > ___ > ___ > ___
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Postlab 2 Assessment Question:Postlab 2 Assessment Question:
Boiling Point & Molecular WeightBoiling Point & Molecular Weight
A mixture of three liquid hydrocarbons was injected into a GCA mixture of three liquid hydrocarbons was injected into a GC
at Tat Tovenoven = 95= 95OOC (isothermal). The resulting chromatogram isC (isothermal). The resulting chromatogram is
below.below.
1) Which compound is the most volatile?1) Which compound is the most volatile?
2) Which compound most likely has the smallest molecular2) Which compound most likely has the smallest molecular
weight?weight?
3) Which compound most likely has the highest boiling point?3) Which compound most likely has the highest boiling point?
4) Which compound spends more time in the liquid phase relative4) Which compound spends more time in the liquid phase relative
to the other two?to the other two?
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4.30 4.40 4.50 4.60 4.70 4.80 4.90 5.00 5.10 5.20 5.30 5.40 5.50 5.60 5.70 5.80 5.90
AA BB CC
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Postlab 2 Assessment Question: Boiling PointPostlab 2 Assessment Question: Boiling Point
GC physically separates liquids based on their differences, amonGC physically separates liquids based on their differences, among otherg other
factors, in boiling point (BP) temperature. Suppose there are thfactors, in boiling point (BP) temperature. Suppose there are threereeliquidsliquids-- A, B, and CA, B, and C-- with different boiling points: B > A > C. The threewith different boiling points: B > A > C. The three
component mixture is injected into a GC instrument whose ovencomponent mixture is injected into a GC instrument whose oven
temperature is held constant,temperature is held constant, i.e.i.e. isothermal. Which chromatogramisothermal. Which chromatogram
below (choices 1, 2, 3 or 4) best represents the order in whichbelow (choices 1, 2, 3 or 4) best represents the order in which the threethe threeliquids exit from the GC?liquids exit from the GC?
1)
2)
= A= A = B= B = C= C
3)
4)
start Time (arbritary units)
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Prelab 3: Oven Temperature, TPrelab 3: Oven Temperature, Tovenoven
Three separate 0.2Three separate 0.2L injections of an unknown colorless liquidL injections of an unknown colorless liquidwere done at three different oven temperatures: 125were done at three different oven temperatures: 125OOC,C,
100100OOC, and 75C, and 75OOC. The three separate chromatograms areC. The three separate chromatograms are
shown below. Match each chromatogram with one of the threeshown below. Match each chromatogram with one of the threeoven temperatures.oven temperatures.
start Time (arbitrary units)
TTovenoven = _______= _______OOCC
Toven = _______O
C
TTovenoven = _______= _______OOCC
l b 3 A Q i
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Postlab 3 Assessment Question:Postlab 3 Assessment Question:
TTovenoven & Molecular Weight& Molecular Weight
Into a test tube are placed four colorless, liquid compounds labInto a test tube are placed four colorless, liquid compounds labeled Aeled A
through D. Their relative molecular weights are:through D. Their relative molecular weights are: BB >> DD >> CC >> AA. On. On
the chromatograms below arrange the four compounds with respectthe chromatograms below arrange the four compounds with respect
to how they elute from the column at Tto how they elute from the column at Tovenoven == 115115
OO
CC andand 150150
OO
CC. All. Allother GC instrument conditions are the same for each analysis.other GC instrument conditions are the same for each analysis.
start Time (arbitrary units)
= A = B = C = D
start Time (arbitrary units)
TTovenoven = 115= 115OOCC
TTovenoven == 150150OOCC
l b
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Postlab 3 Assessment Question: TPostlab 3 Assessment Question: Tovenoven
As a new Environmental Science hire youAs a new Environmental Science hire youre given anre given anassignment involving GC. Your boss doesnassignment involving GC. Your boss doesnt have a cluet have a clue
about GC so gives you the two chromatograms. Bothabout GC so gives you the two chromatograms. Both
represent the same sample. The chromatograms wererepresent the same sample. The chromatograms weresubmitted to the Arizona Department of Environmentalsubmitted to the Arizona Department of Environmental
Quality (ADEQ) but returned because of the absence ofQuality (ADEQ) but returned because of the absence of
the GC operating conditions, namely Tthe GC operating conditions, namely T
ovenoven. Rather than. Rather than
doing the analysis again, you use your newly acquireddoing the analysis again, you use your newly acquired
knowledge of GC to argue that the two chromatogramsknowledge of GC to argue that the two chromatograms
were done at different Twere done at different Tovenoven. Which chromatogram. Which chromatogram-- A orA or
BB-- represents the analysis carried out at the higher ovenrepresents the analysis carried out at the higher oven
temperature? On the next page provide two reasons totemperature? On the next page provide two reasons to
ADEQ that support your conclusion.ADEQ that support your conclusion.
140
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Postlab 3 Assessment Question: TPostlab 3 Assessment Question: Tovenoven
As a new hire at a prestigious environmental consultingAs a new hire at a prestigious environmental consultingfirm, you are asked to conduct a GC analysis of a bottlefirm, you are asked to conduct a GC analysis of a bottle--
containing a colorless, homogeneous liquidcontaining a colorless, homogeneous liquid-- from one offrom one of
your firmyour firms clients. The bottles clients. The bottles label reads onlys label reads onlyAlcoholsAlcohols. Undaunted from such lack of information,. Undaunted from such lack of information,
you inject a small volume of the bottleyou inject a small volume of the bottles liquid contents liquid content
into a GC at Tinto a GC at Tovenoven
= 120= 120OOC (isothermal). The resultingC (isothermal). The resulting
chromatogram shows three peaks. Your boss ischromatogram shows three peaks. Your boss is
incompetent when it comes to GC so asks how you areincompetent when it comes to GC so asks how you are
certain that the bottle does not contain instead four orcertain that the bottle does not contain instead four or
five compounds, presumably all alcohols. What do you dofive compounds, presumably all alcohols. What do you do
to answer your bossto answer your boss question?question? Offer a brief but thoroughOffer a brief but thorough
explanation.explanation.
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Prelab 4: Injector Temperature, TPrelab 4: Injector Temperature, Tinjinj
22--PentanolPentanols boiling temperature is 120s boiling temperature is 120OOC. The redC. The redtriangle below represents 2triangle below represents 2--pentanol when Tpentanol when Tinjinj = 140= 140
OOC.C.
Suppose the same volume of 2Suppose the same volume of 2--pentanol was injected intopentanol was injected into
the GC but with Tthe GC but with Tinjinj = 100= 100OO
C. Using the bottom blank axis,C. Using the bottom blank axis,draw the chromatogram for 2draw the chromatogram for 2--pentanol when Tpentanol when Tinjinj = 100= 100
OOC.C.
Use a black triangle to represent 2Use a black triangle to represent 2--pentanol.pentanol.
start Time (arbitrary units)
start Time (arbitrary units)
TTinjinj = 140= 140OOCC
TTinjinj = 100= 100OOCC
P tl b 4 A t Q ti TP tl b 4 A t Q ti T
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Postlab 4 Assessment Question: TPostlab 4 Assessment Question: Tinjinj
Two separate 0.2L injections of para-xylene (bp = 135-137OC)
were done under identical isothermal oven temperature, carrier gasflow rate, and split ratio conditions but at different injector
temperatures, Tinj: 100OC and 170OC. Three chromatograms are
shown below but only two are realistic. Match a chromatogram
with either Tinj. Which chromatogram represents Tinj 100OC and
which Tinj 170OC? Give a reason for not choosing the remaining
chromatogram.
A.
start Time (arbitrary units)
Tinj = ________OC
B.
C.
Tinj = ________OC
Tinj = ________O
C
P tl b 4 A t Q ti TPostlab 4 Assessment Question T
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Postlab 4 Assessment Question: TPostlab 4 Assessment Question: Tinjinj
Below are a series of chromatograms of 2Below are a series of chromatograms of 2--octanol taken inoctanol taken in
order from top to bottom. Offer an explanation for whyorder from top to bottom. Offer an explanation for whythe chromatograms differ.the chromatograms differ.
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2.0 2.1 2.2 2.3 2.4 2.5 2.6 2.7 2.8 2.9 3.0 3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8 3.9 4.0 4.1 4.2 4.3 4.4
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Student CommentsStudent Comments
Most of the time all we learn in school is very theoretical.Most of the time all we learn in school is very theoretical.We donWe dont apply it or use equipment. It apply it or use equipment. Ive always wanted tove always wanted toat least handle some lab equipment in my life before itat least handle some lab equipment in my life before itbecomes a necessity.becomes a necessity.
Being able to use instrumentation definitely demystifiesBeing able to use instrumentation definitely demystifiescertain elements of science. Itcertain elements of science. Its easier for me to assimilates easier for me to assimilate
knowledge through doing things Iknowledge through doing things I
m learning about.m learning about.
Wanted to let you know that I have seriously consideredWanted to let you know that I have seriously consideredgoing into research. A big part of this was mygoing into research. A big part of this was myintroduction to the use of instrumentation in your class.introduction to the use of instrumentation in your class.
I enjoyed using the expensive machinery [in order] toI enjoyed using the expensive machinery [in order] tounderstand what we were learning in class.understand what we were learning in class.
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AcknowledgementsAcknowledgements
1) National Science Foundation (NSF)1) National Science Foundation (NSF)
Course, Curriculum, &Course, Curriculum, &Laboratory ImprovementLaboratory Improvement
(CCLI) grant (DUE(CCLI) grant (DUE-- 0310264)0310264)
2) All my students2) All my students