Lab Report Beers Law

8/16/2019 Lab Report Beers Law

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Chemical And Biomolecular EngineeringDepartment

Lafayette College

TITLE: Absorbance

SUBMITTED BY: David Angel (A)

LAB A!T"E!S: Brandy West (B), Gabby Montes (C)

DATE #$ LAB %#!&:

DATE DUE: October 17, 201 DATESUBMITTED: October 17, 201

C#U!SE: C!" #12$ "%&eri'ental Design

SECTI#": #

I"ST!UCT#!: oe Woo

ABST!ACT :

*+e ollo-ing re&ort details t+e e%&eri'ental data o t+e relations+i&bet-een absorbance and concentration or t+ree di.erent co''erciallyi'&ortant dyes$ /D C ed 30, D C Bl4e 1, and /D C 5ello- , anddi.erent 'i%t4re o t+e'6 t -as concl4ded t+at t+e relations+i& bet-eent+e variables o interest agreed -it+ t+e 'at+e'atical relations+i& no-n asBeer8s la- only or &4re s&ecies sol4tions6 Mi%t4res o t+e s4bstances didnot agree -it+ said relations+i&6


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A'(tract

*+e ollo-ing re&ort details t+e e%&eri'ental data o t+e relations+i&

bet-een absorbance and concentration or t+ree di.erent co''erciallyi'&ortant dyes$ /D C ed 30, D C Bl4e 1, and /D C 5ello- , anddi.erent 'i%t4re o t+e'6 t -as concl4ded t+at t+e relations+i& bet-een t+evariables o interest agreed -it+ t+e 'at+e'atical relations+i& no-n asBeer8s la- only or &4re s&ecies sol4tions6 Mi%t4res o t+e s4bstances did notagree -it+ said relations+i&6

Introduction

*+e relations+i& bet-een t+e absor&tion o radiant energy by an o&a94e'edi4', 'ore s&eci:cally a li94id sol4tion o a solid, and t+e concentrationo t+e solid in sol4tion -as :rst st4died by t+e Ger'an 'at+e'atician andc+e'ist A4g4st Beer6 n 1; 2 +e &ro&osed t+e 'at+e'atical relations+i&

no-n as Beer8s la- -+ic+ states t+at$


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/or a sol4tion o '4lti&le co'&onents, "96161 e%tends to$

A λi= ∑ ε n , λ i∗ L∗c n ("96 162)

n ot+er -ords, t+e absorbance at a given -avelengt+ o t+e sol4tion -ill begiven by t+e s4''ation o t+e absorbance o t+e individ4al co'&onents ot+e sol4tion6 /or a given 'i%t4re i t+e e%tinction coe cient is no-n oreac+ co'&onent at di.erent -avelengt+s t+en t+eoretically t+econcentrations o eac+ co'&onent can be o4nd by a&&lying t+e Beer8s la-relations+i& at a 'ini'4' o n -avelengt+s, -+ere n is t+e n4'ber oco'&onents in t+e 'i%t4re6 And t+e -avelengt+s are t+e +ig+est absorbance-avelengt+ or eac+ dye6 By &ic ing t+e +ig+est absorbance -avelengt+, t+e+ig+est signal to noise ratio is g4aranteed6

*+is e%&eri'ent +as t+ree 'ain obEectives$

16 *o :nd t+e -avelengt+ o 'a%i'4' absorbance or t+ree 'aEorco''ercially i'&ortant ood dyes and co'&are t+e e%&eri'entalval4e to t+e val4e re&orted in t+e literat4re6

26 *o :nd, by 'eans o a regression, t+e relations+i& bet-een t+eabsorbance o radiant energy and t+e concentration o sol4tions o&4re ood dyes at eac+ o t+e -avelengt+s o 'a%i'4' absorbanceand e%&lore -+et+er Beer8s la- is a&&ro&riately i'&le'ented ordyes, and i so to deter'ine t+e e%tinction coe cient o eac+ o t+edyes at eac+ o t+e -avelengt+s to 4rt+er e%&lore t+e a&&lication oBeer8s la- or 'i%t4res o dyes6

#6 *o e%&lore -+et+er Beer8s la- is an a&&ro&riate 'at+e'aticale%&ression or t+e relations+i& bet-een t+e absorbance o t+e '4ltico'&onent sol4tion at di.erent -avelengt+s and t+e concentration oeac+ co'&onent6

E)perimental Method(

!eagent(:

*+ree co''ercially i'&ortant ood dyes -+ere 4sed in t+is e%&eri'ent to+el& 'eet t+e obEectives o4tlined &revio4sly in t+e introd4ction section6 *+edyes 4sed -ere$ t+e /D C ed 30, /D C Bl4e 1, and /D C 5ello- 6All t+e dyes are ood grade and 'an4 act4red by O!A6

Apparatu(:


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*+e Genesis 2 F&ectro&+oto'eter -as 4sed to 'eas4re absorbance osa'&les at di.erent -avelengt+s6 *+e 'ac+ine +ad to be t4rned on #0'in4tes &revio4s to any 'eas4re'ent beca4se t+e b4lbs need ti'e to -ar'4&

$igure * 6 *+e ront vie- o t+e Genesis 2 s&ectro&+oto'eter6 2

$igure + 6 *+e

&rocess diagra'o t+e

'eas4re'ent &rocess inside o t+e Genesis 2 s&ectro&+oto'eter6 #

2+tt&$ ---6 ran s+os&ital-or s+o&6co' e94i&'ent doc4'ents &+oto'eter 4ser 'an4als F&ectronicH20GenesysH20F&ectro'eterH20 H20 serH20'an4al6&d

# "t6 Al


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/ig4re 2 s+o-s t+e s&eci:c &rocess by -+ic+ t+e Genesis 2s&ectro&+oto'eter o&erates$ *-o la'&s &rod4ce a bea' o lig+t t+atcontains all -avelengt+s6 *+e bea' t+en &asses t+ro4g+ a narro- slit andonly a -avelengt+ o lig+t is selectively let t+ro4g+ to &ass t+ro4g+ t+esa'&le6 *+e re erence detector 'eas4res t+e intensity o t+e bea' o lig+tbe ore it &asses t+ro4g+ t+e sa'&le6 *+e detector t+en 'eas4res t+eintensity o t+e inco'ing lig+t6 Bot+ o t+e intensity 'eas4re'ents areco'&ared to deter'ine t+e absorbance o lig+t6

rocedure:

*+e :rst ste& in t+e e%&eri'ental &roced4re is to &re&are a n4'ber osa'&les o eac+ dye and 'i%t4re o dyes6 *+e :rst ste& in t+is &rocess -asto &re&are a large a'o4nt o a stoc sol4tion -it+ a concentration near0602g I o eac+ dye6 One serial dil4tion (re er to &rocess or serial dil4tion int+e a&&endi%) -as t+en &er or'ed o eac+ o t+e stoc sol4tions by eac+ ot+e 'e'bers o t+e tea'6 *+e concentration o eac+ o t+e res4ltingsa'&les are described in *able 16

Ta'le * 6

Concentration o sa'&lesed Dye Bl4e Dye 5ello- Dye

Fa'&le g I Fa'&le g I Fa'&le g IF1 (stoc ) 060217 F1 (stoc ) 060222 F1 (stoc ) 060221

F2 06010J F2 060111 F2 060111F# 0600 3 F# 0600 K F# 0600F3 060027 F3 06002; F3 06002;F 060013 F 060013 F 060013FK 060007 FK 060007 FK 060007F7 06000# F7 06000# F7 06000#

*+e ne%t ste& in t+e e%&eri'ental &roced4re -as to deter'ine t+e-avelengt+ o 'a%i'4' absorbance or eac+ dye, by testing a stocsol4tion6 *+is -as ac+ieved by 'an4ally c+anging t+e -avelengt+ o'eas4re'ent, and recording t+e absorbance at eac+ o t+e -avelengt+s,-+ile t+e stoc sa'&le -as in t+e c+a'ber o t+e s&ectro&+oto'eter6

Data o absorbance -as t+en collected at a n4'ber o concentrations oreac+ dye in order to e%&lore t+e relations+i& bet-een t+e absorbance ateac+ o t+e t+ree -avelengt+s deter'ined &revio4sly and t+e concentrationo a sa'&le o &4re dye6


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A n4'ber o sa'&les -ere t+en created (re er to &roced4re to &rod4cesa'&les o 'i%t4res in t+e a&&endi%) o 'i%t4re o dyes at di.erent no-nconcentrations and t+en collect data o t+e absorbance at t+e t+ree 'aEor-avelengt+s6 *+e concentrations o t+e sa'&les tested can be o4nd in t+ea&&endi%6

Safety:

/D C ed 30 n general +as less +ealt+ concerns t+an ot+er aLo dyes6 *+ere +ave been clai's t+at ingestion o t+e dye co4ld be related to AD!Db4t evidence is not s4bstantial6

/D C Bl4e 1 -as +andled -it+ care6 t +as t+e ca&acity to ind4ce allergicreactions in individ4als -it+ ast+'a6

/D C 5ello- -as +andled -it+ care6 At least 0601H o t+e &o&4lation issensitive to contact -it+ t+e dye6 artic4larly individ4als -it+ ast+'a andas&irin intolerance6

*+e general sa ety considerations -ere ollo-ed as -ell$ -+en +andling t+e&o-dered or' o t+e dyes gloves '4st be -orn to avoid discovering allergicreactions in gro4& 'e'bers6 *+e &o-dered or' o t+e dyes '4st not becons4'ed or in+aled6 in contact -it+ t+e dye gro4& 'e'bers '4st rinset+e a.ected area -it+ -ater to avoid any &ossible co'&lications6

!e(ult( and Di(cu((ion

*+e res4lts o t+e 'eas4re'ent o t+e absorbance o t+e t+ree di.erent dyesat di.erent -avelengt+s is s+o-n belo-6 *+e stoc sol4tions -ere 4sed or'eas4re'ent6

a)


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b)

c)

$igure , 6 Fcatter &lots o absorbance o eac+ dye at di.erent -avelengt+s6

/ig4re # s+o-s gra&+ically t+e de&endence o absorbance and -avelengt+ o lig+t 'eas4red or t+e stoc sol4tions o t+e t+ree di.erent dyes6 n general,t+e e%&eri'entally deter'ined 'a%i'4' absorbance o eac+ o t+e dyes-as not too di.erent ro' t+e val4e re&orted in t+e literat4re6 *able 2 s+o-st+e divergence o bot+ val4es6

Ta'le +-

Dye Wavelengt+(n')

e&ortedWavelengt+

(n')ed 3J 03Bl4e K#0 K2;

5ello- 3#0 32

Wit+ t+e e%&eri'entally deter'ined 'a%i'4' absorbance -avelengt+ datare&orted in *able 2, e%&eri'ental data o t+e absorbance o eac+ o t+e dyes


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at eac+ o t+e -avelengt+s or di.erent concentrations -as collected6 *ables# and 3 &resent t+e res4lts o a linear regression bet-een absorbance andconcentration or eac+ o t+e dyes at eac+ o t+e -avelengt+s -it+ t+eres&ective J H con:dence intervals6 *+e ra- 'eas4re'ent data and &lotso t+e regressions can be o4nd in t+e a&&endi%6

ed Dye Lg∗cm Bl4e Dye Lg∗cm 5ello- Dye Lg∗cm

3#0n'

K6721 (K6KK3#7,K6777##)

162K (1622;77,162J170)

226#K (226130#,226 737)

3Jn'

2163J (2162720,216713J)

06720J (06KJ3012,06737;20)

16J K (16;;2K2,2602J7#)

K#0n'

0612#J (060;21#11,061K KKJ)

3 6J2 (3 6###;,3K6 0 J)

060 JJ; (0600 ##1#,0612 2; )

Ta'le ,

ed Dye Bl4e Dye 5ello- Dye3#0 n' 060 ; (060 7J;,

060 J0 )

060 ;7 (060 ;3#,

060 J03)

060K10 (060 ;J,

060K#1)

3J n' 060 7; (060 7,060 JJ)

060 K# (060 K107,060 KK2;)

060 70 (060 K27,060 7KJ)

K#0 n' 060 3# (060 #;;JJ, 060 3K;1)

060 K (060 0J,060K22

060 2 (060 3K1#3, 060 ;7#)


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Ta'le .

*able # re&orts t+e slo&e o t+e linear regression -+ile *able 3 re&orts t+einterce&t -it+ t+e 5 a%is6 Nis4ally t+e &lots o t+e linear regressions incl4dedin t+e a&&endi% s+o- t+at t+e linear regression is a airly acc4rate 'odel ort+e relations+i& bet-een absorbance and concentration o t+e dyes6 n act,t+e :t is so good t+at a resid4al analysis -as not even necessary6 !o-everGiven that the samples measure where prepared by three different members, an analysis ofvariation can be performed to evaluate differences in the data with respect to the person that

prepared it and to obviously the concentration. In the appendix box diagrams are reported thatvisually show the small variances on the data. It can be seen that member C usually had higherabsorbance reading, however the deviations were really small.

A 2 -ay A ONA -as atte'&ted, +o-ever beca4se o t+e inde&endencebet-een t+e 'eas4re'ents and t+e individ4al t+at &er or'ed t+e dil4tions,it did not give satis actory res4lts6 /or t+is reason, t-o 1 -ay A ONA -ere&er or'ed instead6 *+e res4lt o t+is calc4lations can be o4nd in t+ea&&endi%6

After a close look of the AN !A data we can conclude that the difference in the measurement between group members was not statistically significant "#$%&&'.'(), this is seen because theconfidence intervals overlap a considerable amount. *he one way AN !A between absorbanceand sample number shows that, to no surprise, the absorbance is highly dependent on the sampleconcentration "sample number).

By e%a'ining t+e res4lting 'at+e'atical e%&ression or t+e linearregressions -e can observe t+at -it+ t+e e%ce&tion o +aving a s'all yinterce&t t+ey agree -it+ Beer8s la- -it+ t+e slo&e corres&onding to t+ee%tinction coe cient6 By e%a'ining t+e y interce&t8s 'agnit4de or eac+ ot+e regressions -e can observe t+at t+ey are relatively constant, -+ic+indicates t+at t+e interce&t is t+e res4lt o a syste'atic error in t+ee94i&'ent6 *+ere ore, or t+e a&&lication o Beer8s la- to 'i%t4res o t+edyes, t+e slo&e o t+e regressions -ere 4sed as t+e e%tinction coe cient oreac+ dye at eac+ -avelengt+ and t+e interce&t -as ignored6

It is important to note that the +( CI of the extinction coefficient of the yellow dye at - 'nmhas ' in it. /hich means that for yellow dye at - ' nm concentration does not influence theabsorbance.


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/ive 'i%t4res o no-n concentration -ere created by eac+ 'e'ber and t+eabsorbance at t+e t+ree 'aEor -avelengt+s -ere 'eas4red6 According toBeer8s la- or 'i%t4res described in "96162$

A λ430= ∑ ε n , λ 430∗ L∗c n

A λ495 = ∑ εn , λ 495∗ L∗c n

A λ630= ∑ εn , λ 630∗ L∗c n

*+e e%&ression t+en ta es into acco4nt t+e absorbance o t+ree co'&onentsat t+ree di.erent -avelengt+s6 *+e res4lting set o t+ree e94ations +as t+ree4n no-ns (concentrations o eac+ co'&onent) t+at can be solved or6 *+e

data or absorbance and a sa'&le o t+e calc4lations o t+e &redictedconcentrations can be o4nd in t+e A&&endi%6 *able s+o-s t+e co'&arisonbet-een t+e act4al concentration or eac+ dye in eac+ 'i%t4re and t+ee%&eri'entally deter'ined concentration$

Ta'le /

Concentrationsg / L

"%&eri'ental

Beersla-

Herror

ed and Bl4eP6,

ed 060013 0600#7 1K36#Bl4e 060013 06002K ; 67

5ello- 0 06001; A

ed and 5ello-

ed 060013 0600#K 1 761


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Bl4e 0 060012 A 5ello- 060013 060033

2136#

5ello- and Bl4eed 0 06002 A

Bl4e 060013 06002K ; 67 5ello- 060013 0600##

1# 67

ed, Bl4e and 5ello- (sa'econcentration)

ed 06001; 060032 1##6#Bl4e 06001J 0600# 76J

5ello- 06001; 06003#

1#;6J

ed, Bl4e and 5ello- (strongGreen)

ed 06001 06003# ##060Bl4e 0600 0600 J 1;60

5ello- 0600 0600K #060

By loo ing at t+e H errors or eac+ o t+e dye concentrations re&orted in *able -e can see t+at Beers la- does not a&&ly to t+e 'i%t4res o dyes as

it a&&lies to t+e &4re dyes6 *+e concentrations obtained by t+e calc4lation o"96162 res4lted in val4es or non e%istent dyes in t+e 'i%t4re6 n ot+er -ords,t+e dyes &resent in t+e 'i%t4res generated t+e ill4sion o a t+ird dye6 t canalso be seen t+at t+e +ig+er t+e concentrations o t+e dyes t+e 'oreacc4rate t+e &rediction o Beer8s la-, t+is is a&&arent ro' t+e res4lts o t+estrong green 'i%t4re6

Green is a &artic4larly co''ercially i'&ortant color6 As can be seen ro' t+eres4lts in *able , a desired +4e o green, -+ic+ -o4ld t+eoretically need to+ave a +ig+ concentration o red dye, can be obtained si'&ly by co'bining

+ig+ concentrations o bl4e and yello- dye6 *+is no-ledge is &otentiallyi'&ortant or t+e ind4stry beca4se it i'&lies t+at t+ere is no need to -astered dye in t+e &rod4ction o t+e desired color6

Source( of error:


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*+e develo&'ent o Beer8s la- +as been incredibly 4se 4l or ind4striala&&lications in &art beca4se o its si'&le linear relations+i& bet-eenabsorbance and concentration6 !o-ever, alt+o4g+ si'&le, t+e 'eas4re oabsorbance is act4ally develo&ed ro' a 'ore co'&le% logarit+'icrelations+i&$ trans'ittance6 n ot+er -ords$

Absorbance = 2− log 10 (Transmittance )

Given t+is logarit+'ic relations+i&, t+e linearity o absorbance brea s do-nat +ig+ concentrations (lo- trans'ittances)6 *+eoretically 360 A t+e linearrelations+i& does not +old any longer 36 Fa'&les o +ig+ concentrations aret+ere ore a so4rce o error6

/or lo- concentration sa'&les t+e so4rces o error incl4de$ lig+t re raction,reQection, and scattering6 All o t+e &revio4s e.ects -ill +ave t+e e.ect oca4sing e%tra absorbance t+at is not ca4sed by t+e sa'&le itsel 6

E)perimental impro0ement(:

ossible e%&eri'ental i'&rove'ents -o4ld rely on red4cing t+e so4rces oerrors6 *+e :rst i'&rove'ent -o4ld be to ens4re t+at t+ere -ere no dye big&articles t+at co4ld scatter lig+t6 *+is co4ld be ac+ieved by 'i%ing t+esol4tions t+oro4g+ly and even 4sing :ner dye &o-der6 Big &articles insol4tion ca4se scattering o lig+t -+ic+ decreases t+e acc4racy o t+e'eas4re'ents6 By 'a ing s4re t+e dyes are t+oro4g+ly dissolved -it+ no&articles in sol4tion t+e scattering -ill be avoided6 *+e second &ossiblei'&rove'ent is to be certain t+ere is no cross conta'ination bet-eensa'&les6 *+is co4ld be ac+ieved by 4sing di.erent &i&ets every ti'e6Anot+er &ossible i'&rove'ent -o4ld be to 4se &er ectly clean and Qa-lessc4vettes in t+e 'eas4re'ent to avoid scattering o lig+t d4e to resid4e in-alls o t+e c4vette and t+e i'&er ections o t+e &lastic6

Conclu(ion(

Based on t+e e%&eri'ental data obtained and &resented in t+e &revio4ssection, -e can 'a e a variety o concl4sions6 *+e :rst concl4sion +as to do-it+ t+e -avelengt+ or 'a%i'4' absorbance o eac+ dye6 t is clear ro'loo ing at /ig4re # t+at t+e -avelengt+ or 'a%i'4' absorbance or eac+dye di.ers ro' t+e -avelengt+ re&orted in t+e literat4re6 *+e e%act reason

3 +tt&$ ---6t4rnerdesigns6co' t2 doc a&&notes F 007 6&d


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t+is is t+e case 'ay +ave to do -it+ t+e &4rity o t+e dye, &ossible4nacco4nted conta'inants, or an error in t+e 4nction o t+e s&ectro'eter6

*+e second concl4sion +as to do -it+ t+e a&&lication o Beer8s la- to &4redyes6 /ro' :g4res -e can observe t+at t+e linear :t is 'ore t+an

acc4rate or t+e dyes tested6 *+e relations+i& o4nd by t+e regressionanalysis see's to be very &recise, given t+at t+e J H C are so s'all6 *+e

act t+at all o t+e dyes syste'atically e%+ibited a si'ilar y interce&t&robably +as to do -it+ a syste'atic 'eas4re'ent ail4re in t+es&ectro&+oto'eter6 n general Beer8s la- see's to be a good :t or t+e data6

*+e t+ird concl4sion +as to do -it+ t+e absorbance o '4lti co'&onentsol4tions o dyes6 By loo ing at *ables -e can see t+at t+e Beer8s la-&rediction or t+e '4lti co'&onent sol4tion +ad a signi:cant error ist+ere ore not a 4se 4l 'at+e'atical relations+i& to deter'ine concentrationso 'i%t4res o dyes6 *+e reason be+ind t+e ina&&licability o Beer8s la-&robably +as to do -it+ synergistic interactions bet-een t+e dyes t+atca4sed t+e absor&tions to be +ig+er t+an t+ey -o4ld +ave been -it+o4t saidinteractions6 Ioo ing at *able (strong green) or instance it can be seent+at t+e &redictions or t+e concentration or bl4e and yello- did not +ave asbig an error as ot+er sa'&les6 !o-ever t+e &rediction or t+e concentrationo t+e red dye +ad a ##0H error -+ic+ s4ggests t+at t+e +ig+ concentrationso yello- and bl4e interacted to &rod4ce t+e e.ect o t+e red dye being&resent6

Literature Cited

*4rner Deigns8 triology laboratory Q4ori'eter a&&lication note onabsorbance$ +tt&$ ---6t4rnerdesigns6co' t2 doc a&&notes F 007 6&d

Milton oy8s Genesys 2 s&ectro&+oto'eter o&erator8s 'an4al$+tt&$ ---6 ran s+os&ital-or s+o&6co' e94i&'ent doc4'ents &+oto'eter 4ser 'an4als F&ectronicH20GenesysH20F&ectro'eterH20 H20 ser

H20'an4al6&d "ncyclo&edia Britannica8s entry on Beer8s la-$ +tt&$ 0---6britannica6co'6libcat6la ayette6ed4 science Beers la-

Appendi)

http://www.turnerdesigns.com/t2/doc/appnotes/S-0075.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.frankshospitalworkshop.com/equipment/documents/photometer/user_manuals/Spectronic%20Genesys%20Spectrometer%20-%20User%20manual.pdfhttp://www.turnerdesigns.com/t2/doc/appnotes/S-0075.pdf


14/28

rocedure to prepare a (erial dilution (et:

Ferial dil4tions o t+e stoc sol4tion '4st be &re&ared by ollo-ing t+e ne%tste&s$

16 'i% 1 'l o stoc sol4tion -it+ 1 'l o D -ater to create F2626 'i% 1 'l o t+e F2 -it+ 1 'l o D -ater to create F#6#6 e&eat 1 2 4ntil F7 is obtained6

roducing (ample( of mi)ture(:

/or 'i%t4res o t-o dyes$ co'bine F3 o one o t+e &4re dye sa'&les to t+eF3 o anot+er o t+e dyes6

/or 'i%t4re o t+ree dyes$ co'bine F3 o t-o dyes t+en ta ing 1 'l o t+e'i%t4re and 'i%ing it -it+ F o t+e t+ird dyes

Concentration de&endence o absorbance o ed Dye at 3#0n'$

06020060106010060006000

0622

06200

0617

061 0

0612

06100

0607

060 0

Concentration( 1g2 L3

A '

( o r ' a n c e

F 06000;J03DF9 10060HD F9( adE ) 10060H

egressionJ H C

!ed Dye Cali'ration lot at .,4 nmAbsorbance ? 060 ; 1 RK6721 Concentrations (g I)

Concentration de&endence o absorbance o ed Dye at 3J n'$


15/28

06020060106010060006000

06

063

06#

062

061

060


A ' ( o r ' a n c e

F 0600#3J0JDF9 10060HD F9( adE ) 10060H

egressionJ H C

!ed Dye Cali'ration lot at .5/ nmAbsorbance ? 060 7;# R2163J Concentrations (g I)

Concentration de&endence o absorbance o ed Dye at K#0n'$

06020060106010060006000

060 ;

060 7

060 K

060

060 3


A ' ( o r ' a n c e

F 06000K ;DF9 K760HDF9(adE) K 62H

egressionJ H C

!ed Dye Cali'ration lot at 6,4 nmAbsorbance ? 060 32J R0612#J Concentrations (g I)


16/28

Concentration de&endence o absorbance o Bl4e Dye at 3#0n'$

060206020060106010060006000

060J0

060;

060;0

0607

06070

060K

060K0


A ' ( o r ' a n c e

F 06000 073DF9 JJ67HDF9(adE) JJ67H

egressionJ H C

Blue Dye Cali'ration lot at .,4nmAbsorbance ? 060 ;73 R162K0 Concentrations (g I)

Concentration de&endence o absorbance o Bl4e Dye at 3J n'$

060206020060106010060006000

0607

06070

060K

060K0

060


A ' ( o r ' a n c e

F 060003##JDF9 JJ63HDF9(adE) JJ63H

egressionJ H C

Blue Dye Cali'ration lot at .5/ nmAbsorbance ? 060 K#7 R06720J Concentrations (g I)


17/28

Concentration de&endence o absorbance o Bl4e Dye at K#0n'$

060206020060106010060006000

162

160

06;

06K

063

062

060


A ' ( o r ' a n c e

F 0600J3 1KDF9 JJ6JHDF9(adE) JJ6JH

egressionJ H C

Blue Dye Cali'ration lot at 6,4 nmAbsorbance ? 060 K 3 R3 6J2 Concentrations (g I)

Concentration de&endence o absorbance o 5ello- Dye at 3#0n'$

060206020060106010060006000

06K

06

063

06#

062

061

060


A '

( o r ' a n c e

F 0600#3;70DF9 10060HD F9( adE ) 1006 0H

egressionJ H C

Yello7 Dye Cali'ration lot at .,4 nmAbsorbance ? 060K0J; R226#K Concentrations (g I)

Concentration de&endence o absorbance o 5ello- Dye at 3J n'$


18/28

060206020060106010060006000

0610

060J

060;

0607

060K


A ' ( o r ' a n c e

F 060011;10DF9 JJ63HDF9(adE) JJ63H

egressionJ H C

Yello7 Dye Cali'ration lot at .5/ nmAbsorbance ? 060 KJ; R16J K Concentrations (g I)

Concentration de&endence o absorbance o 5ello- Dye at K#0n'$

060206020060106010060006000

060 ;

060 7

060 K

060

060 3


A ' ( o r ' a n c e

F 0600103;KDF9 1K6#HDF9(adE) 116JH

egressionJ H C

Yello7 Dye Cali'ration lot at 6,4 nmAbsorbance ? 060 23 R060 JJ; Concentrations (g I)

!a7 E)perimental Data:

Ma%i'4' Absorbanceed Bl4e yello-

-avelengt+

absorbance

-avelengt+

absorbance

-avelengt+

absorbance

# 0 0603J # 0 0602 # 0 0611J#7 0613J #7 06117 #7 06##3300 0611 300 060;K 300 0630332 06131 32 0603 30 0631J3 0 061;K 3 0 0600# 310 063#137 06# ; 37 0602# 31 06 #23;0 06#73 00 0601K 320 0637


19/28

3; 0632; 2 060#; 32 063;K3J0 063JJ 0 060K 3#0 06 333J 06 1 7 06#1 3# 063J#

00 063KK ;0 06#31 330 063 ;03 063K7 ; 06#13 33 063K10 063;7 J0 0631# 3 0 063110 063 1 J 0630J 37 0621J1 063 J K00 063 00 060220 06327 K0 06 3 2 02 06307 K10 06K;3 0 0

0 061;; K1 06;1; 7 07 060J# K20 06;72 K00 0

K00 06002 K2 06J72 2 0K2 0600# K2; 06JJ; K 0 0K 0 0 K#0 160 2 K7 0

K7 0 K# 16001 700 0700 0 K30 06; K 72 072 0 K3 06K; 7 0 07 0 0 K 0 06331 77 077 0 K7 0600J ;00 0;00 0 700 060 3

72 060 27 0 060 377 060 3;00 060 3

ed-avelgt+ 3#0 F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a

n c e Dil4tion A 06203 061#1 060J 0607K 060K7 060K# 060K1Dil4tion B 06203 061#2 060J 0607K 060K7 060K# 060K1

Dil4tion C 0620K 0612J 060JK 0607; 060K; 060K3 060K1

ed-avelgt+ 3J F1 F2 F# F3 F FK F7

concentration g I


20/28

a b s o r b a n c e

Dil4tion A06 1

J062J

K0617

70611

#060;

0607060K

#

Dil4tion B06 1

J062J

K0617

;0611

3060; 0607

1060K

3

Dil4tion C06 2

J062;

; 061;0611

J060;

;0607

2060K

ed-avelgt+ K#0 F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a n c e Dil4tion A

060;

060 0603

0603

0603

0603

0603

Dil4tion B060

;060 060 060 060 060 060

Dil4tion C060

K060 060 060 060 060 060

3

Bl4e-avelgt+ 3#0 F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a

n c e

Dil4tion A 060;7 067# 060K 060K2 060K 060K 060 JDil4tion B 060;7 0607# 060K 060K2 060K 060K 060KDil4tion C 060;K 0607# 060KK 060K# 060K1 060 J 060 J

Bl4e-avelgt+ 3J F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a n c e Dil4tion A 06072 060K3

060K 060 ; 060 7 060 7 060 7

Dil4tion B 06072 060K060

K 060 ; 060 7 060 7 060 7

Dil4tion C 0607# 060K060

K 060 J 060 7 060 7 060 7

Bl4e-avelgt+ K#0 F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a n

c e Dil4tion A 160 J 06 ;# 06#0J 061;1 06117 060;K

0607

Dil4tion B 160K 06 ;3 06#1 061;1 0611; 060;K 0607


21/28

Dil4tion C 160;# 06 ;K 06#17 061;# 0611; 060;060

7

5ello-

-avelgt+ 3#0 F1 F2 F# F3 F FK F7concentration g I

a b s o r b a n

c e

Dil4tion A 06 3J 06#12 061; 06122 060J1 0607 060K7Dil4tion B 06 06#12 061; 06122 060J1 0607 060KK

Dil4tion C 06 7 06#1K 061;J 06127 060J1 06073 060KK

5ello--avelgt+ 3J F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a n c e Dil4tion A

06101 060;

060K;

060K# 060K

060;

0607

Dil4tion B0610

10607 060K

;060K

#060K

1060

J060

7

Dil4tion C 0610607

7060K

;060K

3060

J060

;060

7

5ello--avelgt+ K#0 F1 F2 F# F3 F FK F7

concentration g I

a b s o r b a n c e Dil4tion A 060; 0607 060 060K 060K 060 0603

Dil4tion B060

7060 060 060

K060

K060

K060

3

Dil4tion C060 060 060 060

;060 060 060

ed and Bl4esa'&le A B C

concentrations

ed 060027 1'l06001

3

Bl4e 06002; 1'l06001

3

a b s o r b a

n c e s -avelegnt+

s

3#0 0607 060K; 06073J 060;7 060;1 060;JK#0 0611J 061# 06117


22/28

red and yello-sa'&le A B C

concentrations ed 060027 1'l

06001

3 5ello- 06002; 1'l

060013

a b s o r b a n c e s

-avelegnt+s

3#0 061060J

; 0617

3J 060;7060;

; 060;K

K#0 060 3060

3 060

yello- and bl4esa'&le A B C

concentrations

5ello- 06002; 1 'l06001

3

Bl4e 06002; 1'l06001

3

a b s o r b a n c e

s

-avelegnt+s

3#0 060J7060J

# 060J2

3J 060K1060K

1 060K1K#0 06112 0612 06121

red, bl4e andyello-

sa'&le A B C

c o n c e n t r a t i o n

s

red 0600 3 0606001

;

bl4e 0600 K 0606001

J

yello- 0600 0606001

;

a b s

o r b a

n c e s -avelegn

t+s

3#0 0611# 06113 061K3J 061 06102 0610#K#0 0613 06131 06131

red, bl4e and yello-sa'&le

A B C

c o n red 06010

J061 06001

0


23/28

c e n t r a t i o

n s bl4e

060111 06

06000

yello-06011

1 060600

0

a b s o r b a n

c e s

-avelegnt+s

3#0 061;061;

3 0617;

3J 061120610

7 0610J

K#0 062KK0627

7 0627K

A"#8A re(ult(:


24/28

One way ANOVA of absorbance and group member430 nm

Source DF SS MS F PPerson 2 0.0000 0.0000 0.00 0.999Error 60 0.8215 0.0137Total 62 0.8215

Indi idual 95! "Is For Mean #ased on Pooled StDe$e el % Mean StDe &&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&( 21 0.1221 0.1163 )&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&+# 21 0.1222 0.1165 )&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&+" 21 0.1233 0.1182 )&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&+ &&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&


25/28

0.090 0.120 0.150 0.180Pooled StDe , 0.1170

495 nm


S , 0.1101 -&S , 0.00! -&S )ad/+ , 0.00!

Indi idual 95! "Is For Mean #ased on Pooled StDe$e el % Mean StDe &&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&( 21 0.106 0.1097 )&&&&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&&&+# 21 0.1065 0.1097 )&&&&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&&&+" 21 0.1072 0.1109 )&&&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&&&+ &&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&& 0.075 0.100 0.125 0.150

Pooled StDe , 0.1101

630 nm


S , 0.2 -&S , 0.00! -&S )ad/+ , 0.00!

Indi idual 95! "Is For Mean #ased on Pooled StDe$e el % Mean StDe &&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&

( 21 0.151 0.2 26 )&&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&&+# 21 0.1517 0.2 28 )&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&&+" 21 0.1531 0.2 77 )&&&&&&&&&&&&&&&&&*&&&&&&&&&&&&&&&&+ &&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&& 0.060 0.120 0.180 0.2 0

Pooled StDe , 0.2


26/28


27/28

495 nm Source DF SS MS F PSa le 22 0.727383 0.0330629 8113.59 0.000Error 0 0.0001630 0.00000 1Total 62 0.7275 6

S , 0.002019 -&S , 99.98! -&S )ad/+ , 99.97!

Indi idual 95! "Is For Mean #ased on Pooled StDe$e el % Mean StDe &&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&#lue 1 2 0.07250 0.00071 *#lue 1 1 0.07200 * *#lue 2 3 0.06 67 0.00058 *+#lue 3 3 0.06000 0.00000 *#lue 3 0.05833 0.00058 *#lue 5 3 0.05700 0.00000 *#lue 6 3 0.05700 0.00000 *#lue 7 3 0.05700 0.00000 *-ed 1 3 0.10067 0.00058 *+-ed 2 3 0.07733 0.00252 *+

-ed 3 3 0.06800 0.00000 )*-ed 2 0.06350 0.00071 *+-ed 1 0.06300 * *+-ed 5 3 0.06000 0.00100 *-ed 6 3 0.05833 0.00058 *-ed 7 3 0.05700 0.00000 *

ello4 1 3 0.52233 0.00577 )*ello4 2 3 0.29333 0.00 62 *+ello4 3 3 0.17833 0.00153 *ello4 3 0.11533 0.00321 )*ello4 5 3 0.08600 0.00173 *ello4 6 3 0.07100 0.00100 *ello4 7 3 0.06 00 0.00100 *+

&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&& 0.12 0.2 0.36 0. 8



28/28

630 nm

Source DF SS MS F PSa le 22 3.583197 0.162873 19832.28 0.000Error 0 0.000329 0.000008Total 62 3.583526

S , 0.002866 -&S , 99.99! -&S )ad/+ , 99.99!

Indi idual 95! "Is For Mean #ased on Pooled StDe$e el % Mean StDe &&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&#lue 1 2 1.07150 0.01626 *#lue 1 1 1.05900 * *#lue 2 3 0.58 33 0.00153 *+#lue 3 3 0.31200 0.00 36 *+#lue 3 0.18167 0.00115 *#lue 5 3 0.11767 0.00058 *#lue 6 3 0.08567 0.00058 *#lue 7 3 0.07000 0.00000 *-ed 1 3 0.05667 0.00153 *

-ed 2 3 0.05567 0.00115 *-ed 3 3 0.05500 0.00000 *-ed 2 0.05700 0.001 1 *-ed 1 0.05600 * *-ed 5 3 0.05567 0.00058 *-ed 6 3 0.05533 0.00058 *-ed 7 3 0.05 33 0.00058 *

ello4 1 3 0.05733 0.00115 *ello4 2 3 0.05500 0.00000 *ello4 3 3 0.05 67 0.00058 *ello4 3 0.05 67 0.00058 *ello4 5 3 0.05 67 0.00058 *ello4 6 3 0.05 67 0.00058 *ello4 7 3 0.05 33 0.00058 *

&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&&&&&&&&&'&

0.30 0.60 0.90 1.20


Date post:	05-Jul-2018
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