An Introduction to Signal Processing in Chemical Analysis

An illustrated essay with software available for free download
Last updated June, 2009 PDF format: !na"Process n!.pdf #e$ format:"
Tom %'(a)er Professor *mer tus Department of &hem str+ and , ochem str+ -n )ers t+ of .ar+"and at &o""e!e Par/ *0ma ":

The nterfac n! of ana"+t ca" nstrumentat on to sma"" computers for the purpose of on0 " ne data ac2u s t on has no3 $ecome a"most standard pract ce n the modern chem str+ "a$orator+. -s n! 3 de"+0a)a "a$"e, "o30cost m crocomputers and off0the0she"f add0 n components, t s no3 eas er than e)er to ac2u re data 2u c/"+ n d ! ta" form. In 3hat 3a+s s on0" ne d ! ta" data ac2u s t on super or to the o"d methods such as the chart recorder4 Some of the ad)anta!es are o$) ous, such as arch )a" stora!e and retr e)a" of data and post0run re0p"ott n! 3 th ad5usta$"e sca"e e6pans on. *)en more mportant, ho3e)er, there s the poss $ " t+ of perform n! post0run data ana"+s s and s !na" process n!. There are a "ar!e num$er of computer0$ased numer ca" methods that can $e used to reduce no se, mpro)e the reso"ut on of o)er"app n! pea/s, compensate for nstrumenta" art facts, test h+potheses, opt m 7e measurement strate! es, d a!nose measurement d ff cu"t es, and decompose comp"e6 s !na"s nto the r component parts. These techn 2ues can often ma/e d ff cu"t measurements eas er $+ e6tract n! more nformat on from the a)a "a$"e data. .an+ of these techn 2ues are $ased on "a$or ous mathemat ca" procedures that 3ere not pract ca" $efore the ad)ent of computer 7ed nstrumentat on. It s mportant for chem str+ students to apprec ate the capa$ " t es and the " m tat ons of these modern s !na" process n! techn 2ues. In the chem str+ curr cu"um, s !na" process n! ma+ $e co)ered as part of a course on nstrumenta" ana"+s s 89, 2:, e"ectron cs for chem sts 8;:, "a$orator+ nterfac n! 8<:, or $as c chemometr cs 8=:. The purpose of th s paper s to ! )e a !enera" ntroduct on to some of the most 3 de"+ used s !na" process n! techn 2ues and to ! )e ""ustrat ons of the r app" cat ons n ana"+t ca" chem str+. Th s essa+ co)ers on"+ e"ementar+ top cs and s " m ted to on"+ $as c mathemat cs. For more ad)anced top cs and for a more r !orous treatment of the under"+ n! mathemat cs, refer to the e6tens )e " terature on chemometr cs. Th s tutor a" ma/es use of a free3are s !na"0process n! pro!ram ca""ed SP*&T>-. that 3as used to produce man+ of the ""ustrat ons. ?dd t ona" e6amp"es 3ere de)e"oped n .at"a$, a h !h0performance commerc a" numer ca" comput n! en) ronment and pro!ramm n! "an!ua!e that s 3 de"+ used n research. Para!raphs n !ra+ at the end of each sect on n th s essa+ descr $e the re"ated capa$ " t es of each of these pro!rams.

Signal arithmetic
The most $as c s !na" process n! funct ons are those that n)o")e s mp"e s !na" ar thmet c: po nt0$+0po nt add t on, su$tract on, mu"t p" cat on, or d ) s on of t3o s !na"s or of one s !na" and a constant. Desp te the r mathemat ca" s mp" c t+, these funct ons can $e )er+ usefu". For e6amp"e, n the "eft part of F !ure 9 8# ndo3 9: the top cur)e s the a$sorpt on spectrum of an e6tract of a samp"e of o " sha"e, a / nd of roc/ that s s a source of petro"eum.

Figure 1. A simple point-by--point subtraction of two signals allows the background (bottom curve on the left) to be subtracted from a complex sample (top curve on the left), resulting in a clearer picture of what is really in the sample (right). Th s spectrum e6h $ ts t3o a$sorpt on $ands, at a$out =9= nm and ==0 nm, that are due to a c"ass of mo"ecu"ar foss "s of ch"oroph+"" ca""ed porphyrins. 8Porph+r ns are used as !eomar/ers n o " e6p"orat on:. These $ands are super mposed on a $ac/!round a$sorpt on caused $+ the e6tract n! so")ents and $+ non0porph+r n compounds e6tracted from the sha"e. The $ottom cur)e s the spectrum of an e6tract of a non0porph+r n0$ear n! sha"e, sho3 n! on"+ the $ac/!round a$sorpt on. To o$ta n the spectrum of the sha"e e6tract 3 thout the $ac/!round, the $ac/!round 8$ottom cur)e: s s mp"+ su$tracted from the samp"e spectrum 8top cur)e:. The d fference s sho3n n the r !ht n # ndo3 2 8note the chan!e n @0a6 s sca"e:. In th s case the remo)a" of the $ac/!round s not perfect, $ecause the $ac/!round spectrum s measured on a separate sha"e samp"e. (o3e)er, t 3or/s 3e"" enou!h that the t3o $ands are no3 seen more c"ear"+ and t s eas er to measure prec se"+ the r a$sor$ances and 3a)e"en!ths. In th s e6amp"e and the one $e"o3, the assumpt on s $e n! made that the t3o s !na"s n # ndo3 9 ha)e the same 60a6 s )a"ues, that s, that $oth spectra are d ! t 7ed at the same set of 3a)e"en!ths. Str ct"+ spea/ n! th s operat on 3ou"d not $e )a" d f t3o spectra 3ere d ! t 7ed o)er d fferent 3a)e"en!th ran!es or 3 th d fferent nter)a"s $et3een ad5acent po nts. The 60a6 s )a"ues much match up po nt for po nt. In pract ce, th s s )er+ often the case 3 th data sets ac2u red 3 th n one e6per ment on one nstrument, $ut the e6per menter must ta/e care f the nstruments sett n!s are chan!ed or f data from t3o e6per ments or t3o nstrument are com$ ned. 8Aote: It s poss $"e to use the mathemat ca" techn 2ue of nterpo"at on to chan!e the num$er of po nts or the 60a6 s nter)a"s of

s !na"sB the resu"ts are on"+ appro6 mate $ut often c"ose enou!h n pract ce:. Somet mes one needs to /no3 3hether t3o s !na"s ha)e the same shape, for e6amp"e n compar n! the spectrum of an un/no3n to a stored reference spectrum. .ost " /e"+ the concentrat ons of the un/no3n and reference, and therefore the amp" tudes of the spectra, 3 "" $e d fferent. Therefore a d rect o)er"a+ or su$tract on of the t3o spectra 3 "" not $e usefu". %ne poss $ " t+ s to compute the po nt0$+0po nt rat o of the t3o s !na"sB f the+ ha)e the same shape, the rat o 3 "" $e a constant. For e6amp"e, e6am ne F !ure 2.

Figure 2. Do the two spectra on the left have the same shape !hey certainly do not look the same, but that may simply be due to that fact that one is much weaker that the other. !he ratio of the two spectra, shown in the right part ("indow #), is relatively constant from $%% to &&% nm, with a value of '% ()- %.#. !his means that the shape of these two signals is very nearly identical over this wavelength range. The "eft part 8# ndo3 9: sho3s t3o super mposed spectra, one of 3h ch s much 3ea/er than the other. ,ut do the+ ha)e the same shape4 The rat o of the t3o spectra, sho3n n the r !ht part 8# ndo3 2:, s re"at )e"+ constant from ;00 to <<0 nm, 3 th a )a"ue of 90 C/0 0.2. Th s means that the shape of these t3o s !na"s s the same, 3 th n a$out C/02 D, o)er th s 3a)e"en!th ran!e, and that top cur)e s a$out 90 t mes more ntense than the $ottom one. ?$o)e <<0 nm the rat o s not e)en appro6 mate"+ constantB th s s caused $+ noise, 3h ch s the top c of the ne6t sect on. S mp"e s !na" ar thmet c operat ons such as these are eas "+ done n a spreadsheet, an+ !enera"0purpose pro!ramm n! "an!ua!e, or a ded cated s !na"0process n! pro!ram such as SP*&T>-., 3h ch s a)a "a$"e for free do3n"oad. SP*&T>-. nc"udes add t on and mu"t p" cat on of a s !na" 3 th a constantB add t on, su$tract on, mu"t p" cat on, and d ) s on of t3o s !na"sB norma" 7at on, and a "ar!e num$er of other $as c math funct ons 8"o!, "n, ant "o!, s2uare root, rec proca", etc:. In .at"a$, math operat ons on s !na"s are espec a""+ po3erfu" $ecause the )ar a$"es n .at"a$ can $e e ther scalar 8s n!"e )a"ues:, vector 8" /e a ro3 or a co"umn n a spreadsheet:, represent n! one ent re s !na", spectrum or chromato!ram, or matrix 8" /e a rectan!u"ar $"oc/ of ce""s n a spreadsheet:, represent n! a set of s !na"s. For e6amp"e, n .at"a$ +ou cou"d def ne t3o )ectors a=[1 2 5 2 1] and b=[4 3 2 1 0]. Then to su$tract , from ? +ou 3ou"d 5ust t+pe a-b, 3h ch ! )es the resu"t [-3 -1 3 1 1]. To mu"t p"+ ? t mes , po nt $+ po nt, +ou 3ou"d t+pe a.*b, 3h ch ! )es the resu"t [4 6 10 2 0]. If +ou ha)e an ent re spectrum n the )ar a$"e a, +ou can p"ot t 5ust $+

t+p n! plot(a). ?nd f +ou a"so had a )ector w of 60a6 s )a"ues 8such as 3a)e"en!ths:, +ou can p"ot a )s w $+ t+p n! plot(w,a). The su$tract on of t3o spectra a and b, as n F !ure 9, can $e performed s mp"+ $+ 3r t n! a-b. To p"ot the d fference, +ou 3ou"d 3r te plot(a-b). L /e3 se, to p"ot the rat o of t3o spectra, as n F !ure 2, +ou 3ou"d 3r te plot(a./b). .oreo)er, .at"a$ s a pro!ramm n! "an!ua!e that can automate comp"e6 se2uences of operat ons $+ sa) n! them n scr pts and funct ons.

Signals and noise
*6per menta" measurements are ne)er perfect, e)en 3 th soph st cated modern nstruments. T3o ma n t+pes or measurement errors are reco!n 7ed: systematic error, n 3h ch e)er+ measurement s e ther "ess than or !reater than the EcorrectE )a"ue $+ a f 6ed percenta!e or amount, and random error, n 3h ch there are unpred cta$"e )ar at ons n the measured s !na" from moment to moment or from measurement to measurement. Th s "atter t+pe of error s often ca""ed noise, $+ ana"o!+ to acoust c no se. There are man+ sources of no se n ph+s ca" measurements, such as $u "d n! ) $rat ons, a r currents, e"ectr c po3er f"uctuat ons, stra+ rad at on from near$+ e"ectr ca" apparatus, nterference from rad o and TF transm ss ons, random therma" mot on of mo"ecu"es, and e)en the $as c 2uantum nature of matter and ener!+ tse"f. In spectroscop+, three fundamenta" t+pe of no se are reco!n 7ed: photon no se, detector no se, and f" c/er 8f"uctuat on: no se. Photon no se 8often the " m t n! no se n nstruments that use photomu"t p" er detectors:, s proport ona" to the s2uare root of " !ht ntens t+, and therefore the SA> s proport ona" to the s2uare root of " !ht ntens t+ and d rect"+ proport ona" to the s" t 3 dth. Detector no se 8often the " m t n! no se n nstruments that use so" d0state photod ode detectors: s ndependent of the " !ht ntens t+ and therefore the detector SA> s d rect"+ proport ona" to the " !ht ntens t+ and to the s2uare of the monochromator s" t 3 dth. F" c/er no se, caused $+ " !ht source nsta$ " t+, ) $rat on, samp"e ce"" pos t on n! errors, samp"e tur$u"ence, " !ht scatter n! $+ suspended part c"es, dust, $u$$"es, etc., s d rect"+ proport ona" to the " !ht ntens t+, so the f" c/er SA> s not decreased $+ ncreas n! the s" t 3 dth. F" c/er no se can usua""+ $e reduced or e" m nated $+ us n! spec a" 7ed nstrument des !ns such as dou$"e0$eam, dua" 3a)e"en!th, d ode arra+, and 3a)e"en!th modu"at on. The 2ua" t+ of a s !na" s often e6pressed 2uant tat )e"+ as the signal-to-noise ratio 8SA>: 3h ch s the rat o of the true s !na" amp" tude 8e.!. the a)era!e amp" tude or the pea/ he !ht: to the standard de) at on of the no se. S !na"0to0no se rat o s n)erse"+ proport ona" to the re"at )e standard de) at on of the s !na" amp" tude. .easur n! the s !na"0to0no se rat o usua""+ re2u res that the no se $e measured separate"+, n the a$sence of s !na". Depend n! on the t+pe of e6per ment, t ma+ $e poss $"e to ac2u re read n!s of the no se a"one, for e6amp"e on a se!ment of the $ase" ne $efore or after the occurrence of the s !na". (o3e)er, f the ma!n tude of the no se depends on the "e)e" of the s !na" 8as n photon no se or f" c/er no se n spectroscop+:, then the e6per menter must tr+ to produce a constant s !na" "e)e" to a""o3s measurement of the no se on the s !na". In a fe3 cases, 3here t s poss $"e to mode" the shape of the s !na" e6act"+ $+ means of a mathemat ca" funct on, the no se ma+ $e est mated $+ su$tract n! the mode" s !na" from the e6per menta" s !na".

use can $e made of th s fact $+ measur n! the s !na" o)er and o)er a!a n as fast as pract ca" and add n! up a"" the measurements po nt0$+0po nt. "indow ' (left) is a single measurement of a very noisy signal. %ne of the fundamenta" pro$"ems n s !na" measurement s d st n!u sh n! the no se from the s !na". Some e6amp"es that +ou can do3n"oad and use are these user0def ned funct ons to ca"cu"ate t+p ca" pea/ shapes common"+ encountered n ana"+t ca" chem str+. . . +ou can &tr"0c" c/ on these " n/s to nspect the code:.. @ou can a"so create user0def ned funct ons to automate common"+0 used a"!or thms. and t s one of the most po3erfu" methods for mpro) n! s !na"s. 8If +ou are ) e3 n! th s document on0" ne. such as mean.ften it is possible to average hundreds of measurement. p"us a s !na"0 !enerator that can $e used to !enerate art f c a" s !na"s 3 th Gauss an and Lorent7 an $ands. So f the s !na" can $e measured more than once. and norma""+0d str $uted random no se. ma6. . std. resulting is much more substantial improvement. 3h ch can $e usefu" n mode" n! and s mu"at n! ana"+t ca" s !na"s and test n! measurement techn 2ues. p"ot. and t+p ca" t+pes of random no se 83h teno se.plot(x. . the no se must $e random and the s !na" must occur at the same t me n each repeat.12". !here is actually a broad peak near the center of this signal. and height accurately because the signal-to-noise ratio is very poor (less than '). For th s to 3or/ proper"+.. Th s s ca""ed ensemble averaging. SP*&T>-. ran!e. %nce +ou ha)e created or do3n"oaded those funct ons. "indow # (right) is the average of * repeated measurements of this signal. s ne 3a)es. clearly showing the peak emerging from the noise. 3hen t can $e app" ed. width. !auss an and "orent7 an. ?n e6amp"e s sho3n n F !ure . nc"udes se)era" funct ons for measur n! s !na"s and no se. !he expected improvement in signalto-noise ratio is $ (the s+uare root of *). $+ t+p n! x=[1:256].at"a$ has $u "t0 n funct ons that can $e used for measur n! and p"ott n! s !na"s and no se. 3hereas the !enu ne s !na" s at "east part a""+ reproduc $"e. +ou can use them to p"ot a s mu"ated no s+ pea/ such as n F !ure . Th s s the $as s of f "ter n! and smooth n!. but it is not possible to measure its position.Figure 3. m n. p n/no se:. the s !na" ma+ conta n most"+ "o30fre2uenc+ components and the no se ma+ $e "ocated a h !her fre2uenc es.ut the th n! that rea""+ d st n!u shes s !na" from no se s that random no se s not the same from one measurement of the s !na" to the ne6t.64)#w$ t%!o s%(x)). h st. Somet mes the t3o can $e part"+ d st n!u shed on the $as s of fre2uenc+ components: for e6amp"e.gauss a!(x. .

the true s !na" amp" tudes 8+0a6 s )a"ues: chan!e rather smooth"+ as a funct on of the 60a6 s )a"ues.0po nt smooth 8mH. If the no se s E3h teE 8that s. If the no se n the data s E3h te no seE 8that s. 3h ch s mportant po nt $ecause t preser)es the 60 a6 s pos t on of pea/s and other features n the s !na". and t s a )er+ common 8$ut not necessar+: character st c of s !na"s that are ac2u red $+ automated and computer 7ed e2u pment. then the true s !na" 3 "" not $e much d storted $+ smooth n!. For a =0po nt smooth 8mH=:: for 5 H . 3here S5 the 5th po nt n the smoothed s !na". a$o)e. -sua""+ m s an odd num$er. Th s natura""+ "eads to a smoother s !na". For e6amp"e. 8Th s s espec a""+ cr t ca" for ana"+t ca" and spectroscop c app" cat ons $ecause the pea/ pos t ons are somet mes mportant measurement o$5ect )es:. m. and po nts that are "o3er than the ad5acent po nts are ncreased. then the standard de) at on of the no se rema n n! n the s !na" after the f rst pass of an un3e !hted s" d n!0a)era!e smooth 3 "" $e appro6 mate"+ s o)er the s2uare root of m 8s/s2rt8m::.:: for 5 H 2 to n09. Th s smooth s more effect )e at reduc n! h !h0fre2uenc+ no se n the s !na" than the s mp"er rectan!u"ar smooth. 3hereas man+ / nds of no se are seen as rap d. 3here m s a pos t )e nte!er ca""ed the smooth width. @5 the 5th po nt n the or ! na" s !na". Smooth n! usua""+ reduces the no se n a s !na". that s. 3here m s the smooth 3 dth. S m "ar smooth operat ons can $e constructed for an+ des red smooth 3 dth. to n02. random chan!es n amp" tude from po nt to po nt 3 th n the s !na". Th s s e2u )a"ent to t3o passes of a . Noise reduction.Smoothing In man+ e6per ments n ph+s ca" sc ence. for a . Aote that 3e are assum n! here that the 60a6 s nter)a"s of the s !na" s un form. Aote that a!a n n th s case. In smooth n!. then the . e)en"+ d str $uted o)er a"" fre2uenc es: and ts standard de) at on s s. The triangular smooth s " /e the rectan!u"ar smooth. The s mp"est smooth n! a"!or thm s the rectangular or unweighted sliding-average smoothB t s mp"+ rep"aces each po nt n the s !na" 3 th the a)era!e of m ad5acent po nts.0po nt rectan!u"ar smooth. In the "atter s tuat on t s common pract ce to attempt to reduce the no se $+ a process ca""ed smoothing. Th s s a"so assumed n man+ of the other s !na"0process n! techn 2ues descr $ed n th s essa+. that the d fference $et3een the 60a6 s )a"ues of ad5acent po nts s the same throu!hout the s !na". e6cept that t mp"ements a 3e !hted smooth n! funct on. and s m "ar"+ for other smooth 3 dths. e)en"+ d str $uted o)er a"" fre2uenc es: and ts standard de) at on s s. and n s the tota" num$er of po nts n the s !na". the 3 dth of the smooth m s an odd nte!er and the smooth coeff c ents are s+mmetr ca""+ $a"anced around the centra" po nt. Smoothing algorithms. the data po nts of a s !na" are mod f ed so that nd ) dua" po nts that are h !her than the mmed ate"+ ad5acent po nts 8presuma$"+ $ecause of no se: are reduced. ?s "on! as the true under"+ n! s !na" s actua""+ smooth. $ut the no se 3 "" $e reduced.

The opt mum cho ce of smooth 3 dth depends upon the 3 dth and shape of the s !na" and the d ! t 0 . on"+ a s" !ht mpro)ement o)er t3o passes 8tr an!u"ar smooth:.0po nt rectan!u"ar smooth s def ned on"+ for 5 H 2 to n09. respect )e"+. and width... The other approach s to use pro!ress )e"+ sma""er smooths at the ends of the s !na". The r !ht ha"f s the same pea/ after under!o n! a tr an!u"ar smooth n! a"!or thm.. height. Smooth n! operat ons can $e app" ed more than once: that s. area. he !ht. $ut a"so the !reater the poss $ " t+ that the s !na" 3 "" $e d storted $+ the smooth n! operat on. there 3 "" $e 8m09:/2 po nts at the $e! nn n! of the s !na" and 8m09:/2 po nts at the end of the s !na" for 3h ch a comp"ete m03 dth smooth can not $e ca"cu0 "ated. $ecause there are no data po nts $efore the f rst po nt or after the "ast po nt. K. There s not enou!h data n the s !na" to def ne a comp"ete . For e6amp"e.0po nt smooth for the f rst po nt n the s !na" 85 H 9: or for the "ast po nt 85 H n: . for e6amp"e to use 2. In some cases th s can $e usefu" f there s a !reat dea" of h !h0fre2uenc+ no se n the s !na".. !he left half of this signal is a noisy peak. Edge effects and the lost points problem. The no se s !reat"+ reduced 3h "e the pea/ tse"f s hard"+ chan!ed..and <. . n0. ? s mp"e e6amp"e of smooth n! s sho3n n F !ure <. 3 dth. Figure 4. 8That's the approach ta/en n the f !ures n th s paper:.J/s2rt8m:. for an m0 3 dth smooth. and therefore a smooth can not $e ca"cu"ated for the f rst t3o po nts or for the "ast t3o po nts. 3here m s the smooth 3 dth. !he noise is greatly reduced while the peak itself is hardly changed. 2.standard de) at on of the no se rema n n! n the s !na" after one pass of a tr an!u"ar smooth 3 "" $e appro6 mate"+ sI0. n02. po nt smooths for s !na" po nts 9. n09. The "ar!er the smooth 3 dth. (o3e)er. Smooth n! ncreases the s !na"0to0no se rat o and a""o3s the s !na" character st cs 8pea/ pos t on. etc. . In !enera".K/s2rt8m:. a pre) ous"+0smoothed s !na" can $e smoothed a!a n. %ne s to accept the "oss of po nts and tr m off those po nts or rep"ace them 3 th 7eros n the smooth s !na". The "eft ha"f of th s s !na" s a no s+ pea/. #hat to do4 There are t3o approaches. the !reater the no se reduct on. the no se reduct on for 3h te no se s "ess "ess n each success )e smooth. Examples of smoothing. a =0po nt smooth s def ned on"+ for 5 H .. to n02.. and for po nts n. =. espec a""+ 3hen computer0automated methods of "ocat n! and measur n! pea/s are $e n! emp"o+ed. S m "ar"+.. three passes of a rectan!u"ar smooth reduces 3h te no se $+ a factor of appro6 mate"+ sI0. !he right half is the same peak after undergoing a smoothing algorithm. The "ater approach ma+ $e prefera$"e f the ed!es of the s !na" conta n cr t ca" nformat on.. making it easier to measure the peak position. $ut t ncreases e6ecut on t me.. Aote n the e2uat ons a$o)e that the ..: to $e measured more accurate"+.

. the or ! na" pea/ 8 n red: has a true he !ht of 9. In the f !ure on the r !ht.ecause the pea/ 3 dth s J0 po nts. Th s ! )es a much more rea" st c mpress on of the performance of smooth n! Optimization of smoothing. the cr t ca" factor s the smoothing ratio. then smooth rat os $e"o3 0. the rat o $et3een the smooth 3 dth m and the num$er of po nts n the ha"f03 dth of the pea/. . po nts. and =9 po nts. the smooth ratios of these three smooths are "ar!er: 0. #h ch s the $est smooth rat o4 It depends on the purpose of the pea/"a$. Aote that e)en n the 3orst case. The three super mposed !reen " nes are the resu"ts of smooth n! th s pea/ 3 th a tr an!u"ar smooth of 3 dth 8from top to $ottom: K. In the f !ure on the "eft. respect )e"+.0 and a ha"f03 dth of . If the o$5ect )e of the measurement s to measure the true pea/ he !ht and 3 dth.7at on nter)a". 3h ch s hard to est mate ) sua""+ $ecause there are so fe3 "o30fre2uenc+ c+c"es n the s !na" record. For the "ar!est smooth. respect )e"+. @ou can see that the pea/ d stort on effect 8reduct on of pea/ he !ht and ncrease n pea/ 3 dth: s !reater for the narro3er pea/ $ecause the smooth rat os are h !her. 8In the e6amp"e on the . the pea/ has a 8true: he !ht of 2. Smooth rat os of !reater than 9. and =9/J0 H 0. to demonstrate the "o30fre2uenc+ no se that rema ns n the s !na" after smooth n!. For pea/0t+pe s !na"s. Th s s ""ustrated n the Interact )e Smooth n! modu"e for .09. the pea/ 3 dth s s" !ht"+ ncreased. The f !ures a$o)e sho3 e6amp"es of the effect of three d fferent smooth 3 dths on no s+ !auss an0shaped pea/s. In !enera". 3h ch nc"udes a E>esamp"eE contro" that s3aps the no se n the s !na" 3 th d fferent random no se samp"es. Th s causes the ) e3er to underestimate the contribution of low-fre+uency noise. and 9. 8It s a"so "ess no s+ than the e6amp"e on the "eft.==.9.29. ?s the smooth 3 dth ncreases. It's mportant to po nt out that smooth n! resu"ts such as ""ustrated n the f !ure a$o)e ma+ $e decept )e"+ mpress )e $ecause the+ emp"o+ a single sample of a no s+ s !na" that s smoothed to d fferent de!rees.0 are se"dom used $ecause of e6cess )e pea/ d stort on.. po nts. the no se s pro!ress )e"+ reduced $ut the pea/ he !ht a"so s reduced s" !ht"+.0 and there are J0 po nts n the ha"f03 dth of the pea/. 2=/J0 H 0.: The three super mposed !reen " nes are the resu"ts of the same three tr an!u"ar smooths of 3 dth 8from top to $ottom: K. 2=. 2=.ut $ecause the pea/ 3 dth n th s case s on"+ .L<.. 0. Th s error can $e remed ed $+ ta/ n! a "ar!e num$er of ndependent samp"es of no s+ s !na".2 shou"d $e used. the smooth ratios of these three smooths are K/J0 H 0.KL. the pea/ pos t ons are not effected 8assum n! that the or ! na" pea/s 3ere s+mmetr ca" and not o)er"apped $+ other pea/s:. and =9 po nts. ncreas n! the smooth n! rat o mpro)es the s !na"0to0no se rat o $ut causes a reduct on n amp" tude and n ncrease n the $and3 dth of the pea/. The red " ne s the or ! na" unsmoothed pea/. .

. for e6amp"e f the "ocat on of ma6 ma. the pea/ he !ht reduct on caused $+ smooth n! s not so mportant. to prepare a n cer0"oo/ n! !raph c of a s !na" for ) sua" nspect on or pu$" cat on. and 82: f the s !na" 3 "" $e su$se2uent"+ processed $+ an a"!or thm that 3ou"d $e ad)erse"+ effected $+ the presence of too much h !h0fre2uenc+ no se n the s !na". 8The no se s reduced $+ appro6 mate"+ the s2uare root of the smooth 3 dth:. n a popu"ar techn 2ue for pea/ f nd n! and measurement. are n t a""+ ser ous"+ naccurate $ecause of the no se. enta " n! at "east some chan!e n s !na" shape and amp" tude. the pea/ he !ht reduct on of the standard s !na"s 3 "" $e e6act"+ the same as that of the samp"e s !na"s and the effect 3 "" cance" out e6act"+.. 3hereas the smoothed pea/ 3 th a smooth rat o of 0. other3 se a "ar!e s+stemat c error ma+ resu"t..ut f the o$5ect )e of the measurement s to measure the pea/ pos t on 860a6 s )a"ue of the pea/:. ?s the smooth 3 dth s ncreased. ?n attempt to measure the pea/ amp" tude and pea/ 3 dth of the no s+ s !na". the or ! na" pea/ 8red " ne: has a pea/ he !ht !reater than the true )a"ue 2. $ut the pos t on. If these re2u rements conf" ct. In pract ca" ana"+t ca" chem str+. .= to 9.ut one common s tuat on 3here +ou shou"d not smooth s !na"s s pr or to "east0s2uares cur)e f tt n!. When should you smooth a signal? There are t3o reasons to smooth a s !na": 89: for cosmet c reasons. he !ht. $ecause n most cases ca" $rat on s $ased on the s !na"s of standard so"ut ons. In such cases smooth 3 dths from 0. sho3n at the $ottom of the ) deo. care must $e used n the des !n of a"!or thms. or nf"ect on po nts n the s !na" s to $e automat ca""+ determ ned $+ detect n! 7ero0cross n!s n der )at )es of the s !na". e)en f hea)+ smooth n! s necessar+ to pro) de re" a$"e d scr m nat on a!a nst no se pea/s... a$so"ute pea/ he !ht measurements are se"dom re2u redB ca" $rat on a!a nst standard so"ut ons s the ru"e. to app"+ exactly the same s !na" process n! steps to the standard s !na"s as to the samp"e s !na"s.0 and 3 der than 200. . m m ma.0 and pea/ 3 dth of 200. and 3 dth of each pea/ s determ ned $+ "east0s2uares cur)e0f tt n! of a se!ment of or ! na" unsmoothed data n the ) c n t+ of the 7ero0cross n!.: It s )er+ mportant.3m): demonstrates the effect of tr an!u"ar smooth n! on a s n!"e Gauss an pea/ 3 th a pea/ he !ht of 9. much "ar!er smooth rat os can $e emp"o+ed f des red. a$o)e a smooth 3 dth of a$out <0 8smooth rat o 0.. .+te ) deo 8Smooth. Thus.0 $ecause of the no se. The n t a" 3h te no se amp" tude s 0. e)en thou!h the s !na"0to0no se .09 has a pea/ he !ht that s much c"oser to the correct )a"ue:.2:.0 can $e used f necessar+ to further mpro)e the s !na"0to0no se rat o. If the same s !na" process n! operat ons are app" ed to the samp"es and to the standards. $ecause a"" smooth n! a"!or thms are at "east s" !ht"+ E"oss+E."eft. Th s 9J0second. ! ) n! an n t a" s !na"0to0 no se rat o of a$out . pea/s are "ocated $+ detect n! do3n3ard 7ero0cross n!s n the smoothed f rst der )at )e.. ho3e)er. ho3e)er. In 2uant tat )e ana"+s s app" cat ons. . the s !na"0to0no se rat o mpro)es and the accurac+ of the measurements of pea/ amp" tude and pea/ 3 dth are mpro)ed. $ecause smooth n! has no effect at a"" on the pea/ pos t on of a s+mmetr ca" pea/ 8un"ess the ncrease n pea/ 3 dth s so much that t causes ad5acent pea/s to o)er"ap:. (o3e)er. For e6amp"e. the pea/ parameters e6tracted $+ cur)e f tt n! are not d storted. Video Demonstration. the smooth n! causes the pea/ to $e shorter than 9. 8>emem$er: the o$5ect )e of a 2uant tat )e ana"+t ca" spectrophotometr c procedure s not to measure a$sor$ance $ut rather to measure the concentrat on of the ana"+te.

8@ou can "ea)e off the "ast t3o nput ar!uments: &asts'oot$(*. ! )es a pseudo0Gauss an 8e2u )a"ent to . 8Th s demonstrat on 3as created n . nc"udes rectan!u"ar and tr an!u"ar smooth n! funct ons for an+ num$er of po nts. >un SmoothS" derTest.%)g%). d+/"a$ Interact )e Smooth n! m0f "es.rat o cont nues to mpro)e as the smooth 3 dth s ncreased. the s mp"est a"!or thm for comput n! a f rst der )at )e s: . +ou can ctr"0c" c/ on th s " n/ to nspect the code:.7 p.= us n! EInteract )e Smooth n! for . ?ssum n! that the 60 nter)a" $et3een ad5acent po nts s constant. fastsmooth t+p ca""+ ! )es much faster e6ecut on t mes.w) smooths 3 th t+peH9 and ed!eH0:. Smoothing in"a$ L. The ar!ument Eed!eE contro"s ho3 the Eed!esE of the s !na" 8the f rst 3/2 po nts and the "ast 3/2 po nts: are hand"ed. 8In th s mode the e"apsed t me s ndependent of the smooth 3 dth. The smoothed s !na" s returned as the )ector EsE. SP*&T>-. Fastsmooth s a .w. Interact )eSmooth n!.w. The user0def ned funct on fastsmooth mp"ements a"" the t+pes of smooths d scussed a$o)e. 3 dth. the ed!es are 7ero. +ou ma+ do3n"oad the comp"ete set of . If ed!eH0. that s. The ar!ument EaE s the nput s !na" )ectorB E3E s the smooth 3 dthB Et+peE determ nes the smooth t+pe: t+peH9 ! )es a rectan!u"ar 8s" d n!0a)era!e or $o6car:B t+peH2 ! )es a tr an!u"ar 8e2u )a"ent to 2 passes of a s" d n! a)era!e:B t+peH. &an $e used 3 th an+ smooth n! funct on.t(p%) smooths 3 th ed!eH0 and &asts'oot$(*. If +ou ha)e access to that"a$ s a .9 second. Inc"udes a se"f0conta ned nteract )e demo of the effect of smooth n! on pea/ he !ht. 3h ch s nterpreted as the slope of the tan!ent to the s !na" at each po nt. the ed!es are smoothed 3 th pro!ress )e"+ sma""er smooths the c"oser to the end. If ed!eH9.000 po nt s" d n! a)era!e n "ess than"a$ funct on of the form s=&asts'oot$(a. &ompared to con)o"ut on0$ased smooth a"!or thms. Th s ! )es the fastest e6ecut on t me:. 8If +ou are ) e3 n! th s document on0" ne. 8In th s mode the e6ecut on t me ncreases 3 th ncreas n! smooth 3 dths:. 3 th s" ders that a""o3 +ou to ad5ust the smooth n! parameters cont nuous"+ 3h "e o$ser) n! the effect on +our s !na" d+nam ca""+. The f rst der )at )e of a s !na" s the rate of chan!e of + 3 th 6. Interact )e Smooth n! for .000 po nt s !na" 3 th a"a$E modu"e:. Differentiation The s+m$o" c d fferent at on of funct ons s a top c that s ntroduced n a"" e"ementar+ &a"cu"us courses.t(p%. and s !na"0to0no se rat o. espec a""+ for "ar!e smooth 3 dthsB t can smooth a"a$ modu"e for nteract )e smooth n! for t me0 ser es s !na"s.m to see ho3 t 3or/s. The numer ca" d fferent at on of d ! t 7ed s !na"s s an app" cat on of th s concept that has man+ uses n ana"+t ca" s !na" process n!. 892 M$+tes: so that +ou can e6per ment 3 th a"" the )ar a$"es at 3 "" and tr+ out th s techn 2ue on +our o3n s !na". passes of a s" d n! a)era!e:.

The second derivative s the der )at )e of the der )at )e: t s a measure of the curvature of the s !na". that s. 3here O'5 and @'5 are the O and @ )a"ues of the 5th po nt of the der )at )e. ts der )at )e s pos t )eB 3here a s !na" s"opes do3n. s the f rst der )at )e of # ndo3 2. The s !na" n each of the four 3 ndo3s s the f rst der )at )e of the one $efore tB that s. n H num$er of po nts n the s !na". ? common"+ used )ar at on of th s a"!or thm computes the a)era!e s"ope $et3een three ad5acent po nts: 8for 2 N 5 Nn09:. # ndo3 . # ndo3 . The s mp"est a"!or thm for d rect computat on of the second der )at )e n one step s 8for 2 N 5 Nn09:. the rate of chan!e of the s"ope of the s !na". @ou can pred ct the shape of each s !na" $+ reca"" n! that the der )at )e s s mp"+ the s"ope of the or ! na" s !na": 3here a s !na" s"opes up. a"thou!h these der )at )es can a"so $e computed s mp"+ $+ ta/ n! success )e "o3er order der )at )es. # ndo3 2 s the f rst der )at )e of # ndo3 9. Th s corresponds to the maximum n ts f rst der )at )e 8# ndo3 2: and to the -ero-crossing 8po nt 3here the s !na" crosses the 60a6 s !o n! e ther from pos t )e to ne!at )e or vice versa: n the second der )at )e n # ndo3 . ts der )at )e s ne!at )eB and 3here a s !na" has 7ero s"ope. !asic "roperties of Deri#ati#e Signals The f !ure on the "eft sho3s the resu"ts of the success )e d fferent at on of a computer0 !enerated s !na". and so on. 0<. C9 for the <th der )at )e. ts der )at )e s 7ero. S m "ar"+. s the second der )at )e of # ndo3 9. The s !mo da" s !na" sho3n n # ndo3 9 has an inflection point 8po nt 3here 3here the s"ope s ma6 mum: at the center of the 6 a6 s ran!e. C2. Th s $eha) or can $e usefu" for prec se"+ "ocat n! the nf"ect on po nt n a s !mo d s !na". It can $e ca"cu"ated $+ app"+ n! the f rst der )at )e ca"cu"at on t3 ce n success on. 0<. 09 for the th rd der )at )e and C9.8for 9N 5 Nn09:. CL. 02. h !her der )at )e orders can $e computed us n! the appropr ate se2uence of coeff c ents: for e6amp"e C9.. and DO s the d fference $et3een the O )a"ues of ad5acent data po nts. $+ comput n! the "ocat on of the 7ero0cross n! n ts .

*ach se!ment s no3 c"ear"+ seen as a separate step 3hose he !ht 8+0a6 s )a"ue: s the s"ope. 3here the $rea/s $et3een then fa"". t s found that that the amp" tude of the nth der )at )e of a pea/ s n)erse"+ proport ona" to the nth po3er of ts 3 dth. The t3o $ands ha)e the same amp" tude 8pea/ he !ht: $ut one of them s e6act"+ t3 ce the 3 dth of the other. and the effect $ecomes more not cea$"e at h !her der )at )e orders. Th s s !na" s t+p ca" of the t+pe of s !na" recorded n amperometr c t trat ons and some / nds of therma" ana"+s s and / net c e6per ments: a ser es of stra !ht " ne se!ments of d fferent s"ope. The f !ure on the "eft sho3s the resu"ts of the success )e d fferent at on of t3o computer0!enerated Gauss an $ands. Thus d fferent at on n effect d scr m nates a!a nst 3 der pea/s and the h !her the order of d fferent at on the !reater the d scr m nat on. Aote that n th s e6amp"e the steps n the der )at )e s !na" are not comp"ete"+ f"at. ?nother mportant propert+ of the d fferent at on of pea/0t+pe s !na"s s the effect of the pea/ 3 dth on the amp" tude of der )at )es. The o$5ect )e s to determ ne ho3 man+ se!ments there are. and the s"opes of each se!ment. ?s +ou can see. In !enera". $ecause the s"ope d fferences are sma"" and the reso"ut on of the computer screen d sp"a+ s " m t n!.second der )at )e. $pplications of Differentiation ? s mp"e e6amp"e of the app" cat on of d fferent at on of e6per menta" s !na"s s sho3n n F !ure =. The +0a6 s no3 ta/es on the un ts of d+/d6. The tas/ s much s mp"er f the f rst der )at )e 8s"ope: of the s !na" s ca"cu"ated 8F !ure =. r !ht:. nd cat n! that the " ne se!ments n the or ! na" s !na" 3ere not perfect"+ stra !ht. Th s s d ff cu"t to do from the ra3 data. t s more not cea$"e n the der )at )e . Th s s most " /e"+ due to random no se n the or ! na" s !na". . ?"thou!h th s no se 3as not part cu"ar"+ e) dent n the or ! na" s !na". S m "ar"+. the wider pea/ has the smaller der )at )e amp" tude. the "ocat on of ma6 mum n a pea/0t+pe s !na" can $e computed prec se"+ $+ comput n! the "ocat on of the 7ero0cross n! n ts f rst der )at )e. Th s $eha) or can $e usefu" n 2uant tat )e ana"+t ca" app" cat ons for detect n! pea/s that are super mposed on and o$scured $+ stron!er $ut $roader $ac/!round pea/s.

but its numerically calculated derivative (dx)dy). !he endpoint is the point of greatest slope2 this is also an inflection point. c"ose to the theoret ca" )a"ue of 20 mL. The dotted " nes sho3 that the 7ero cross n! fa""s at a$out 99. with volume in m/ on the 0-axis and p. . # th a 3ea/ ac d such as th s. on the 1-axis. un"ess the d fferent at on a"!or thm nc"udes smooth n! that s carefu""+ opt m 7ed for each app" cat on. shows that the line actually has several approximately straight-line segments with distinctly different slopes and with well-defined breaks between each segment. the t trat on cur)e has a s !mo da" shape and the endpo nt s nd cated $+ the inflection point. The 7ero cross n! of the second der )at )e corresponds to the endpo nt and s much more prec se"+ measura$"e. ? c"ass c use of second d fferent at on n chem ca" ana"+s s s n the "ocat on of endpo nts n potent ometr c t trat on. where the curvature of the signal is -ero. It s common"+ o$ser)ed that d fferent at on de!rades s !na"0to0no se rat o. it is difficult to locate this point precisely from the original titration curve. as the the -ero crossing. !he endpoint is much more easily located in the second derivative. "ith a weak acid such as this. Aote that n the second der )at )e p"ot. the po nt 3here the s"ope s ma6 mum and the cur)ature s 7ero.a6 ma and 7ero cross n!s are usua""+ much eas er to "ocate prec se"+ than nf"ect on po nts. The endpo nt s the po nt of !reatest s"opeB th s s a"so an nf"ect on po nt. 3 th )o"ume n mL on the O0a6 s and p( on the @0a6 s. Aumer ca" a"!or thms for d fferent at on are as numerous as for smooth n! and must $e carefu""+ chosen to contro" s !na"0to0no se de!radat on. . shown on the right. and the second der )at )e 3 "" e6h $ t a -ero-crossing at that po nt.Figure 5.< mL. t s d ff cu"t to "ocate th s po nt prec se"+ from the or ! na" t trat on cur)e. !he signal on the left seems to be a more-or-less straight line. $oth the 60a6 s and the +0a6 s sca"es ha)e $een e6panded to sho3 the 7ero cross n! po nt more c"ear"+. In most t trat ons. titration curve of a very weak acid with a strong base. F !ure L sho3s a p( t trat on cur)e of a )er+ 3ea/ ac d 3 th a stron! $ase. Figure 6 !he signal on the left is the p. The )o"umetr c e2u )a"ence po nt 8the Etheoret ca"E endpo nt: s 20 mL. The f rst der )at )e of the t trat on cur)e 3 "" therefore e6h $ t a maximum at the nf"ect on po nt. plotted on the right. 3here the cur)ature of the s !na" s 7ero. The second der )at )e of the cur)e s sho3n n # ndo3 2 on the r !ht.

3h ch sho3s a s mu"ated -F spectrum 8a$sor$ance )s 3a)e"en!th n nm:. the amp" tude of a der )at )e s proport ona" to the amp" tude of the or ! na" s !na". In order for th s techn 2ue to 3or/. Th s effect s cons dera$"+ enhanced n the second der )at )e. Fer+ often n the pract ca" app" cat ons of spectrophotometr+ to the ana"+s s of comp"e6 samp"es.0) s $"e a$sorpt on. Th s s the $as s for the app" cat on of d fferent at on as a method of correct on for $ac/!round s !na"s n 2uant tat )e spectrophotometr c ana"+s s. !radua""+ cur)ed $ac/!round. the compound to $e measured: are super mposed on a $road. u. referred to as derivative spectroscopy. as a techn 2ue for the correct on for rre"e)ant $ac/!round a$sorpt on and as a 3a+ to fac " tate mu"t component ana"+s s. t s necessar+ that the $ac/!round a$sorpt on $e $roader 8that s. d fferent at on ma+ $e emp"o+ed as a !enera" 3a+ to d scr m nate a!a nst $road spectra" features n fa)or of narro3 components. as a 2ua" tat )e f n!erpr nt n! techn 2ue to accentuate sma"" structura" d fferences $et3een near"+ dent ca" spectraB 8$: spectra" reso"ut on enhancement. . $ut th s turns out to . Th s s ""ustrated $+ the f !ure on the "eft. sho3n on the r !ht.ecause of the fact that the amp" tude of the nth der )at )e of a pea/0shaped s !na" s n)erse"+ proport ona" to the nth po3er of the 3 dth of the pea/.e. part cu"ar"+ n nfra0red.ost commerc a" spectrophotometers no3 ha)e $u "t0 n der )at )e capa$ " t+. 3 th the !reen cur)e represent n! the spectrum of the pure ana"+te and the red " ne represent n! the spectrum of a m 6ture conta n n! the ana"+te p"us other compounds that ! )e r se to the "ar!e s"op n! $ac/!round a$sorpt on. . . ha)e "o3er cur)ature: than the ana"+te spectra" pea/.ecause d fferent at on s a " near techn 2ue. 3h ch a""o3s 2uant tat )e ana"+s s app" cat ons emp"o+ n! an+ of the standard ca" $rat on techn 2ues:. 8. the spectra" $ands of the ana"+te 8 . the d fferent at on of spectra s a 3 de"+ used techn 2ue. In th s case the spectra of the pure ana"+te and of the m 6ture are a"most dent ca".ac/!round of th s t+pe can $e reduced $+ d fferent at on. and ref"ectance spectrophotometr+. Der )at )e methods ha)e $een used n ana"+t ca" spectroscop+ for three ma n purposes: 8a: spectra" d scr m nat on. as a techn 2ue for ncreas n! the apparent reso"ut on of o)er"app n! spectra" $ands n order to more eas "+ determ ne the num$er of $ands and the r 3a)e"en!thsB 8c: 2uant tat )e ana"+s s. f"uorescence. $+ means of dua" 3a)e"en!th or 3a)e"en!th modu"at on des !ns. Some nstruments are des !ned to measure the spectra" der )at )es opt ca""+.Deri#ati#e Spectroscopy In spectroscop+. The f rst der )at )es of these t3o s !na"s are sho3n n the centerB +ou can see that the d fference $et3een the pure ana"+te spectrum 8!reen: and the m 6ture spectrum 8red: s reduced.).

%race $nalysis %ne of the 3 dest uses of the der )at )e s !na" process n! techn 2ue n pract ca" ana"+t ca" 3or/ s n the measurement of sma"" amounts of su$stances n the presence of "ar!e amounts of potent a""+ nterfer n! mater a"s. to sa+ that $% miles per hour s !reater than #% miles. ?n e6amp"e of the mpro)ed a$ " t+ to detect trace component n the presence of stron! $ac/!round nterference s sho3n n F !ure K. For e6amp"e. It s somet mes 8m sta/en"+: sa d that d fferent at on E ncreases the sens t ) t+E of ana"+s s. . espec a""+ 3hen the ana"+te s !na" s sma"" compared to the $ac/!round and f there s a "ot of uncontro""ed )ar a$ " t+ n the $ac/!round. . The un ts of the or ! na" spectrum are absorbanceB the un ts of the f rst der )at )e are absorbance per nm. ?n o$) ous $enef t of the suppress on of $road $ac/!round $+ d fferent at on s that variations n the $ac/!round amp" tude from samp"e to samp"e are a"so reduced.: @ou can. for nstance. compare the signal-tobackground ratio and the signal-to-noise ratio. t s not )a" d to compare the amp" tudes of s !na"s and the r der )at )es $ecause the+ ha)e d fferent un ts.$e a rather common s tuat on. and so"ut on tur$ d t+. second 8and somet mes e)en h !her0order: der )at )es are often used for such purposes. Th s can resu"t n mpro)ed prec s on or measurement n man+ nstances. (o3e)er.easurement prec s on s often de!raded $+ samp"e0to0samp"e $ase" ne sh fts due to non0 spec f c $road$and nterfer n! a$sorpt on. In such app" cat ons t s common that the ana"+t ca" s !na"s are 3ea/. @ou can't compare absorbance to absorbance per nm an+ more than +ou can compare miles to miles per hour. . 8It's mean n!"ess. d rt or f n!erpr nts on the cu)ette 3a""s. @ou can see ho3 t 3ou"d $e tempt n! to sa+ someth n! " /e that $+ nspect n! the three f !ures a$o)eB t does seems that the s !na" amp" tude of the der )at )es s !reater 8at "east !raph ca""+: than that of the or ! na" ana"+te s !na". and the the un ts of the second der )at )e are absorbance per nm#. no s+. t 3ou"d $e )a" d to sa+ that the s !na"0to0$ac/!round rat o s ncreased n the der )at )es. and super mposed on "ar!e $ac/!round s !na"s.ecause of the r !reater d scr m nat on a!a nst $road $ac/!round. ho3e)er.ase" ne sh fts from these sources are usua""+ e ther 3a)e"en!th0 ndependent 8" !ht $"oc/a!e caused $+ $u$$"es or "ar!e suspended part c"es: or e6h $ t a 3ea/ 3a)e"en!th dependence 8sma""0part c"e tur$ d t+:. . n the a$o)e e6amp"e. non0reproduc $"e cu)ette pos t on n!. Therefore t can $e e6pected that d fferent at on 3 "" n !enera" he"p to d scr m nate re"e)ant a$sorpt on from these sources of $ase" ne sh ft. mperfect cu)ette transm ss on match n!.

Th s s essent a""+ the same spectrum as n F !ure K. !he fourth derivative (right) shows that a peak is still there. !he spectrum on the left shows a weak shoulder near the center due to a small concentration of the substance that is to be measured (e. !he background has been almost completely suppressed and the analyte peak now stands out clearly. Th s use of s !na" d fferent at on has $ecome 3 de"+ used n 2uant tat )e spectroscop+. $ut ne)erthe"ess the s !na"0to0no se rat o s suff c ent"+ !ood for a reasona$"e 2uant tat )e measurement. the active ingredient in a pharmaceutical preparation). The $ac/!round has $een a"most comp"ete"+ suppressed and the ana"+te pea/ no3 stands out c"ear"+. s"op n! $ac/!round o$scures the pea/ and ma/es 2uant tat )e measurement )er+ d ff cu"t. but much reduced in amplitude (note the smaller y-axis scale). ?n e)en more dramat c case s sho3n n F !ure J. The 2uest on s: s there a detecta$"e amount of ana"+te n th s spectrum4 Th s s 2u te mposs $"e to sa+ from the norma" spectrum. sta$ " 7ers. e6cept that the concentrat on of the ana"+te s "o3er. care must $e ta/en to opt m 7e s !na"0to0no se rat o of the nstrument as much as poss $"e. !he fourth derivative of this spectrum is shown on the right. In that app" cat on the ana"+te 3ou"d t+p ca""+ $e the act )e n!red ent n a pharmaceut ca" preparat on and the $ac/!round nterferences m !ht ar se from the presence of f ""ers. f"a)or n! or co"or n! a!ents. but in the case the peak is so weak that it can not even be seen in the spectrum on the left. n trace ana"+s s app" cat ons. Figure 8. %f course.Figure 7. The spectrum on the "eft sho3s a 3ea/ shou"der near the center due to the ana"+te.g. 3t is difficult to measure the intensity of this peak because it is obscured by the strong background caused by other substances in the sample. facilitating measurement. t . 4imilar to 5igure 6. %he &mportance of Smoothing Deri#ati#es For the successfu" app" cat on of d fferent at on n 2uant tat )e ana"+t ca" app" cat ons. emu"s f ers. The s !na"0to0no se rat o s )er+ !ood n th s spectrum. $ut nspect on of the fourth der )at )e 8r !ht: sho3s that the ans3er s yes. The fourth der )at )e of th s spectrum s sho3n on the r !ht. fac " tat n! measurement. $ut n sp te of that the $road. part cu"ar"+ for 2ua" t+ contro" n the pharmaceut ca" ndustr+. $uffers. Some no se s c"ear"+ e) dent here. or other e6c p ents.

3 thout suff c ent smooth n!. # ndo3 9 sho3s a Gauss an $and 3 th a sma"" amount of added no se. smooth n!. th s 3on't $e a pro$"em. the s !na"0to0no se rat o of the unsmoothed second der )at )e 8# ndo3 2: s so poor +ou can not e)en see the s !na" ) sua""+. ho3e)er. Th s s ""ustrated n the f !ure on the "eft.. a$o)e: s much "ess than ts "ess0smoothed )ers on 8# ndo3 . 3h ch a""o3s 2uant tat )e ana"+s s app" cat ons emp"o+ n! an+ of the standard ca" $rat on techn 2ues:. the s !na"0to0no se rat o of the der )at )e can $e su$stant a""+ poorer than the or ! na" s !na". . (o3e)er. Th s effect s e)en more str / n! n the second der )at )e. (o3e)er.0. The opt mum )a"ues of smooth rat o for der )at )e s !na"s s appro6 mate"+ 0. #hat s mportant. 3 th ade2uate amounts of smooth n!. ts smooth rat o 8rat o of the smooth 3 dth to the 3 dth of the or ! na" pea/:. the s !na"0to0no se rat o of the smoothed der )at )e can $e $etter than that of the unsmoothed or ! na". n order to opt m 7e the s !na"0to0no se rat o. For a second der )at )e. sho3 the f rst der )at )e of that s !na" 3 th ncreas n! smooth 3 dths. 8. and <.s essent a" to use d fferent at on n com$ nat on 3 th suff c ent smooth n!. and the num$er of t mes the s !na" s smoothed. as sho3n $e"o3. three app" cat ons of a s mp"e rectan!u"ar smooth or t3o app" cat ons of a tr an!u"ar smooth s ade2uate. For a f rst der )at )e. It ma/es no d fference 3hether the smooth operat on s app" ed $efore or after the d fferent at on. t3o app" cat ons of a s mp"e rectan!u"ar smooth or one app" cat on of a tr an!u"ar smooth s ade2uate. . Such hea)+ amounts of smooth n! resu"t n su$stant a" attenuat on of the der )at )e amp" tudeB n the f !ure on the r !ht a$o)e. the amp" tude of the most hea) "+ smoothed der )at )e 8 n # ndo3 <.ecause d fferent at on and smooth n! are $oth " near techn 2ues. s the nature of the smooth. and measurement procedure as s app" ed to the un/no3n samp"es. ?s +ou can see. as "on! as the standard 8ana"+t ca": cur)e s prepared us n! the e6act same der )at )e. the amp" tude of a smoothed der )at )e s proport ona" to the amp" tude of the or ! na" s !na". The !enera" ru"e s: for the nth der )at )e. use at "east nC9 app" cat ons of rectan!u"ar smooth 8or ha"f that num$er of tr an!u"ar smooths:.= to 9.:. # ndo3s 2. In th s case.

and der )<. and t3o nteract )e )ers ons. )ar a$"e $ac/!"a$ funct ons and scr pts for nteract )e d fferent at on of t me0 ser es s !na""a$"a$ F "e *6chan!e. These are a"" deta "ed $e"o3. and 3 dth of each pea/ $+ "east0s2uares cur)e0f tt n!.000. se"f0conta ned demos to sho3 ho3 t 3or/s. 8Th s s usefu" pr mar "+ for s !na"s that ha)e se)era" data po nts n each pea/. . Differentiation in atlab Deri#ati#e'based "ea( )inding and easurement in atlab Th s sect on descr $es a fast. smooth 3 dth. not for sp /es that ha)e on"+ one or t3o po nts:.(a)B the nput ar!ument s a s !na" )ector EaE. se)era" se"f0conta ned demos to sho3 ho3 t 3or/s. Determ nes the pos t on.SP*&T>-. custom 7a$"e .m:B 82: an nteract )e /e+press0operated funct on 8 pea/. The rout ne s a)a "a$"e n three d fferent )ers ons: 89: the $as c f ndpea/s funct on 8f ndpea/"a$ funct on of the form )=)%+ .at"a$. Interact )e s" ders a""o3 +ou to contro" a"" the parameters. 3 th s" ders that a""o3 +ou to ad5ust the der )at )e order. +ou can ctr"0c" c/ on these " n/s to nspect the code. and the d fferent ated s !na" s returned as the )ector EdE. >un Interact )eDer )at )eTest to see ho3 t 3or/s. @ou can a"so do3n"oad t from the .0po nt centra"0d fference method.m. a s mp"e second der )at )e us n! the .7 pE that a"so nc"udes support n! funct ons.=.: a scr pt us n! mouse0contro""ed s" ders for nteract )e contro".at"a$ s a co""ect on of . 3h ch demonstrates the app" cat on of d fferent at on to the detect on of pea/s super mposed on a stron!. a f rst der )at )e us n! the 20po nt centra"0d fference method. us n! a <0po nt formu"a.3m). der )2. nc"udes f rst and second der )at )e funct ons. It detects pea/s $+ "oo/ n! for do3n3ard 7ero0 cross n!s n the smoothed f rst der )at )e. a <th der )at )e us n! a =0po nt formu"!round2. then d fferent ates and smooths t. a th rd der )at )e der ).m: for ad5ust n! the pea/ detect on cr ter a n rea"0t me to opt m 7e for an+ part cu"ar pea/ t+peB and 8. If +ou are ) e3 n! th s document on0" ne. D fferent at on funct ons such as descr $ed a$o)e can eas "+ $e created n . and measures the s !na" ran!e and s !na"0to0no se rat o 8SA>:. adds random no se and a )ar a$"e $ac/!round. &tr"0c" c/ here to do3n"oad the PIP f "e EInteract )eDer )at )e. It can f nd and measure 9000 pea/s n a 9. &" c/ here to do3n"oad the PIP f "e EPea/f nder. ?"so nc"udes Der )at )eDemo. and sca"e e6pans on cont nuous"+ 3h "e o$ser) n! the effect on +our s !na" d+nam ca""+. Generates a s !na" pea/.7 pE that nc"udes support n! funct ons. Some s mp"e e6amp"es that +ou can do3n"oad nc"ude: der ). he !"a$ rout ne for "ocat n! and measur n! the pea/s n no s+ t me0ser es data sets. Th s 3as used to create the ) deo demonstrat on Der )at )e.000 po nt s !na" n J seconds. 3h ch can $e app" ed success )e"+ to compute der )at )es of an+ order. Interact )e Der )at )e for . *ach of these s a s mp"e . >e2u res .

The e6tens )e o)er"ap of the $ands ma/es the accurate measurement of the r ntens t es and pos t ons mposs $"e. The component $ands ha)e $een art f c a""+ narro3ed so that the ntens t es and pos t ons can $e measured. part"+ o)er"app n!: $ands. f the $ands 3ere narro3er. 3here >5 s the reso"ut on0enhanced s !na". n # ndo3 9. super mposed on the negative of ts second der )at )e n !reen:. the s !na"0to0no se rat o s de!raded.Resolution enhancement F !ure 9 sho3s a spectrum that cons sts of se)era" poor"+0reso")ed 8that s. (ere use can $e made of resolution enhancement a"!or thms to art f c a""+ mpro)e the apparent reso"ut on of the pea/s. (ere's ho3 t 3or/s. s !na"0 to0no se de!radat on. the component bands are narrowed so that the intensities and positions can be measured. . that s. The opt mum cho ce depends upon the 3 dth. It s "eft to the user to se"ect the 3e !ht n! factor / 3h ch ! )es the $est trade0off $et3een reso"ut on enhancement. shape. or 32/L for pea/s of Lorent7 an shape. %ne of the s mp"est such a"!or thms s $ased on the 3e !hted sum of the or ! na" s !na" and the ne!at )e of ts second der )at )e. 3n the resulting signal. and $ase" ne undershoot. @ s the or ! na" s !na". and d ! t 7at on nter)a" of the s !na". a computer0!enerated pea/ 83 th a Lorent7 an shape: n red. and / s a user0se"ected 3e !ht n! factor. a reasona$"e )a"ue for / s 32/2= for pea/s of Gauss an shape. The f !ure sho3s. A resolution enhancement algorithm has been applied to the signal on the left to artificially improve the apparent resolution of the peaks. right. Figure 9. e)en thou!h the s !na"0to0no se rat o s )er+ !ood. (o3e)er. Th n!s 3ou"d $e eas er f the $ands 3ere more comp"ete"+ reso")ed. 8?s a start n! po nt. 3here 3 s the num$er of data po nts n the ha"f03 dth of the component pea/s:. The resu"t of the app" cat on of th s a"!or thm s sho3n on the r !ht n F !ure 9. @'' s the second der )at )e of @.

so that +ou can e6per ment 3 th a"" the )ar a$"es at 3 "" and tr+ out th s techn 2ue on +our o3n s !na"s:. *esolution enhancement in atlab. Video"a$ reso"ut on0enhancement m0f "es 89L M$+tes:. the 2nd der )at )e factor 8Factor 9: s ad5usted.7 p. the area under the pea/: $ecause the tota" area under the cur)e of the der )at )e of a pea/0shaped s !na" s 7ero 8the area under the ne!at )es "o$es cance"s the area under the pos t )e "o$es:. >e2u res .+te ) deo (ResEnhance3. The s !na" cons sts of four o)er"app n!. the ar!uments k2 and k4 are 2nd and <th der )at )e 3e !ht n! factors. of the pea/. poor"+0reso")ed Lorent7 an $ands. and an ncrease n he !ht. 3hereas th s reso"ut on enhancement techn 2ue can not $e e6pected to do that. sho3n n # ndo3 2. Th s 3or/s $est 3 th Lorent7 an0shaped pea/sB 3 th Gauss an0shaped pea/s.k4.wmv) demonstrates the . nc"udes th s s mp"e reso"ut on0enhancement a"!or thm. It can hand"e s !na"s of ) rtua""+ an+ "en!th. then the smooth . Th s 9=0second. The resu"t. and SmoothWidth s the 3 dth of the $u "t0 n smooth.K .at"a$ funct on enhance. the reso"ut on enhancement s "ess dramat c 8on"+ a$out"a$ L. The Interact )e >eso"ut on *nhancement for . Aote: ?nother techn 2ue that can ncrease the reso"ut on of o)er"app n! pea/s s decon)o"ut on. 3here signal s the or ! na" s !na" )ector. Decon)o"ut on of the $roaden n! funct on from the $roadened pea/s s n pr nc p"e capa$"e of e6tract n! the under"+ n! pea/s shapes. " m ted on"+ $+ the memor+ n +our computer.The second der )at )e s amp" f ed 8$+ mu"t p"+ n! t $+ an ad5usta$"e constant: so that the ne!at )e s des of the n)erted second der )at )e 8from appro6 mate"+ O H 0 to 900 and from O H 9=0 to 2=0: are a m rror ma!e of the s des of the or ! na" pea/ o)er those re! ons. 9. In th s 3a+.m. +ou ma+ do3n"oad a set of .m 8ctr"0c" c/ on th s " n/ to nspect the code: uses a s" !ht"+ more ad)anced a"!or thm that e6tends the a$o)e approach $+ add n! n a sma"" amount of the <th der )at )e of the s !na": > H @ 0 /2@'' C /<@'''' Th s funct on has the form: Enhancedsignal=enhance(signal. s a su$stant a" 8a$out =0D: reduct on n the 3 dth. ?n nterest n! propert+ of th s procedure s that t does not chan!e the tota" pea/ area 8that"a$. and the smooth 3 dth: 3h "e o$ser) n! the effect on the s !na" output d+nam ca""+. 3 th ad5usta$"e 3e !ht n! factor and der )at )e smooth n! 3 dth. 3hen the or ! na" pea/ s added to the n)erted second der )at )e. 8If +ou ha)e access to .k2. <th der )at )e factor /<. The user0def ned . F rst. >eso"ut on*"a$ has s" ders that a""o3 +ou to ad5ust the reso"ut on enhancement parameters 82nd der )at )e factor /2. 3h ch s app" ca$"e 3hen the $roaden n! funct on respons $"e for the o)er"ap of the pea/s s /no3n. the t3o s !na"s 3 "" appro6 mate"+ cance" out n the t3o s de re! ons $ut 3 "" re nforce each other n the centra" re! on 8from O H 900 to"a$ nteract )e reso"ut on enhancement funct on Interact )e>es*nhance. SP*&T>-.SmoothWidth). The reso"ut on0 enhanced s !na" s returned n the )ector Enhancedsignal.. then the <th der )at )e factor 8Factor 2: s ad5usted.

!he signal on the left (x 7 time2 y 7 voltage).. The power spectrum s a s mp"e 3a+ of sho3 n! the tota" amp" tude at each of these fre2uenc esB t s ca"cu"ated as the s2uare root of the sum of the s2uares of the coeff c ents of the s ne and cos ne components. It s often usefu" to descr $e the amp" tude and fre2uenc+ of such per od c components e6act"+. The Four er transform s s mp"+ the coeff c ents of these s ne and cos ne components. as the nterfero!ram formed $+ p"ott n! the detector s !na" )s m rror d sp"acement n a scann n! . short pu"se of rad o fre2uenc+ ener!+ produces a free nduct on deca+ s !na" that s the Four er transform of the resonance spectrum.>:. 3here D6 s the nter)a" $et3een ad5acent 60a6 s )a"ues and n s the tota" num$er of po nts. !he smaller peak at '#% .-.. " /e a s ne 3a)e. suggesting that much of the noise is caused by stray pick-up from the 8% .-) probably comes from the same source. The second po nt 86H9: corresponds to the fundamenta" fre2uenc+. The 60a6 s s the harmonic number. n/2 t mes the fundamenta" fre2uenc+ fH9/nD6. In Four er transform nuc"ear ma!net c resonance spectroscop+ 8FTA. 3h ch s a 3a+ of e6press n! a s !na" as a sum of s ne and cos ne 3a)es.. It can $e sho3n that an+ ar$ trar+ d screte"+ samp"ed s !na" can $e descr $ed comp"ete"+ $+ the sum of a f n te num$er of s ne and cos ne components 3hose fre2uenc es are 0. Harmonic analysis and the Fourier Transform Some s !na"s e6h $ t per od c components that repeat at f 6ed nter)a"s throu!hout the s !na". e6c tat on of the samp"e $+ an ntense. The concept of the Four er transform s n)o")ed n t3o )er+ mportant nstrumenta" methods n chem str+.. the Four er transform of the spectrum s measured d rect"+ $+ the nstrument. 3hether or not the data appear per od c. chae"son nterferometer. . In $oth cases the spectrum s reco)ered $+ n)erse Four er transformat on of the measured s !na". etc. which was expected to contain a single peak.3 dth 8Smooth: s ad5usted.. Figure 10. In Four er transform nfrared spectroscop+ 8FTI>:. (armon c ana"+s s s con)ent ona""+ $ased on the 5ourier transform. ?n e6amp"e of a pract ca" app" cat on of the use of the po3er .power line.9. and f na""+ the Factor 2 s t3ea/ed a!a n. t s poss $"e to ana"+7e an+ ar$ trar+ set of data nto per od c components. ?ctua""+. The f rst po nt 86H0: s the 7ero0fre2uenc+ 8constant: component. is clearly very noisy.2.(the second harmonic of 8% .-) shows a strong component at 8% . ? s !na" 3 th n po nts ! )es a po3er spectrum 3 th on"+ 8n/2:C9 po nts. the ne6t po nt to t3 ce the fundamenta" fre2uenc+. !he po er spectrum of this signal (x-axis 7 fre+uency in .

m:. . Th s pro!ram s usefu" for teach n! and demonstrat n! the po3er spectra of d fferent t+pes of s !na"s and the effect of s !na" durat on and samp" n! rate. s" ders for rea" t me contro". In such cases the un ts of the 60a6 s of the po3er spectrum are s mp"+ the rec proca" of the un ts of the 60a6 s of the or ! na" s !na" 8e. #hen Sound s"a$ has $u "t0 n funct ons for comput n! the Four er transform 8FFT and IFFT: and the po3er spectrum"a$.!. s" ders contro" the !"o$a" )ar a$"es f9 and f2 3h ch are used to contro" the fre2uenc+. • The Signal s" der se"ects from 90 d fferent pre0 pro!rammed s !na"sI. The *ate s" der contro"s the samp"e rate 8(7:. nm09 for a s !na" 3hose 60a6 s s n nm:. and aud o 3a)eform output. +ou can ctr"0c" c/ here to do3n"oad the PIP f "e EPo3erSpectrumDemo99. The )+ and ). each t me a s" der s mo)ed. Interactive Power S ectrum Demo for !atlab Th s s d ! ta" s !na" !enerator 8s mu"ator: for . the s !na" 8 n the )ar a$"e E+E: s sent to the # ndo3s #?F* aud o de) ce. • The %ime s" der contro"s the tota" durat on of s !na" 8seconds:.ore !enera""+. . 3 th po3er spectrum d sp"a+.7 pE. 8@ou can chan!e the s !na" def n t ons n >edra3Spectrum. the s !na" s a t me0ser es s !na" 3 th t me as the ndependent )ar a$"e.T::. such as an opt ca" spectrum. e)en 3hen Sound s %FF. 3h ch a"so nc"udes support n! funct ons and se"f0conta ned demos to sho3 ho3 t 3or/s. • The Sound s" der has on"+ t3o sett n!s: %FF 8do3n: and %A 8up:. t me. In the e6amp"e sho3n here. • • If +ou are ) e3 n! th s document on0""a$ F "e *6chan!e. SP*&T>-. nc"udes a po3er spectrum funct"a$ L. t s a"so poss $"e to compute the Four er transform and po3er spectrum of an+ s !na". >e2u res . and/or amp" tude of the d fferent pre0pro!rammed s !na"s as descr $ed $e"o3. I The 90 pre0pro!rammed s !na"s are: S ne 3a)e of fre2uenc+ f9 8(7: and phase f2B S ne 3a)e $urst of fre2uenc+ f9 8(7: and durat on f2B S2uare 3a)e of fre2uenc+ f9 8(7: and phase f2B Sa3tooth 3a)e of fre2uenc+ f98(7:B <<0 (7 carr er amp" tude modu"ated $+ s ne 3a)e of fre2uenc+ f9 8(7: and amp" tude f2B <<0 (7 carr er fre2uenc+ modu"ated $+ s ne 3a)e of fre2uenc+ f9 8(7: and amp" tude f2B S ne 3a)e of fre2uenc+ f9 8(7: modu"ated $+ Gauss an pu"se of 3 dth f2B >andom 3h te no seB S ne 3a)e of fre2uenc+ f9 8(7: and amp" tude f2 p"us random 3h te no seB S ne 3a)e s3eep from 0 to f9 8(7:.spectrum as a d a!nost c too" s sho3n n F !ure 90. as 3e"" as for3ard and re)erse Four er transformat on. 3here the ndependent )ar a$"e m !ht $e )o"ts. #hen Sound s %FF. @ou can a"so do3n"oad t from the . . 3here the ndependent )ar a$"e m !ht $e 3a)e"en!th or 3a)enum$er. no sound s p"a+ed 3hen a s" der s mo)ed. or an e"ectrochem ca" s !na". (o3e)er.=. c" c/ n! on the Sound s" der p"a+s the s !na" once for each c" c/. phase.

?"thou!h th s seems to $e a round0a$out method. Th s funct on can $e used to create )er+ !enera" t+pe of f "ters and smooth n! funct ons. Figure 11. F rst. .Convolution &on)o"ut on s an operat on performed on t3o s !na"s 3h ch n)o")es mu"t p"+ n! one s !na" $+ a de"a+ed )ers on of another s !na". the Four er transform of each s !na" s o$ta ned. !he resulting convoluted spectrum (bottom center) shows that the two lines near x7''% and '#% will not be resolved but the line at x7&% will be partly resolved. !he 9aussian function has already been rotated so that its maximum falls at x7%. and repeat n! the process for d fferent de"a+s. nc"udes con)o"ut on and auto0corre"at on 8se"f0con)o"ut on: funct ons. The e6amp"e of F !ure 99 sho3s ho3 t can $e used to pred ct the $roaden n! effect of a spectrometer on an atom c " ne spectrum. such as s" d n!0a)era!e and tr an!u"ar smooths. . SP*&T>"a$ has a $u "t0 n funct on for con)o"ut on: con). t turns out to $e faster then the sh ft0and0mu"t p"+ a"!or thm 3hen the num$er of po nts n the s !na" s "ar!e. !onvolution is used here to determine how the atomic line spectrum in "indow ' (top left) will appear when scanned with a spectrometer whose slit function (spectral resolution) is described by the 9aussian function in "indow # (top right). In pract ce the ca"cu"at on s usua""+ performed $+ mu"t p" cat on of the t3o s !na"s n the Four er doma n. Then the t3o Four er transforms are mu"t p" ed $+ the ru"es for comp"e6 mu"t p" cat on and the resu"t s then n)erse Four er transformed. &on)o"ut on can $e used as a )er+ po3erfu" and !enera" a"!or thm for smooth n! and d fferent at on. nte!rat n! or a)era! n! the product. &on)o"ut on s a usefu" process $ecause t accurate"+ descr $es some effects that occur 3 de"+ n sc ent f c measurements. such as the nf"uence of a "o30pass f "ter on an e"ectr ca" s !na" or of a spectrometer on the shape of a spectrum.

%.e. Figure 12. !he signal in the bottom left is the result of deconvoluting the derivative spectrum (top right) from the original spectrum (top left). 3n this example. the convolution series is seen to be (#. "indow # (top right) is the first derivative of this spectrum produced by an (unknown) algorithm in the software supplied with the spectrometer. for e6amp"e. "indow ' (top left) is a uv-visible absorption spectrum recorded from a commercial photodiode array spectrometer (0-axis< nanometers2 1-axis< milliabsorbance). !he response function ("indow #. center) is usually either calculated on the basis of some theoretical model or is measured experimentally as the output signal produced by applying an impulse (delta) function to the input of the system. -#. with its maximum at x7%. >xpressed in terms of the smallest whole numbers. . !he result (right) shows a closer approximation to the real shape of the peaks2 however. "econvolution is used here to remove the distorting influence of an exponential tailing response function from a recorded signal ("indow '.. to re)erse the s !na" d stort on effect of an e"ectr ca" f "ter or of the f n te reso"ut on of a spectrometer. A different application of the deconvolution function is to reveal the nature of an unknown data transformation function that has been applied to a data set by the measurement instrument itself. computat ona": 3a+ to re)erse the resu"t of a con)o"ut on occurr n! n the ph+s ca" doma n. left) that is the result of an unavoidable :. The pract ca" s !n f cance of decon)o"ut on s that t can $e used as an art f c a" 8 . :otating and expanding it on the x-axis makes the function easier to see (bottom right). !his therefore must be the convolution function used by the differentiation algorithm in the spectrometer=s software. -'. 5igure '$. the signal-to-noise ratio is unavoidably degraded. the decon)o"ut on of one s !na" from another s usua""+ performed $+ dividing the t3o s !na"s n the Four er doma n. is deconvoluted from the original signal . ('. !his simple example of ?reverse engineering? would make it easier to compare results from other instruments or to duplicate these result on other e+uipment. low-pass filter action in the electronics. !he response function.Deconvolution Decon)o"ut on s the con)erse of con)o"ut on n the sense that d ) s on s the con)erse of mu"t p" cat on.T3o e6amp"es of the app" cat on of decon)o"ut on are sho3n n F !ures 92 and 9. In fact.

!he power spectrum is expanded in the 0 and 1 directions to show more clearly the low-fre+uency region ("indow '. Figure 14. . . $ecause n decon)o"ut on the under"+ n! pea/ shape s un/no3n $ut the $roaden n! funct on s assumed to $e /no3nB 3hereas n terat )e "east0s2uares cur)e f tt n! the under"+ n! pea/ shape s assumed to $e /no3n $ut the $roaden n! funct on s un/no3n. the Fourier filter function can be used to delete the higher harmonics and to reconstruct the signal from the first #% harmonics. The user tr es to f nd a cut0off fre2uenc+ that 3 "" a""o3 most of the no se to $e e" m nated 3h "e not d stort n! the s !na" s !n f cant"+. SP*&T>-. second from left) shows that high-fre+uency components dominate the signal. Fourier filter The Four er f "ter s a t+pe of f "ter n! or smooth n! funct on that s $ased on the fre2uenc+ components of a s !na". Note. but its po er spectrum ("indow #. ?n e6amp"e of the app" cat on of the Four er f "ter s ! )en n F !ure 9<.at"a$ has $u "t0 n funct on for decon)o"ut on: decon).The term Edecon)o"ut onE s somet mes a"so used for the process of reso") n! or decompos n! a set of o)er"app n! pea/s nto the r separate components $+ the techn 2ue of terat )e "east0s2uares cur)e f tt n! of a putat )e pea/ mode" to the data set. "orking on the hypothesis that the components above the #%th harmonic are noise.#hen app"+ n! decon)o"ut on to e6per menta" data. Th s can $e contro""ed $ut not comp"ete"+ e" m nated $+ smooth n! and $+ constra n n! the decon)o"ut on to a fre2uenc+ re! on 3here the s !na" has a suff c ent"+ h !h s !na"0to0 no se rat o. The assumpt on s made here that the fre2uenc+ components of the s !na" fa"" predom nant"+ at "o3 fre2uenc es and those of the no se fa"" predom nant"+ at h !h fre2uenc es. nc"udes a decon)o"ut on funct on. a )er+ ser ous s !na"0to0no se de!radat on common"+ occurs. to remo)e the effect of a /no3n $roaden n! or "o30pass f "ter operator caused $+ the e6per menta" s+stem. then n)erse transform n! the resu"t. $ecause the h !h fre2uenc+ components of the $roaden n! operator 8the denom nator n the d ) s on of the Four er transforms: are t+p ca""+ )er+ sma"". ?n+ no se added to the s !na" $+ the s+stem after the $roaden n! or "o30pass f "ter operator 3 "" $e !reat"+ amp" f ed 3hen the Four er transform of the s !na" s d ) ded $+ the Four er transform of the $roaden n! operator. It 3or/s $+ ta/ n! the Four er transform of the s !na". resu"t n! n a !reat amp" f cat on of h !h fre2uenc+ no se n the resu"t n! decon)o"uted s !na". right). The process s actua""+ conceptua""+ d st nct from decon)o"ut on. then cutt n! off a"" fre2uenc es a$o)e a user0spec f ed " m t. !he signal at the far left seems to be only random noise.

or . m "" sec.m s a more f"e6 $"e Four er f "ter that can ser)e as a "o3pass. or m crosecB 'centerfre2uenc+' and 'fre2uenc+3 dth' are the center fre2uenc+ and 3 dth of the f "ter n (7.mode) 3here + s the t me0ser es s !na" )ector. In chromato!raph c ana"+s s one often has the pro$"em of measur n! the the area under . Integration and ea" area measurement The s+m$o" c nte!rat on of funct ons and the ca"cu"at on of def n te nte!ra"s are top cs that are ntroduced n e"ementar+ &a"cu"us"a$ funct on FouF "ter. h !hpass. The s mp"e numer c nte!rat on of a d ! ta". and cut0off rate: 3h "e o$ser) n! the effect on the s !na" output d+nam ca""+. >e2u res . The reason for th s s that pea/ area s "ess sens t )e to the nf"uence of pea/ $roaden n! 8d spers on: mechan sms. 3h ch ar se from man+ sources. 3here the area under each pea/ or mu"t p"et s proport ona" to the num$er of e2u )a"ent h+dro!ens respons $"e for that pea/. M(7. 'samp" n!t me' s the tota" durat on of samp"ed s !na" n sec. Pea/ area measurements are )er+ mportant n chromato!raph+. and more uns+mmetr ca". Set mode H 0 for $and0pass f "ter. SP*&T>-. FouF "ter returns the f "tered s !na" n r+. Quant tat on n chromato!raph+ s customar "+ carr ed out on the $as s of pea/ area rather than pea/ he !ht measurement. mode H 9 for $and0re5ect 8notch: f "ter. Th s s a common"+ used method n proton A.!he result (far right) shows the signal contains two bands at about x7#%% and x7$%% that are totally obscured by noise in the original signal. The user0def ned . Therefore pea/ area measurements are often found to $e more re" a$"e than pea/ he !ht measurement. . 3e"" reso")ed:.!.(7. or $andre5ect 8notch: f "ter 3 th )ar a$"e cut0off rate.ut th s 3or/s 3e"" on"+ f the pea/s are 3e"" separated from each other 8e. It can hand"e s !na"s of ) rtua""+ an+ "en!th. $roader."re# encywidth. The Interact )e Four er F "ter for .at"a$ funct on has s" ders that a""o3 +ou to ad5ust the Four er f "ter parameters 8center fre2uenc+. cause chromato!raph c pea/s to $ecome shorter. the he !ht of each of 3h ch s proport ona" to the area under that pea/.sam!lingtime.=. $andpass. If shape H 9.> spectroscop+. $ut ha)e " tt"e effect on the tota" area under the pea/.sha!e. These $roaden n! effects. f "ter 3 dth. 3 "" con)ert a ser es of pea/s nto a ser es of steps. respect )e"+B 'Shape' determ nes the sharpness of the cut0"re# ency. nc"udes a s mp"e Four er "o30pass f "ter funct on 3 th ad5usta$"e harmon c cut0off. " m ted on"+ $+ the memor+ n +our computer. The numer ca" nte!rat on of d ! t 7ed s !na"s f nds app" cat on n ana"+t ca" s !na" process n! ma n"+ as a method for measur n! the areas under the cur)es of pea/0t+pe s !na"s.!. the f "ter s Gauss anB as shape ncreases the f "ter shape $ecomes more and more rectan!u"ar. The pea/ area rema ns proport ona" to the tota" 2uant t+ of su$stance pass n! nto the detector. $+ S mpson's ru"e. e. (as the form ry=Fo Filter("a$ L.

%n"+ for the f rst0order po"+nom a" s the p"ot of O )s @ " near. and the t3o )ert ca" " nes. F !ure 9= sho3s a ser es of four computer0s+nthes 7ed Gauss an pea/s that a"" ha)e the same he !ht. a"thou!h )er+ ted ous to do $+ hand. desp te the poor reso"ut on. or a cu$ c 8@ H a C bO C cO2 C dO. The !raph sho3s a s mp"e stra !ht0" ne e6amp"e 8@ H a C bO:. 3 dth. ? po"+nom a" e2uat on e6presses the dependent )ar a$"e @ as a po"+nom a" n the ndependent )ar a$"e O. Curve fitting A# $inear $east S%uares The o$5ect )e of cur)e f tt n! s to f nd a mathemat ca" e2uat on that descr $es a set of data and that s m n ma""+ nf"uenced $+ random no se. @ s a " near funct on of the parameters a. and area.the cur)e of the pea/s 3hen the+ are not 3e"" reso")ed or are super mposed on a $ac/!round.c. The "eft and r !ht $ounds of the pea/ are usua""+ ta/en as the )a""e+s 8m n ma: $et3een the pea/s. $ut the separat on $et3een the pea/s on the r !ht s nsuff c ent to ach e)e comp"ete reso"ut on. -s n! th s method t s poss $"e to est mate the area of the second pea/ n th s e6amp"e to an accurac+ of a$out 0. Figure 15. using the perpendicular drop method. and d. 3here a s the intercept and b s the slope:. not $ecause the p"ot of O )s @ s " near. The c"ass ca" 3a+ to hand"e th s pro$"em s to dra3 t3o )ert ca" " nes from the "eft and r !ht $ounds of the pea/ do3n to the 60a6 s and then to measure the tota" area $ounded $+ the s !na" cur)e. @eak area measurement for overlapping peaks. or h !her0order po"+nom a". a 3e""0/no3n mathemat ca" procedure for f nd n! the coeff c ents of po"+nom a" e2uat ons that are a E$est f tE to a set of O.D and the second and th rd pea/s to an accurac+ of $etter than <D. In a"" these cases. The $"ue dots are the data po nts. The "east0s2uares a"!or thm computes the )a"ues of a 8 ntercept: and b 8s"ope: of the stra !ht .@ data. Th s s often ca""ed the the perpend cu"ar drop method. or a 2uadrat c 8@ H a C bO C cO2:.b. the 60a6 s 8+H0 " ne:. For e6amp"e. a sma"" set of data 3here O H concentrat on and @ H nstrument read n!..:. a"so ca""ed Epo"+nom a" "east s2uaresE. The dea s ""ustrated for the second pea/ from the "eft n F !ure 9=. The most common approach s the E" near "east s2uaresE method. and t s an eas+ tas/ for a computer. !his s 3h+ th s s ca""ed E" nearE "east0s2uares f t. for e6amp"e as a stra !ht " ne 8@ H a C bO. 8Somet mes th s t+pe of cur)e f tt n! s ca""ed Ecur) " nearE:.

The "east s2uares procedure a"so ca"cu"ates >2. C bO. a"so ca""ed the correlation coefficient.0000 the $etter. Th s s a fundamenta""+ non0" near pro$"em $ecause @ s a non0" near funct on of the parameter b. sho3n $+ the !reen so" d " ne n the f !ure. the d fferences $et3een the + )a"ues n the or ! na" data and the + )a"ues computed $+ the f t e2uat on. "ess than that 3hen the f t s mperfect. the po nts n F !ure 9L are from the s mu"at on of an e6ponent a" deca+ 8O H t me. (o3e)er. The po nts do not a"" fa"" on the stra !nt " ne $ecause of random no se n the data. +ou'd !et f )e s" !ht"+ d fferent ans3ers. and then the "east0s2uares method can $e app" ed to the resu"t n! " near e2uat on. (ad there $een a "ar!er num$er of po nts n th s data set. for e6amp"e a -0shaped or an S0shaped cur)e. >2 s e6act"+ 9. For th s set of data." ne that s a "east0s2uares $est f t to the data po nts. $+ ta/ n! the natura" "o! of $oth s des of the e2uat on. a H 9 and b H 09. from 3h ch )a"ues of a and b are eas "+ ca"cu"ated. so the presense of the no se caused th s part cu"ar measurement of s"ope to $e off $+ a$out 2D. +ou'd !et the e6act same ans3er e)er+ t me:. If the res dua"s are comp"ete"+ random and sho3 no s+stemat c )ar at on. 3here a s the @0)a"ue at O H 0 and b s the deca+ constant. then the f t s as !ood as poss $"e for that "e)e" of no seB $ut f the res dua" p"ot sho3s a re!u"ar cur)e.9JJ9L O:. and b. 8If +ou 3ere to ! )e th s set of data to f )e d fferent peop"e and as/ them to est mate the $est0f t " ne ) sua""+. The $est f t e2uat on. In th s part cu"ar e6amp"e.!. the EtrueE )a"ue of the s"ope s 90 and of the ntercept s 7ero. Least0s2uares $est f ts can $e ca"cu"ated $+ some hand0he"d ca"cu"ators. the ca"cu"ated )a"ues of s"ope and ntercept 3ou"d pro$a$"+ ha)e $een $etter:.999. the "east0s2uare method s more o$5ect )e and eas er to automate. The c"oser to 9. the s"ope s 9. ? s mp"e 3a+ to e)a"uate a cur)e f t s to "oo/ at a p"ot of the Eres dua"sE. ta/ n! the "o! or the rec proca" of the data:. $ut t +ou !a)e the data set to f )e d fferent computer pro!rams. that s a H 0. )er+ . s @ H 0. "n8@: s a " near funct on of "n8a. spreadsheets. the coefficient of determination. An exponential least-s+uares fit (solid line) applied to a noisy data set (points) in order to estimate the decay constant.9J9K and b H 00. 3e o$ta n "n8@: H "n8a.0000 3hen the f t s perfect. For e6amp"e. ?"thou!h t s poss $"e to est mate the $est0f t stra !ht " ne $+ ) sua" est mat on and a stra !hted!e. 8In th s e6amp"e. @ H s !na" ntens t+: that has the mathemat ca" form @ H a e6p8bO:. %ransforming non'linear relationships In some cases a fundamenta""+ non0" near re"at onsh p can $e transformed nto a form that s amena$"e to po"+nom a" cur)e f tt n! $+ means of a coord nate transformat on 8e. and ded cated computer pro!rams 8see $e"o3 for deta "s:. so t can $e f t $+ the "east s2uares method.9JJ9L. 3h ch s an nd cator of the E!oodness of f tE.K92L and the ntercept s 0.9J9K e6p80 0. In th s e2uat on. t ma+ mean that another f tt n! funct on 3ou"d $e more appropr ate.

and cB the pea/ he !ht s ! )en $+ %xp(a--*(b/(2*-)). In pract ce th s means that an+ non07ero $ase" ne must $e su$tracted from the data set $efore app"+ n! th s method. 3hen f tt n! an+th n! $ut a stra !ht " ne 3as d ff cu"t. . The most mportant re2u rement s that the mode" $e !ood. The resu"t n! data set.35/03/(s0+t(2)*s0+t(--)). ta/ n! the natura" "o! of the data 8upper r !ht: produces a para$o"a that can $e f t 3 th a 2uadrat c "east0s2uares f t 8sho3n $+ the $"ue " ne n the "o3er "eft:. that s. +ou ma+ ha)e to resort to the more d ff cu"t Aon0" near Iterat )e &ur)e F tt n! method:. and the or ! na" Gauss an pea/ s !na" must ha)e a 7ero $ase" ne.2). the pea/ pos t on $+ -b/(2*-). b. ho3e)er. Aote: n order for th s method to 3or/ proper"+. and 3 dth: can $e ca"cu"ated from the three 2uadrat c coeff c ents a. 8%n"+ a fe3 non0" near re"at onsh ps can $e hand"ed th s 3a+. that s. From the three coeff c ents of the 2uadrat c f t 3e can ca"cu"ate much more accurate )a"ues of the Gauss an pea/ parameters. %ther e6amp"es of non0" near re"at onsh ps that can $e " near 7ed $+ coord nate transformat on nc"ude the "o!ar thm c 8@ H a "n8bO:: and po3er 8@HaOb: re"at onsh ps. sho3n $+ the red po nts n the upper "eft. Thus. It s st "" used toda+ to e6tend the ran!e of funct ona" re"at onsh ps that can $e hand"ed $+ common " near "east0s2uares rout nes. ?n e6amp"e s sho3n n the f !ure on the "eft.ethods of th s t+pe used to $e )er+ common $ac/ n the da+s $efore computers.c"ose to the e6pected )a"ues. 3h ch 3or/s for data sets 3 th 7eros and ne!at )e num$ers and a"so for data 3 th mu"t p"e o)er"app n! pea/s. ?nother e6amp"e of the use of transformat on to con)ert a non0" near re"at onsh p nto a form that s amena$"e to po"+nom a" cur)e f tt n! s the use of the natura" "o! 8"n: transformat on to con)ert a Gauss an pea/. $ecause the under"+ n! mode"s are not a"3a+s /no3n 3 th certa nt+. (o3e)er. ma6 mum pos t on. us n! th s scr pt. that the e2uat on se"ected for the mode" accurate"+ descr $es the under"+ n! $eha) or of the s+stem 8e6cept for no se:. e)en n the presence of su$stant a" random no se. must tend to 7ero far from the pea/ center. %ften that s the most d ff cu"t aspect. The p"ot n the "o3er r !ht sho3s the resu"t n! Gauss an f t 8 n $"ue: d sp"a+ed 3 th the or ! na" data 8red po nts:. t s poss $"e to !et !ood est mates of the parameters of the under"+ n! e2uat on. . s the non0" near terat )e cur)e f tt n! method:. too fe3 for accurate measurement of pea/ he !ht. has on"+ L data po nts on the pea/. The s !na" s a Gauss an pea/ 3 th a pea/ he !ht of e6act"+ 900 un ts. To f t an+ ar$ trar+ custom funct on. and the pea/ ha"f03 dth $+ 2. 3h ch can $e f t 3 th a second order po"+nom a" 82uadrat c: funct on 8+ H a C bx C c62:. ?"" three parameters of the Gauss an 8he !ht.9 un ts on the 60a6 s. The ma6 mum of those L po nts s at 6HJK and has an amp" tude of 9=. pos t on. 8? more !enera" approach to f tt n! Gauss an pea/s. a pea/ pos t on of 900 un"a$. nto a para$o"a of the form 062. The accurac+ s " m ted on"+ $+ the no se n the data 8a$out 9D n th s e6amp"e:. and 3 dth. 3h ch has the fundamenta" funct ona" form e6p8062:. and a ha"f03 dth of 900 un ts. not that c"ose to the true )a"ues of pea/ pos t on and he !ht 8900:. the data set must not conta n an+ 7eros or ne!at )es po nts. sho3n at the $ottom of the f !ure. Th s f !ure 3as created n . $ut t s samp"ed on"+ e)er+ .

@ou can a"so do3n"oad the spreadsheets p ctured a$o)e.umd.ods:. the LIA*ST funct on n $oth *6ce" and %pen%ff ce &a"c can $e used to compute po"+nom a" and other cur) " near "east0s2uares f ts.odt: and a"so a )ers on for 2uadrat c 8para$o" c: f ts 8http://terpconnect. E>2E. &ur)e f tt n! a"!or thms t+p ca""+ accept a set of ar$ trar "+0spaced 60a6 s )a"ues and a correspond n! set of +0a6 s )a"ues.htm":. that automate the computat on of those e2uat ons and a"so p"ot the data and the $est0f t " ne.0000 f the f t s perfect and "ess than that f the f t s mperfect:: n$=$n m%er$o"$&.edu/~toh/spectrum/LeastS2uares.ethods n ?$sorpt on Spectroscop+ and *rror propa!at on n ?na"+t ca" &a" $rat on. n %pen%ff ce &a"c format. For the app" cat on to ana"+t ca" ca" $rat on.+ data set can $e computed us n! on"+ $as c ar thmet c.umd. there s no re2u rement that the 60a6 s nter)a" $et3een data po nts $e un form. . (ere are the re"e)ant e2uat ons for comput n! the s"ope and ntercept of the f rst0order $est0f t e2uat on. see &a" $rat on &ur)e F tt n! .Note: In the cur)e f tt n! techn 2ues descr $ed here and n the ne6t t3o sect ons. re2u r n! on"+ that +ou t+pe n 8or paste n: the 60+ data.*(ssr)ssy)$ "opular spreadsheets ha)e fac " t es for comput n! po"+nom a" "east0s2uares cur)e f ts. 3h ch s an nd cator of the E!oodness of f tE.y$data$!oints$ s m&$=$'& s my$=$'y s m&y$=$'&(y s m&2$=$'&(& mean&$=$s m&$)$n meany$=$s my$)$n slo!e$=$(n(s m&y$*$s m&(s my)$)$(n(s m&2$*$s m&(s m&$ interce!t$=$meany*(slo!e(mean&) ssy$=$'(y*meany)+2 ssr$=$'(y*interce!t*slo!e(&)+2 R2$=$. there are spec f c )ers ons of these spreadsheets that a"so ca"cu"ate the concentrat ons of the un/no3ns 8http://terpconnect. For e6amp"e. For some e6amp"es of app" cat ons to ana"+t ca" chem str+. ath details The "east0s2uares $est f t for an 6. There s one spreadsheet for " near f ts 8http://terpconnect. as s the assumpt on n man+ of the other s !na" process n! techn 2ues pre) ous"+ co)"s/&a" $rat on&ur)e. + H ntercept C s"opeI6. as 3e"" as the coeff c ent of determ nat on. 8 >2 s 9.

Th s method s 3 de"+ used n mu"t 03a)e"en!th techn 2ues such as d ode0arra+. . Definitions: ? H ana"+t ca" s !na" c9. For a 2uadrat c f t 8nH2:. etc. s e.+. etc.y. 3h ch s the s"ope of a p"ot of ? )s c.&)).: s d rect"+ proport ona" to concentrat on.+ data po nts n the )ectors E6E and E+E.0 f the f t s perfect and 7ero f there s no s+stemat c re"at on at a"" $et3een 6 and +. >2 s a"3a+s $et3een 7ero and one: t s 9. 2 for a 2uadrat c 8para$o"a: f t. f ts t to a po"+nom a" of order Epo"+orderE. for a"" n spec es. c H mo"ar concentrat on s9. c2 H spec es 9.&&).) s the 62 term 8EcE:.!olyval(coe". coe"(. and po3er re"at onsh ps. .-o%-. c. spec es 2. For a stra !ht0" ne f t 8nH9:.y. nc"udes "east0s2uares cur)e f tt n! for po"+nom a"s of order 9 throu!h =.&&. 8&tr"0c" c/ to ) e3 th s funct on:. ? 8such as a$sor$ance n a$sorpt on spectroscop+. and then us n! 66 rather than 6 to e)a"uate and p"ot the f t: !lot(&. e)a"uated at more f ne"+0d ) ded )a"ues of 6.-*r-).. 3hen the spectra of the components o)er"ap cons dera$"+. coe"(2) s the 6 term 8E$E: and coe"(3) s the constant term 8EaE:. and d sp"a+s the f t coeff c ents and the !oodness0of0f t measure >2 n the upper "eft corner of the !raph.&). w H num$er of 3a)e"en!ths at 3h ch s !na" s measured $ssumptions: The ana"+t ca" s !na". ? H ec The tota" s !na" s the sum of the s !na"s for each component n a m 6ture: ?tota" H ?c9 C ?c2 C .at"a$ has s mp"e $u "t0 n funct ons for "east0s2uares cur)e f tt n!: po"+f t and po"+)a". and ref"ectance n ref"ectance spectroscop+. etc. and automated scann n! spectrometers. 3a)e"en!th 2. The !raph on the "eft 3as !enerated $+ the user0def ned funct on p"otf t86. "o!ar thm c. samp"e 2.po"+order:. for e6amp"e "ity=!olyval(coe". The f t e2uat on can $e e)a"uated us n! the funct on po"+)a".s2 H samp"e 9. For e6amp"e.y*!olyval(coe". 8#hen the num$er of data po nts s sma"". then the coeff c ents for the "east0s2uares f t are ! )en $+ coe"=!oly"it(&.-*r-):..&.y. The proport ona" t+ constant. coe"(. f +ou ha)e a set of 6. p"us e6ponent a". p"ots the data and the f t. Th s 3or/s for an+ order of po"+nom a" f t 8EnE:. Curve fitting B: Multicomponent Spectroscopy The spectroscop c ana"+s s of m 6tures. s H num$er of samp"es n H num$er of d st nct chem ca" spec es n the m 6ture.: s the s"ope 8E$E: and coe"(2) s the ntercept 8EaE:. +ou can !et a smoother p"ot of the f tted e2uat on. 3h ch accepts the data n the )ectors E6E and E+E. $+ def n n! &&=lins!ace(min(&). @ou can p"ot the data and the f tted e2uat on to!ether us n! the !lot funct on: !lot(&. 3h ch p"ots the data as $"ue c rc"es and the f tted e2uat on as a red " ne. re2u re spec a" ca" $rat on methods $ased on a t+pe of " near "east s2uares ca""ed mu"t p"e " near re!ress on. f"uorescence ntens t+ n f"uorescence spectroscop+. @ou can p"ot the res dua"s $+ 3r t n! !lot(&.-o%-.&). l2 H 3a)e"en!th 9.!olyval(coe".n). Four er transform.SP*&T>-. 3here EnE s the order of the po"+nom a" f t: n H 9 for a stra !ht0" ne f t. e H ana"+t ca" sens t ) t+ l9.

for a"" n spec es. . photon no se and detector no se:. f w R n. In most pract ca" app" cat ons.ut then the e2uat on can not $e so")ed $+ matr 6 n)ers on. If w H n. Then t fo""o3s that: ?l9Hec9. C . then 3e ha)e a s+stem of n e2uat ons n n un/no3ns 3h ch can $e so")ed $+ pre0mu"t p"+ n! $oth s des of the e2uat on $+ e-1. In a$sorpt on T 09 09 T 09 . Th s has the effect of ntroduc n! nto the so"ut on an add t ona" component 3 th a f"at spectrumB th s s referred to as T$ac/!round correct onU. e e s a s2uare matr 6 of order n. Th s set of " near e2uat ons s 3r tten compact"+ n matr 6 form: $ H e/ 3here $ s the w0"en!th )ector of measured s !na"s 8 .l9 cc.. Thus: T -1 T T -1 T T -1 T / H 8e e : e $ T In th s e6press on. then +ou measure ts spectrum $ and see/ to ca"cu"ate the concentrat on )ector /. th s s not a )er+ $ ! matr 6 to n)ert. n s t+p ca""+ on"+ 2 to =. F rst..ecause rea" e6per menta" spectra are su$5ect to random no se 8e.l2 cc2 C ec.. ? so"ut on can $e o$ta ned n th s case $+ pre0mu"t p"+ n! $oth s des of the e2uat on $+ the e6press on 8eTe:-1eT: -1 8e e: e $ H 8e e: e e/ H 8e e: 8e e:/ T -1 T . If +ou ha)e a samp"e so"ut on conta n n! un/no3n amounts of those spec es. the num$er of spec es. Second. calibration. the s !na" spectrum: of the m 6ture.l2 cc. T3o e6tens ons of the &LS method are common"+ made.l9 cc2 C ec. a co"umn of 9s s added to the e matr 6..l9 cc9 C ec2. In !enera". . the so"ut on 3 "" $e more prec se f s !na"s at a "ar!er num$er of 3a)e"en!ths are used. Th s method s app" ca$"e to the 2uant tat )e ana"+s s of a m 6ture of spec es 3hen the spectra of the nd ) dua" spec es are /no3n. . ?l2Hec9. t s common to perform a weighted "east s2uares so"ut on that de0 emphas 7es 3a)e"en!th re! ons 3here prec s on s poor: T -1 T / = 8e V e : e V $ 3here V s an w 6 w d a!ona" matr 6 of )ar ances at each 3a)e"en!th. $ac/!round. In order to so")e the a$o)e matr 6 e2uat on for /. the matr 6 n)erse of e.!. the num$er of 3a)e"en!ths w must $e e2ua" to or !reater than the num$er of spec es n. and so on for a"" 3a)e"en!ths. and us n! the propert+ that an+ matr 6 t mes ts n)erse s un t+: /He $ ..e. no matter ho3 man+ 3a)e"en!ths are used. e s the n 6 w rectan!u"ar matr 6 of the /no3n e0)a"ues for each of the n spec es at each of the w 3a)e"en!ths. n order to account for the fact that the prec s on of measurement ma+ )ar+ 3 th 3a)e"en!th. the more 3a)e"en!ths are used the more effect )e"+ the random no se 3 "" $e a)era!ed out 8a"thou!h t 3onSt he"p to use 3a)e"en!ths n spectra" re! ons 3here none of the components produce ana"+t ca" s !na"s:. $ecause the e matr 6 s a w 6 n matr 6 and a matr 6 n)erse e6 sts on"+ for s2uare matr ces..e.easurement of the spectra of /no3n concentrat ons of the separate spec es a""o3s the r ana"+t ca" sens t ) t+ e at each 3a)e"en!th to $e determ ned. and " !ht scatter n!.ut the 2uant t+ 8e e: 8e e: s a matr 6 t mes ts n)erse and s therefore un t+.l2 cc9 C ec2./lassical 0east S1uares 2/0S. n order to account for $ase" ne sh ft caused $+ dr ft. and / s the n0"en!th )ector of concentrat ons of a"" the spec es. The determ nat on of the opt mum 3a)e"en!th re! on must usua""+ $e determ ned emp r ca""+. C .

If the num$er of samp"es n the ca" $rat on set s !reater than the num$er of 3a)e"en!ths. 3h ch are s m "ar to $ut not n the ca" $rat on set.m demonstrates the c"ass ca" "east s2uares procedure for a s mu"ated a$sorpt on spectrum of a =0component m 6ture. the notat on s a " tt"e d fferent: transpose of a matr 6 1 s 12. %pen%ff ce &a"c. 3here the prec s on of measurement s poor n spectra" re! ons 3here the a$sor$ance s )er+ h !h 8and the " !ht "e)e" and s !na"0to0no se rat o therefore "o3:. Thus the so"ut on to the c"ass ca" "east s2uares method a$o)e s 3r tten 3 = !. for e6amp"e *6ce". ILS s a method that can $e used to measure the concentrat on of an ana"+te n samp"es n 3h ch the spectrum of the ana"+te n the samp"e s not /no3n $eforehand.l9 C ml2?s9. &n#erse 0east S1uares 2&0S. 3hose spectra are /no3n $ut 3hose concentrat ons n the m 6ture are un/no3n.. and s the w0 "en!th )ector of ca" $rat on coeff c ents. ""ustrated on the "eft.spectroscop+. and so on for a"" s samp"es.. calibration.. In matr 6 form /H$ 3here / s the s0"en!th )ector of concentrat ons of the ana"+te n the s samp"es. 3h ch must $e "ess than s:. #hereas the c"ass ca" "east s2uares method mode"s the s !na" at each 3a)e"en!th as the sum of the concentrat ons of the ana"+te t mes the ana"+t ca" sens t ) t+.ut atlab s rea""+ the natura" computer approach to mu"t component ana"+s s $ecause t hand"es a"" t+pes of matr 6 math so eas "+. (ere s an e6amp"e of a mu"t component ca" $rat on ca" $rat on performed n a spreadsheet en) ronment. In .l2 C ml2?"a$. the "east0s2uares so"ut on s: H 8$ $: $ / T 8Aote that $ $ s a s2uare matr 6 of s 7e w. Th s 3 "" ser)e as the calibration set. @ou measure the spectrum of each of the samp"es n th s ca" $rat on set and put these data nto a w 6 s matr 6 of measured s !na"s $. and matr 6 mu"t p" cat on s des !nated $+ I. suppose that +ou ha)e a set of standard samp"es that are t+p ca" of the t+pe of samp"e that +ou 3 sh to $e a$"e to measure and 3h ch conta n a ran!e of ana"+te concentrat ons that span the ran!e of concentrat ons e6pected to $e found n other samp"es of that t+pe.l2 C . In th s e6amp"e the dots represent the o$ser)ed spectrum of the m 6ture 83 th no se: and the f )e co"ored $ands represent the f )e components n the m 6ture. the num$er of 3a)e"en!ths. To!ether these data a""o3 +ou to ca"cu"ate the ca" $rat on )ector $+ so") n! the a$o)e e2uat on. .(1). @ou then measure the ana"+te concentrat ons n each of the samp"es $+ some re" a$"e and ndependent ana"+t ca" method and put those data nto a s0"en!th )ector of concentrat ons /.l2 C . for a"" w 3a)e"en!ths.(42*4)*42*13 3here 4 s the rectan!u"ar matr 6 of sens t ) t es at each 3a)e"en!th for each component The scr pt >e!ress onDemo.ost modern spreadsheets ha)e $as c matr 6 man pu"at on capa$ " t es and can $e used for mu"t component ca" $rat on.l9 C ml2?s2.l2 C ml2?s9. t s common to use the transm ttance T or ts s2uare T2 as 3e !ht n! factors. . Ao3. from the measured spectra of the samp"es: /H$ &"ear"+ th s 3 "" 3or/ 3e"" on"+ f the ana"+t ca" samp"es are s m "ar to the ca" $rat on set.. the n)erse of 1 s !. Th s ca" $rat on )ector can $e used to compute the ana"+te concentrat ons of other samp"es. cs2 H ml9?s2. $ s the w 6 s matr 6 of measured s !na"s at the w 3a)e"en!ths n the s samp"es. The $"ac/ " ne represents the E$est f tE to the o$ser)ed spectrum ca"cu"ated $+ the T -1 T . the n)erse "east s2uares methods use the re)erse approach and mode"s the ana"+te concentrat on c n each samp"e as the sum of the s !na"s ? at each 3a)e"en!th t mes ca" $rat on coeff c ents m that e6press ho3 the concentrat on of that spec es s re"ated to the s !na" at each 3a)e"en!th: cs9 H ml9?s9.

and therefore s t " /e"+ that near0 nfrared spectroscop+ shou"d do a prett+ !ood 5o$ of est mat n! the prote n content of s m "ar un/no3n samp"es. 3h ch can $e measured much more 2u c/"+ on 3heat paste preparat ons. Th s cont nues unt " the f tt n! error s "ess than the spec f ed error. In th s e6amp"e there are =0 ca" $rat on samp"es measured at L 3a)e"en!ths. In th s e6amp"e the concentrat ons of the f )e components are measured to an accurac+ of a$out 9D re"at )e 8" m ted $+ the no se n the o$ser)ed spectrum:. reference method. means that terat )e methods are more 3 de"+ used no3 than e)er $efore. the pro!ram s+stemat ca""+ chan!es one or more of the parameters and "oops $ac/ around to step . corre"ates to the r prote n content. $ut t s " m ted to s tuat ons 3here the dependent )ar a$"e can $e mode"ed as a po"+nom a" 3 th " near coeff c ents. The most !enera" 3a+ of f tt n! an+ mode" to a set of data s the terat )e method. Th s sounds " /e a $rute0force approach. The In)erse Least S2uares 8ILS: techn 2ue s demonstrated $+ th s scr pt and the !raph $e"o3. th s method 3as on"+ !rud! n!"+ app" ed.. . #e sa3 that n some cases a non0" near s tuat on can $e con)erted nto a " near one $+ a coord nate transformat on. 3here "ar!e num$ers of samp"es of a s m "ar pred cta$"e t+pe must $e ana"+7ed. Iterat )e methods proceed n the fo""o3 n! !enera" 3a+: 89: the operator se"ects a mode" for the dataB 82: f rst !uesses of a"" the non0" near parameters are madeB 8. n an+ case.: a computer pro!ram computes the mode" and compares t to the data set. coup"ed 3 th ad)ances n computer speed and a"!or thm eff c enc+. %ne popu"ar techn 2ue for do n! th s s ca""ed the Aelder-Bead Bodified 4implex. The purpose of th s ca" $rat on s to esta$" sh 3hether near0 nfrared ref"ectance spectroscop+. The /e+ s that the un/no3n samp"es must $e s m "ar to the ca" $rat ons samp"es 8e6cept for the prote n content:. Curve fitting C: Non-linear Iterative Curve Fitting ("spectral deconvolution" or "peak deconvolution" The " near "east s2uares cur)e f tt n! descr $ed n E&ur)e F tt n! ?E s s mp"e and fast. th s s a )er+ common ana"+t ca" s tuat on n commerce. a / nd of Etr a" and errorE procedure n 3h ch the parameters of the mode" are ad5usted n a s+stemat c fash on unt " the e2uat on f ts the data as c"ose as re2u red. Th s s essent a""+ a . Th s s a rea" data set der )ed from the near nfrared 8AI>: ref"ectance spectroscop+ of a!r cu"tura" 3heat samp"es ana"+7ed for prote n content. the resu"t n! coord nate transformat on of the no se n the data can resu"t n naccurac es n the parameters measured n th s 3a+. (o3e)er.ut ts !reat !enera" t+. $ut "a$or ous and t me0consum n!. $ut th s s poss $"e on"+ n some spec a" cases and. at "east for th s set of =0 3heat samp"es. The samp"es had a"read+ $een ana"+7ed $+ a re" a$"e. These resu"ts nd cate that t does. n the da+s $efore!ram. and t's true that. ca"cu"at n! a f tt n! errorB 8<: f the f tt n! error s !reater that the re2u red f tt n! accurac+.

"ac/$od+DataF t.m. depend n! on the "en!th of the f rst0!uess EstartE )ector:. an terat )e f t m !ht ta/e fract ons of a second 3here a re!ress on 3ou"d ta/e fract ons of a m "" second. ? s mp"e e6amp"e s f tt n! the $"ac/$od+ e2uat on to the spectrum of an ncandescent $od+ for the purpose of est mat n! ts co"or temperature. us n! the f tt n! funct on f t!auss2.+ to a mode" descr $ed n a pre) ous"+0created funct on ca""ed "it" nction. 3r t n! options 7 optimset(=!ol0=. Iterat )e cur)e f tt n! a"so ta/es "on!er than " near re!ress on 0 3 th t+p ca" modern persona" computers. the pea/ pos t on and the pea/ 3 dth 8the pea/ he !ht s a " near parameter and s determ ned $+ re!ress on n " ne 9 of the f tt n! funct on f t!auss2. Spreadsheets and stand'alone programs.IAS*?>&( 3 th the f tt n! funct on f t$"ac/$od+. and returns the f tt n! error.o!tions. In th s case there are t3o non0" near parameters. The custom demonstrat on scr pt Demof t!auss. In th s case there s on"+ one non" near parameter. The parameters of the f t are returned n the )ector EparametersE. the ent re process t+p ca""+ ta/es on"+ a fract on of a second.IAS*?>&( that uses the Ae"der0. p"ac n! the e6per menta""+ measured spectrum n the )ectors E3a)e"en!thE and Erad anceE and then ca"" n! F.m. The scr pt .').m. us n! the f rst !uesses n the )ector start and stopp n! at the to"erance def ned $+ the optimset funct on.m. a$o)e: to shorten the t me re2u red to f t the funct on to the re2u red de!ree of accurac+. atlab has a con)en ent and eff c ent funct on ca""ed F. For e6amp""a$ command 3 ndo3:. th s s fast enou!h for man+ purposes. St "".m demonstrates the techn 2ue. Aote: the term Espectra" decon)o"ut onE or E$and decon)o"ut onE s often used to refer to th s techn 2ue.m and s returned n the !"o$a" )ar a$"e EcE:. ?"" these funct ons ca"" the user0def ned pea/ shape funct on !auss an.%. 3h ch mode"s the sum of t3o e6ponent a" deca+s. There are a num$er of do3n"oada$"e non0 " near terat )e cur)e f tt n! adds0ons and macros for *6ce" and %pen%ff ce. # th contemporar+ persona" computers. Th s s eas "+ e6tended to f tt n! t3o o)er"app n! Gauss ans n Demof t!auss2.m demonstrates f tt n! a Gauss an funct on to a set of data. ?nother app" cat on s demonstrated $+ . temperature. 5ust t+pe Ef tdemoE n the .start. 8To see th s. such as "orent7 an.m or .&.3a+ of or!an 7 n! and opt m 7 n! the chan!es n parameters 8step <.m and ts f tt n! funct on f tfun. as 3e"" as some stand0a"one free3are and commerc a" pro!rams that perform th s funct on. The ma n d ff cu"t+ of the nteract )e methods s that the+ somet me fa " to con)er!e at an opt mum so"ut on n d ff cu"t cases. n the same order that the+ appear n EstartE. S m "ar procedures can $e def ned for other pea/ shapes s mp"+ $+ ca"" n! the correspond n! pea/ shape funct on. 8Aote: n order for scr pts " /e Demof t!"a$'s $u "t0 n demo f tdemo. It 3or/s n con5unct on 3 th a user0def ned Ef tt n! funct onE that computes the mode".m.m 8sho3n on the "eft: us n! the same f tt n! funct on 83h ch eas "+ adapts to an+ num$er of pea/s.y) performs an nterat )e f t of the data n the )ectors 6. compares t to the data.ead method. The standard approach to hand"e th s s to restart the a"!or thm 3 th another set of f rst !uesses.$ !arameter=F/01SE2R34(-"it" nction-.

. 3 dths. Lorent7 an. a"" the funct ons that the+ ca"" must $e "oaded nto .at"a$. a mode" of the nstrumenta""+0$roadened transm ss on spectrum s f t to the o$ser)ed transm ss on data. nc"ud n! those 3 th non0 nte!er and non0un form 60)a"ues. 3h ch uses an unconstra ned non0" near opt m 7at on a"!or thm to decompose a comp"e6.m:. #ccurac$ and precision of pea% parameter measurement There are four ma5or sources of error n measur n! the pea/ parameters 8pea/ pos t ons. he !hts. &nteracti#e "ea( )itter 8http://terpconnect. If +ou ha)e the 3ron! mode" for +our pea/s. Lo! st c.m and !auss an. n this application n a$sorpt on spectroscop+. to do3n"oad them to +our computer for use 3 th n . For e6amp"e.Demof t!auss2. Note.=:. @ou can create +our o3n f tt n! funct ons for an+ purposeB the+ are not " m ted to s n!"e a"!e$ra c e6press ons.. and areas: $+ terat )e cur)e f tt n!: a. The+ accept s !na"s of an+ "en!th. and a )ers on 3 th mouse0contro""ed s" ders 83h ch re2u res .at"a$ $eforehand. These pro!rams do not re2u re the s !na" process n! or opt m 7at on too"$o6es. n order to e6tend the d+nam c ran!e and ca" $rat on " near t+ $e+ond the norma" " m ts.umd. and e6ponent a""+0$roadened Gauss an mode"s 8e6panda$"e to other shapes:. the resu"ts can not $e . There are three d fferent )ers ons. us n! a f tt n! funct on that performs Four er con)o"ut on of the transm ss on spectrum 3 th the s" t funct on of the spectrometer. o)er"app n!0pea/ s !na" nto ts component parts.m to 3or/ on +our )ers on of .+ou can r !ht0 c" c/ on an+ of the m0f "e " n/s a$o)e and se"ect Sa#e 0in( $"a$"a$ pea/ f tt n! pro!rams for t me0ser es s !na"s. Pearson. odel errors. and can f ts !roups of pea/s 3 th Gauss an. a /e+press operated nteract )e )ers on. $ut can $e arbitrary comp"e6 mu"t 0step a"!or thms. The o$5ect )e s to determ ne 3hether +our s !na" can $e represented as the sum of fundamenta" under"+ n! pea/s )ePea/F"a$.htm: Th s s a ser es of . n th s case f t!auss2. a command " ne )ers on.

79.4<$$$$$$$.8.. .7>=.) FitRes lts$= .7:=< /eanFitError$= :. ! )es a f tt n! error of 9. the pea/ parameters of the pea/s that are f t ma+ not + e"d accurate parameter measurements.3) FitRes lts$= . f +our actua" pea/s are Lorent7 an n shape..:. th s pea/ ! )es a LD f tt n! error and he !ht and 3 dth errors of JD and 20D. 3 th a he !ht of 9. respect )e"+.... $ut s actua""+ t3o pea/s at 6H< and 6H= 3 th pea/s he !hts of 9. 3hen f t 3 th a Lo! st c mode"..:.+2). as +ou can see here.. and pea/ area:..$$$$$$$.$$$$$$$..: ?nother source of mode" error occurs f +ou ha)e the 3ron! num$er of pea/s n +our mode".7772$$$$$$ 7. as "on! as the pea/ s s+mmetr ca" and not h !h"+ o)er"app n! 3 th other pea/s . us n! the . a s n!"e so"ated Gauss an pea/ at 6H=.3<$$$$$$$2.47<: #hen f t 3 th a Lorent7 an mode" 8sho3n on the r !ht:. For e6amp"e.D and LD. pea/ he !ht.=< 8The f t resu"ts are.3.< /eanFitError$= .<.7.=4. pea/ num$er. For e6amp"e.. 55$6FitRes lts.000 and 3 dths . 55$ 6FitRes lts. pea/ pos t on.$$$$$$$:.. from "eft to r !ht.e6pected to $e accurateB for nstance.=..<D and he !ht and 3 dth errors or .000 f ts a Gauss an mode" ) rtua""+ perfect"+. respect )e"+: 55$6FitRes lts...$$$$$$$$$$$$:$$$$$$$$$$$$. $ut +ou f t them 3 th a Gauss an mode".. $ut the parameter errors are not e+ual to the f tt n! error 8that 3ou"d 5ust $e too eas+:.ut th s same pea/.:.77.=><3 So c"ear"+ the "ar!er the f tt n! errors./eanFitError9=!eak"it(6&-$y-9.2$$$$$$$"a$ user0def ned pea/f t funct on: 55$&=678.. f the data actua""+ has t3o pea/s $ut +ou tr+ to f t t 3 th on"+ one pea/.==24 /eanFitError$= 7./eanFitError9=!eak"it(6&-$y-9.2) FitRes lts$= . In the e6amp"e sho3n on the r !ht.. pea/ 3 dth. the "ar!er are the parameter errors. the parameters errors depend not 5ust on the f tt n! error $ut a"so on the data dens t+ 8num$er of data po nts n the 3 dth of each pea/: and on the e6tent of pea/ o)er"ap.y=e&!(*(&*:).. e)en f the mode" s 3a+ 3ron!.. are measured accurate"+. It's comp" cated./eanFitError9=!eak"it(6&-$y-9. or vice versa.:2$$$$$$$$.$$$$$$$$$$$$:$$$$$$$. 8To ma/e matters 3orse.7. c"ear"+ the pea/ width and area are the parameters most suscept $"e to errors.7. ?"so. The pea/ positions.. the s !na" looks " /e one pea/.

The s mp"est assumpt on.<<<.77./eanFitError9=!eak"it(6&-$y-9....:. that s.<<<3.:$$$$$$$..) FitRes lts$= .) FitRes lts$= .$$$$$$$. +ou !et: 55$&=678.<=<.ut e)en so there s often a res dua" $ac/!round that can not $e e" m nated comp"ete"+ e6per menta""+.3 . %rd nar "+ the e6per ment protoco" s des !ned to m n m 7e the $ac/!round or to compensate for the $ac/!round. rather than the random scatter of po nts that 3ou"d dea""+ $e o$ser)ed f a pea/ s accurate"+ f t..4:$$$$$$$. $ut often th s $ac/!round s a $road.==42 /eanFitError$= 7.....ode" errors resu"t n a E3a)+E structure n the res dua" p"ot 8"o3er pane" of the f !ure:. 55$6FitRes lts.ut a f t 3 th two pea/s 8sho3n on the r !ht: s much $etter and + e"ds accurate parameters for $oth pea/s: RR 6FitRes lts.+2)?e&!(*(&*:). If +ou f t th s s !na" 3 th a s n!"e0pea/ mode"... for e6amp"e $+ su$tract n! the s !na" from a E$"an/E from the s !na" from an actua" spec men.m funct on s turned on 8% /e+:.:.$$$$$$$3. .$$$$$$$$$$4.:. sa)e for the random no se.777<$$$$$$$..8.<<<:$$$$$$7.7..2..:>23 /eanFitError$= $$$$$$7.of 9. can $e appro6 mated as a stra !ht " ne n the "oca" re! on of !roup of pea/s $e n! f t to!ether./eanFitError9=!eak"it(6&-$y-9. a stra !ht0" ne $ase" ne connect n! the t3o ends of the s !na" se!ment n the upper pane" 3 "" $e automat ca""+ su$tracted as the pan and 7oom contro"s are used to so"ate the !roup of o)er"app n! pea/s to $e f t. 3h ch s used $+ the Interact )e Pea/ F tter.$$$$$$$. t "ted.79.$$$$$$$. s that the $ac/!round s "oca""+ " near. !ac(ground correction errors. 8Th s s one !ood reason for not smooth n! +our data $efore f tt n!:. and the pea/s are comparat )e"+ narro3 features super mposed on that $ac/!round. The pea/s that are measured n most measurement nstruments are often super mposed on a non0spec f c $ac/!round. .==7= 2$$$$$$$4.777< . b.y=e&!(*(&*4).>2$$$$$$$3. #hen the auto7ero mode of the pf. The or ! n and shape of that $ac/!round depends on the spec f c measurement method...+2).LL=.:>>=$$$$$$$2.7.. or cur)ed shape... There are )ar ous soph st cated methods descr $ed n the " terature for est mat n! and su$tract n! the $ac/!round n such cases.

m 83h ch re2u res the pea/f t. and therefore e6h $ t poorer parameter measurements:. and 82: the ma!n tude of the no se dur n! one samp"e m !ht not $e t+p ca"B the no se m !ht ha)e $een random"+ !reater or sma""er than a)era!e dur n! that t me. . The assumpt on s ord nar "+ made that the scatter s e2ua""+ a$o)e and $e"o3 the true s !na".LD and the percent re"at )e standard de) at on of the parameters ran!es from 0. ?n+ e6per menta" s !na" has a certa n amount of random no se. Th s s e6act"+ 3hat the scr pt Demo@eakfit. 3h ch means that the nd ) dua" data po nts scatter random"+ a$o)e and $e"o3 the r mean )a"ues.m2 ($) 5it with a three-peak 9aussian model. t 3 "" conta n another ndependent samp"e of the no se.ut the prec s on and the accurac+ of the measured parameters also depend on 3h ch parameter t s 8pea/ pos t ons are a"3a+s measured more accurate"+ than the r he !hts. and ca"cu"ate the standard error of each parameter. It's eas+ to demonstrate that. as e6pected. compute the mode"s parameters from each f t.m funct on: does for s mu"ated no s+ pea/ s !na"s such as those ""ustrated $e"o3. and areas: and on the pea/ he !ht and e6tent of pea/ o)er"ap 8the t3o "eft0most pea/s n th s e6amp"e are not on"+ 3ea/er $ut a"so more o)er"apped that the r !ht0most pea/. In th s e6amp"e. If another record n! of the s !na" s made. Th s means that the mathemat ca" Epropa!at on of errorE methods. *andom noise in the signal. the a)era!e f tt n! error prec s on and the re"at )e standard de) at on of the parameters ncreases d rect"+ 3 th the random no se "e)e" n the s !na". 3 dths. so that the "on!0term a)era!e approaches the true mean )a"ueB the no se Ea)era!es to 7eroE as t s often sa d. The pract ca" pro$"em s that an+ ! )en record n! of the s !na" conta ns on"+ one f n te samp"e of the no se. f t each of those separate"+. These no se samp"e are not nf n te"+ "on! and therefore do not represent the true "on!0term nature of the no se. . (') :aw data with peaks superimposed on baseline2 (#) Caseline automatically subtracted by the auto-ero mode in ipf.>xample of an experimental chromatographic signal. 5rom left to right. the f tt n! error s 9. c. 3h ch see/ to est mate the " /e"+ error n the mode" parameters $ased on the no se n the s !na". Th s presents t3o pro$"ems: 89: an nd ) dua" samp"e of the no se 3 "" not Ea)era!e to 7eroE and thus the parameters of the $est0f t mode" 3 "" not necessar "+ e2ua" the true )a"ues.0=D for the pea/ pos t on of the "ar!est pea/ to 92D for the pea/ area of the sma""est pea/. ? $etter 3a+ to est mate the parameter errors s to record mu"t p"e samp"es of the s !na". 3 "" $e su$5ect to error 8underestimating the error f the no se happens to $e lower than a)era!e and overestimating the errors f the no se happens to $e larger than a)era!e:.

then the resu"t n! ncrease n the num$er of data po nts n each pea/ shou"d he"p reduce the effect of no se. but the relative standard deviation of peak position. with the same noise level but with another peak overlapping it. yielding a relative fit error of &.8D . $ut the a)era!e parameter error drops to 0. &terati#e fitting errors. ?s a demonstrat on. ($) !he addition of a third peak further reduces the fit error to '. height. -n" /e mu"t p"e " near re!ress on cur)e f tt n!. and '. but also upon other peaks that are overlapping it. height.*D and relative standard deviation of peak position.m.m to create a s mu"ated o)er"app n! pea/ s !na" " /e that sho3n a$o)e r !ht. $ecause t uses s" !ht"+ d fferent start n! )a"ues each t me the s !na" s f t 8$+ press n! the ) /e+ n pf. height.&D (because the addition if the second peak increases overall signal amplitude). $+ press n! the 4 /e+. us n! the scr pt DemoPea/f t. The Interact )e Pea/ F tter ma/es t eas+ to test th s. and width of %. d. t's poss $"e to chan!e the nter)a" $et3een 6 )a"ues and thus the tota" num$er of data po nts n the s !na". %.*ED. (#) !he same peak.= and the a)era!e parameter error s 0. *)en $etter. or to ta/e mu"t p"e read n!s at each 60a6 s )a"ues.!he parameter errors depend not only on the characteristics of the peaks in +uestion. reduces the relative fit error to #. respectively. about the same as with two peaks. and &D . # th . because the third peak does not overlap the first one significantly. the f tt n! error s 0. # th a no se "e)e" of 9D and K= po nts n the s !na". and width of the first peak are still %. the f tt n! error s essent a""+ the same. but with poorer precision for the first peak. ED.a seemingly better fit. and $. If the e6per ment ma/es t poss $"e to reduce the 60a6 s nter)a" $et3een po nts. the pf. 5rom left to right< (') a single peak at x7'%% with a peak height of '. for e6amp"e:.m funct on .JD. but increases the relative standard deviation of peak position.% and width of $% is fit with a 9aussian model.FD.FD.F&D. and width to %..#D.ED . E.00 po nts n the s !na" and the same no se "e)e".<D. terat )e methods ma+ not con)er!e on the e6act same mode" parameters each t me the f t s repeated 3 th s" !ht"+ d fferent start n! )a"ues 8f rst !uesses:. %ne 3a+ to reduce the effect of no se s to ta/e more data.8&D. su!!est n! that the accurac+ of the measured parameters )ar es n)erse"+ 3 th the s2uare root of the num$er of data po nts n the pea/s.

*. no se.T. and $ac/!round.s "ent"+ computes 90 f ts 3 th d fferent start n! )a"ues and ta/es the one 3 th the "o3est f tt n! of the f !ures n th s essa+ are screen ma!es from S.: the errors depend on the data dens t+ 8num$er of ndependent data po nts n the 3 dth of each pea/: and on the e6tent of pea/ o)er"ap 8the parameters of so"ated pea/s are eas er to measure than h !h"+ o)er"apped pea/s:. a . . Th s s !enera""+ a !ood assumpt on. In man+ pract ca" cases there s enou!h random no se n the s !na"s that the terat )e f tt n! errors 3 th n one samp"e of the s !na" are sma"" compared to the random no se errors $et3een samp"es.S d fference $et3een the mode" and the data: s m n m 7ed. >emem$er that the )ar a$ " t+ n measured pea/ parameters from f t to f t of a s n!"e samp"e of the s !na" s not a !ood est mate of the prec s on or accurac+ of those parameters. a Esamp"eE s a s n!"e record n! of s !na":. 8In th s case. 3h ch sho3s t+p ca" percent parameters errors as a funct on of f tt n! error for the "eft0most pea/ n one samp"e of the s mu"ated s !na" !enerated $+ DemoPea/f t. 8S !na" "rocess n! for E6per menta" /hem str+ %each n! and *esearch/ 5n )ers t+ of ar+"and:. for the s mp"e reason that those resu"ts represent on"+ one samp"e of the s !na"..&. the parameter errors 8the d fference $et3een the actua" parameters and the parameters of the $est0f t mode": 3 "" a"so $e m n m 7ed.-.P. rather than $+ random no se n the s !na".>. So.. 3e can ma/e the fo""o3 n! o$ser)at ons a$out the accurac+ of mode" parameters: 89: the parameter errors are d rect"+ proport ona" to the no se n the data and to the f tt n! error 8$ut s not e2ua" to the f tt n! error:B 82: the errors are t+p ca""+ "east for pea/ pos t on and 3orse for pea/ 3 dth and areaB 8. ? $as c assumpt on of an+ cur)e f tt n! operat on s that the f tt n! error 8the >. A endi&# Software details '( SP)CTR*! . to sum ntosh pro!ram 3e ha)e de)e"oped for teach n! s !na" process n! to chem str+ students. The samp"e0to0samp"e )ar at ons are " /e"+ to $e much !reater than the 3 th n0samp"e )ar at ons due to the terat )e cur)e f tt n!.m 8sho3n n the pre) ous sect on:. as demonstrated $+ the !raph to the r !ht. The )ar a$ " t+ of the f tt n! error here s caused $+ random sma"" )ar at ons n the f rst !uesses.

for e6amp"e $+ reduc n! no se. achine *e1uirements. $ut the pro!ram cou"d $e used n an+ f e"d n 3h ch nstrumenta" measurements are used: e. mu"t p" cat on. rec proca". rotate. !roup po nts $+ 2s. The pro!ram and ts assoc ated tutor a" 3as or ! na""+ de)e"oped for students of ana"+t ca" chem str+. compensat n! for nstrumenta" art facts.?n+ . s des !ned for post0run 8rather than rea"0t me: process n! of Espectra"E or t me0 ser es data 8+ )a"ues at e2ua""+0spaced 6 nter)a"s:. an+ standard pr nter. and decompos n! a comp"e6 s !na" nto ts component parts. has $een tested on most . for &est !hemistr$ software and for &est "esign. P& users can run SP*&T>-.9. en! neer n!.. re)erse O0a6 s. su$tract on. can $e used $oth as a research too" and as an nstruct ona" a d n teach n! s !na" process n! techn 2ues.. ant "o!. $ ochem str+. e6tract su$set of po nts. "o!. a!r cu"tura" sc ntosh mode" 3 th m n mum 9 . Features • • • • • • • • • • • • • • • • >eads one0 or t3o0 co"umn 8+0on"+ or 60+: te6t data ta$"es 3 th e ther ta$ or space separators D sp"a+s fast. test n! h+potheses. 3 th mouse0contro""ed sett n! of start and stop po nts Four er transformat on Po3er spectra Four er f "ter n! &on)o"ut on and decon)o"ut on &ross0 and auto0corre"at on . chem str+. etc. norma" 7e. nc"ud n!: nspect and ed t nd ) dua" data po nts.+te >?. super mpose. nterpo"ate. "a$e"ed p"ots n standard res 7a$"e 3 ndo3s 3 th fu"" 60 and +0a6 s sca"e e6pans on and a mouse0 contro""ed measurement cursor ?dd t on. The pro!ram enhances the nformat on content of nstrument s !na"s. chromato!rams. med ca" ntosh emu"ator pro!ram runn n! on a .ac ntosh mode"s and on a"" )ers ons of the operat n! s+stem throu!h %S J. standard de) at on. $ o"o!+. ph+s cs. s ne 3a)e and norma""+0d str $uted random no se ? num$er of other usefu" funct ons. and d ) s on of t3o s !na"s T3o / nds of smooth n!. e"ectrochem ca" s !na"s.!:GB was the winner of two >DG. &o"or screen des ra$"e. Three / nds of d fferent at on Inte!rat on >eso"ut on enhancement Interpo"at on Fused pea/ area measurement $+ perpend cu"ar drop or tan!ent s/ m methods. en) ronmenta" and earth sc ences. or mater a"s test n!.. "n. concatenate.B)A. set O a6 s )a"ues. us n! a . a$so"ute )a"ue. 7ero f "".SP*&T>-. ant "n. s2uare root SP*&T>-. SP*&T>-. such as spectra.:3@!A/ national software awards in '**%. h sto!ram.u "t0 n s !na" s mu"ator 3 th Gauss an and Lorent7 an $ands. c" n ca" ps+cho"o!+. $r d!e se!ment. 4@>.!. mpro) n! reso"ut on.

Signal processing tutorial.? " $rar+ of prerecorded data f "es for use 3 th the s !na" process n! tutor a". Se"f0!u ded tutor a" on the app" cat ons of s !na" process n! n ana"+t ca" chem str+.umd.>*&T>-. Aote: .+e: S !na"s are stored nterna""+ as extended-precision rea" )ar a$"es and there s a " m t of 902< po nts per s !na". crosoft #ord.anua". #r teAo3. . crosoft #ord. . 3h ch s pre0"oaded nto the *6ecutor .ac ntosh 3ord processors:. *eference manual.on" " s/ II 8http://$as " s/ ntosh 3ord processors:. %utorial s mu"ator. sh ft0c" c/ on the do3n"oad " n/s a$o)e to $e! n the do3n"oad. $ut a""o3s s !na"s 3 th data more po nts.99e.*&T>-. SP*&T>*&T> There are t3o )ers ons: S"E/%*5 +. do3n"oad the E(QOE f "es to +our & dr )e. and most other fu""0featured . then open the do3n"oaded (QO f "es 3 th Stuff t *$as " s/2/:.ac8http://333. There s a"so a documentat on pac/a!e 8"ocated at http://terpconnect. "aunch *6ecutor.htm": c. *6p"a ns each menu se"ect on and descr $es the a"!or thms and mathemat ca" formu"ae for each operat on.umd. %he full #ersion of S"E/%*5 +. The SP*&T>-. If +ou are us n! the ?>DI *6ecutor .ac#r te. . S"E/%*5 te.h26. and most other fu""0featured . &"ar s#or/s. Stuff t *6pander 3 "" automat ca""+ decode and decompress the do3n"oaded f "es. &urrent"+ a)a "a$"e .$ ntosh en)<9J&/S !na"Process n!.net/:. The do3n"oad address of th s )ers on n (QO format s http://terpconnect.c5$.ac3r te format 8&an $e opened from 3 th n .h26: cons st n! of: a. If +ou are do3n"oad n! on a # ndo3s P&.ecause t 3as de)e"oped for academ c teach n! app" cat on 3here the most modern and po3erfu" mode"s of computers ma+ not $e a)a "a$"e.anua" s a"so a)a "a$"e separate"+ n PDF format at http://terpconnect. These are p"a n dec ma" asc 8ta$0de" m ted: data f " ntosh emu"ators nc"ude Soft.umd.h26.+ is no6 a#ailable as free6are3 and can $e do3n"oaded from http://terpconnect. The do3n"oad address of th s )ers on n (QO format s http://terpconnect. . and ). 3as des !ned to $e E"ean and meanE 0 that s. a"" th s shou"d happen comp"ete"+ automat ca""+. Th s )ers on s "ess prec se n ts ca"cu"at ons 8has more numer ca" round0off error: than the other )ers on. &"ar s#or/s.umd. t has a s mp"e . These f "es are $ nhe6 encoded: use Stuff t *6pander to decode and decompress as usua".# ndo3s mach ne.or!/:. .edu/~toh/spectrum/SP*&T>-.emu" ntosh. Th s tutor a" s a"so a)a "a$"e on the #e$ at 8http://terpconnect.pdf. #r"oad. .+b: S !na"s are stored nterna""+ as single-precision rea" )ar a$"es and there s a " m t of <000 po nts per s !na".99$.as " s/ II JIT 8http://!3eno"e. >eference .ac ntosh0t+pe user nterface and )er+ sma"" memor+ and .ac 8http://333.demo. If +ou are do3n"oad n! on a .ac3r te format 8&an $e opened from 3 th n . Th s )ers on performs a"" ts ca"cu"at ons n e6tended prec s on and thus has the $est d+nam c ran!e and the sma""est numer c round0off errors. The t3o )ers ons are other3 se dent ca".

@rinciples of 3nstrumental Analysis. 99J<. and . 3nstrumental Analysis. . Th s pro!ram s free and ma+ $e free"+ d str $uted.3 / ped a. For e6amp"e: a. and 3 "" e)en run on o"der monochrome mode"s 83 th some cramp n! of screen space:. ..e+ers 8*d. &hr st an and James *.?TL?. *n!"e3ood &" ffs. Ph "ade"ph a. 9KL<W9J00. Dou!"as ?. 99J9.or!/3 / /. for a !enera" descr pt on.u$. =. ?""+n and . #.htm" f. and Gar+ (or" c/.math3or/s. . atlab Summary and %utorial. L.u$c.?. 99JL. It 3 "" 3or/ 2u te 3e"" on .:. *n/e. $ "ractical &ntroduction to atlab 8http://333.hemistry. Prent ce (a"". Mo3a"s/ . &.. . It does not re2u re a math co0processor. ?.edu/~ms!oc/en/ ntro/ ntro. atlab /hemometrics &ndex http://333. 8http://333. >.d s/ space re2u rements.math. . +( !atlab . S/oo!. %'(a) der/ca)ers/. #ent7e"" and &hr stopher D.uf". %'>e ""+.htm": ntosh )H. . Gates and Jordan .hemometrics. See http://en. <.htm" *eferences 9. John # "e+ and Sons.en"o Par/."a$ s a h !h0performance commerc a" numer ca" comput n! en) ronment and pro!ramm n! "an!ua!e that s 3 de"+ used n research and educat on. 8c: 99J9 T. Stephen &. Pa!es J<L0J=9.?TL?. ulti#ariate /ur#e *esolution.cs. There are se)era" $as c on0" ne tutor a"s and co""ect on of samp"e code.en5am"come. /aboratory Automation using the 3CB @.htm":"p/mat"a$0tutor a"/: d. $n &ntroductory 7uide to $%0$!.at"a$Gu de/!u de. Pa!es J9L0JK0. &hr st e G./&hemometr cs/ n de6.uhammad ?. p.math. Second *d t on. . ntosh mode"s as o"d as the . http://333.a"mstadt. AJ. Ae3 @or/. b.drShP7(e@!:. n >ncyclopedia of Analytical . Pa!es K. . co""ect ons or other arch )"a$centra"/" n/Ve6chan!e/. $%0$! %utorial for Ne6 5sers 8http://333. 99JL. Gar+ D. De$orah L I""man. Sharaf. >lectronic Beasurements for 4cientists. S !na" Process n! n ?na"+t ca" &hem str+. Th rd *d t on. (o3ard F. It ma+ $e nc"uded on &D0>%. 8http://333. 99K<. .ruce >. Peter D.0KL. .+outu$e.ro3n. Th s s a narrated <0m nute ) deo ntroduct on for ne3 users.

http://sc ence. 8Do3n"oada$"e chapter $+ chapter n PDF format from http://333.hemistry and in 9eneral 4cientific Data Analysis.oo/s..a+ 999J:.htm": 9<. chae" Po"to3s/ .!"e""a/mat"a$G-Is/: 9=. *fstath ou. Scott Fan . # "e+0F&( 8.3 dener.htm": .ecn. Fe" n!er.!r/?pp"ets/?pp"etVInde62. 8http://users. &S!N'+80=29L<<J<<. Sm$oo/.or!/ ppa!e/e/ pdata/2009/9. 2000 8http://m+3e$. Stat st cs for ?na"+t ca" &hem str+.htm:.htm". *ducat ona" . and &hemometr cs. 2009:. &enter for S !na" and Ima!e Process n! 8&SIP:. &am$r d!e -n )ers t+ PressB 9 ed t on 8Fe$ruar+ 9=. Data ?na"+s s and S !na" Process n! n &hromato!raph+.dsp!u de. &hemometr cs: Stat st cs and &omputer ?pp" cat on n ?na"+t ca" &hem str+.ouman.ow to use >xcel in Analytical . 99.chem.htm: J. . Aumer ca" ?na"+s s for &hem ca" *n! neers. D ! ta" S !na" Process n! Demonstrat ons n .atth as %tto.cher c.John # "e+ X Sons Ltd.$/ stats/stats. S !na" Process n!. &h"a$ G-Is. 92. . The Sc ent st and *n! neer's Gu de to D ! ta" S !na" Process n!. ?. 9.ramer. Some parts ) e3a$"e n Goo!"e . *"se) er Sc ce 899 . >o$ert de Le) e. *ducat ona" ?pp"ets n ?na"+t ca" &hem*/ee<.uoa. &onstant nos *. &har"es . Purdue -n )ers t+ 8http://333.pdf: K.da". PDF e6cerpt . Jan ?""e$ach.arch"a$. Ste)en #.J/demos/Demos. and . 90.ece. Th s s a much more !enera" treatment of the top c. 8http://333. Taechu" Lee 8http://333./"ecture. Geor! a Inst tute of Techno"o!+.

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