certified uv/vis reference materials - dialab · certified uv/vis reference materials guidelines //...

CertifiedUV/Vis Reference Materials Guidelines

// easy handlinG// Reliable measuRement Results // inteRnational compaRability

High Precision in Spectro-Optics

// Glass filteRs

// liquid filteRs

// ceRtified RefeRence plates

accredited accordance todin en iso 17025

tRaceable

3

contents

1. IntroductIon page 4–10

1.1 hellma analytics calibration laboratory page 4

1.2 certified test equipment page 6

1.3 Glass filter applications page 6

1.4 liquid filter applications page 6

1.5 dakks calibration certificate page 8

1.6 expanded absorbance range – from 0 to < 3.1 for uV-Vis reference materials page 11

2. Glass fIlters page 12–21

2.1 holmium oxide glass filter for checking wavelength accuracy page 12

2.2 didymium glass filter for checking wavelength accuracy page 13

2.3 didymium glass filter for checking photometric accuracy page 14

2.4 neutral density glass filters for checking photometric accuracy page 15

2.5 Glass filter sets page 16

2.6 General usage guidelines for glass filters page 17

2.7 calibration with glass filters page 18–21

3. lIquId fIlters page 22–35

3.1 holmium oxide liquid filter for checking wavelength accuracy page 22

3.2 potassium dichromate liquid filter for checking photometric accuracy page 23

3.3 liquid filters for checking for stray light page 24

3.4 liquid filters for checking spectral resolution page 25 3.5 liquid filter set compliant with the european pharmacopoeia page 26

3.6 liquid filter set compliant with the united states pharmacopeia page 27

3.7 General usage guidelines for liquid filters page 27 3.8 calibration with liquid filters page 28–35

4. certIfIed reference plates page 36–41

4.1 Reference plate for checking photometric accuracy page 36

4.2 Reference plate for checking photometric and wavelength accuracy page 37

4.3 General usage guidelines for reference plates page 38

4.4 calibration with reference plates page 39–41

5. recertIfIcatIon page 42–43

6. faqs page 44–45

7. Glossary page 46

8. recommendatIons for further readInG page 47

9. product overvIew page 48–50

accRedited to

din en iso 17025

although checking measuring equipment to ensure thatresults are accurate has long been common practice foranalytical balances, it still tends to take something ofa backseat where spectrophotometers are concerned.Spectrophotometers are important instruments that playa major role in health care, the life sciences, environmentalanalysisandprocessessuchasproductioncontrolandensu-ring product quality. Over the last two years, many labora-torieshavebecomeconsiderablymoreawareof theneed tocheck their spectrophotometers, making it all the more im-portant to know that these precision tools are also subjectto mandatory checks under DIN eN ISO 9001. The standardclearly stipulates that measuring equipment be calibratedor verified, either at regular intervals or before use, usingmeasurementstandards thatcanbe tracedback to interna-tional or national standards. For an overview of the measu-rementstandardsforUV-Visspectrophotometers,pleasere-fertoourreferencematerialsinthisbrochure.anincreasingnumber of laboratories are turning to this easy method forensuringhighstandardsofwork–notonlytosatisfyrequire-ments in time for theirnextaudit,butalso tobesafe in theknowledge that theyare takingaccuratemeasurementsandthus basing their actions and responses on correct results.We are delighted that our products are helping to achie-ve this. Details of our product range, as well as usage gui-delines,helpful tips,and recommendations,areall includedin this Handbook. We VeRy much hope that you enjoy ReadinG it!

1.1 hellma analytics calibration laboratory: accredited to din en iso 17025

Our lab is a DakkS calibration laboratory and is accredited toDIN eN ISO 17025, a comprehensive quality management sy-stem that acts as a seamless continuation of other systemssuchas ISO9000.Byachieving thisaccreditation,wehavede-monstratedproofofexpertiseinthecalibrationactivitiesthatweperform and are authorized to issue internationally recognizedDakkS calibration certificates. accreditation is the key to highqualitymeasurements, internationalcomparability,andtrust inboththeworkofthecalibrationlaboratoryandthetransparencyofresults.

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1. IntroductIon

dear Readers,

din en iso 17025

accReditationstands for trust – and trust is

built on reliability(in latin accredere means giving credibility

to something)

accRedited to

din en iso 17025

birgit Kehl,HeadofHellmaanalyticscalibrationlaboratory

76

1.2 certified test equipment

Qualityassuranceandqualitycontrolregulations,suchasISO9000,gLp,gMp,andpharmacopoeias,requirecompaniestoverify the consistently excellent performance of any spectro-photometerinuse.Thetwomostimportantfactorsforobtai-ningprecisespectrophotometerdataarethephotometricac-curacy(absorbanceaccuracy)andwavelengthaccuracyofthespectrophotometer,whichshouldbetestedonaregularbasis.

IntheHellmaanalyticscalibrationlaboratory,whichisacc-redited toDINeN ISO17025,wemanufacturecertified re-ferencematerialsbasedon the regulatorycodes issuedbyNIST(NationalInstituteofStandardsandTechnology),aSTM(americanSocietyforTestingandMaterials)andpharmaco-poeias (ph.eur.,DaB,USp).all certifiedmeasurement re-sultscanbetracedbacktoNISTstandardreferencemateri-als(SRMs).(photometricaccuracy:SRM®930eandSRM®1930,wavelengthaccuracy:SRM®2034).

choose between glass filters and liquid filters for your reference materials:

1.3 Glass filter applications

glass filters are certified reference materials made of glassmanufactured specifically for calibration. They are, above all,extremelyrobust.allglassfilterscertifiedbyHellmaanalyticsare traceable to NIST primary standards. Certified glass fil-tersaresuitableforcheckingthefollowingparametersofyourspectrophotometer:

» Wavelength accuracy » photometric accuracy (absorbance)

1.4 liquid filter applications

Liquidfiltersarecertifiedliquidreferencematerialsthataremanufacturedincompliancewithpharmacopoeiasand/orNISTstandardsandfilledintoquartzglasscuvettesundercontrolledconditions.Thecuvettesarethenpermanentlysealedtobecomeairtight.Liquidfiltershavethedistinctad-vantageofequatingtorealmeasuringconditions.CertifiedHellmaanalyticsliquidfiltersaresuitableforcheckingthefollowingparametersofyourspectrophotometer:

» Wavelength accuracy » photometric accuracy (absorbance) » stray light levels » spectral resolution

You should regularly check your UV/Vis spectrophotome-ter forallof these factors,especiallyphotometricandwa-velength accuracy, while observing the relevant require-mentsinyourdevicehandbook.Thankstotheireaseofuseand long service life, certified Hellma analytics referencematerialsprovideanexcellentaidforallroutinechecks.

hellma analytics’ dakks-certified refe-rence materials comply with the provi-sions stipulated by quality management systems and pharmacopoeias, meeting the highest quality requirements and ensuring the international comparability of measurement results.

1. IntroductIon

hellma analytics RefeRence mateRials in accoRdance With the most impoRtant set of Rules and ReGulations

mateRial checKinG of RanGe ph.euR. dab usp 851 astm

Glass filteRs

holmium oxide glass Wavelengthaccuracy UV/Vis

didymium glass filter Wavelengthaccuracy UV/Vis

didymium glass filter photometricaccuracy UV

neutral density glass photometricaccuracy Vis

liquid filteRs

holmium oxide solution Wavelengthaccuracy UV/Vis

potassium dichromate solution photometricaccuracy UV/Vis

toluene in hexane Spectralresolution UV

potassium chloride solution Straylight UV

sodium iodide solution Straylight UV

sodium nitrite solution Straylight UV

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Kalibriergegenstand: Kalibrierstandard-Satz, bestehend aus drei Neutralglas-filtern NG11, NG5 und NG4.

Calibration Object: Set of calibration filters, consisting of three neutral density glass filters NG11, NG5 and NG4.

Kalibrierverfahren: Messung der optischen Dichte. Diese Kalibrierstandards wurden gegen Luft als Referenz gemessen.

Calibration Method:Measurement of optical density. These calibration standards were measured using air as reference.

Messtechnische Bedingungen bei der Kalibrierung: Conditions of Calibration: Die in diesem Kalibrierschein angegebenen Werte wurden mit dem verwendeten Spektralphotometer und den nach-folgenden Einstellungen ermittelt:

The following settings were used on the spectrometer employed to obtain the data quoted on this calibration certificate:

UV/VISModus der Ordinatenskala: Optische Dichte (Abs) Spaltbreite: 1,00 nm Spaltmodus: Fix Integrationszeit: 3,0 s

UV/VISOrdinate mode: Optical density (Abs)

mn 00.1 :tilS xiF :edoM tilS

Integration time: 3.0 s

Für die Kalibrierung dieses Kalibriergegenstandes wurde ein UV/VIS/NIR-Spektralphotometer Varian Cary-5000 mit der Seriennummer UV1101M202 eingesetzt.

This calibration object was calibrated on a UV/VIS/NIR spectrophotometer Varian Cary-5000 with serial number UV1101M202.

Dieses Gerät wird regelmäßig auf die Einhaltung seiner Spezifikationen überprüft. Datum der letzten technischen Überprüfung: 27. November 2014

This instrument is regularly checked for the compliance with its specifications. Most recently technical check: 27 November 2014

Für die regelmäßige Überprüfung der photometrischen Richtigkeit werden die Bezugsnormale des NIST SRM 930e Filter Nr. 2115, gültig bis März 2015 und SRM 1930 Filter Nr. 202, gültig bis März 2016 eingesetzt.

A set of NIST SRM 930e Filter No. 2115, valid until March 2015 and SRM 1930 Filter No. 202, valid until March 2016 standard reference materials is used to regularly check the photometric accuracy of the spectrophotometer.

Zur regelmäßigen Überprüfung der Wellenlängenrichtigkeit wurde das intrinsische Bezugsnormal

Hellma UV5 S.Nr. 0861 / 87450-PTB-14, gültig bis Dezember 2015 eingesetzt.

The intrinsic standard reference material Hellma UV5 serial no. 0861 / 87450-PTB-14, valid until

December 2015 is used to regularly check the wavelength accuracy.

Zusätzlich werden die Emissionslinien von Deuterium, Quecksilber und Argon zur Überprüfung der Wellen-längenrichtigkeit verwendet.

In addition, the emission lines of deuterium, mercury and argon are used to check the wavelength accuracy.

:snoitidnoC latnemnorivnE :negnugnidebsgnubegmU

Die Messungen wurden bei einer Umgebungstemperatur von 22°C ± 2°C und einer relativen Luftfeuchtigkeit von 30% bis 65% durchgeführt.

Measurements were performed at an ambient temperature of 22°C ± 2°C and a relative humidity of 30% to 65%.

1.5 dakks calibration certificate

after careful production, reference materials are certifiedintheHellmaanalyticscalibrationlaboratory(accreditedtoDIN eN ISO 17025) using a high-performance UV-Vis/NIRspectrophotometer. Reference materials are only conside-

redtobecertifiediftheyhavebeenissuedwithaDakkSca-libration certificate and bear a calibration mark. Using themeasurement valuesdocumentedandcertified in thecali-brationcertificate,userscancheckandcalibratetheirspec-trophotometersaccordingly.

1 description of calibrated item

2 measurement conditions

3 type of device used to carry out

measurements

4 type, serial number and validity of

calibrated nist/ptb reference

standards used to regularly check

reference photometers; details of

additional checking methods

5 ambient conditions during

measurement

1 name of accreditation body

2 name and address of issuing

laboratory

3 description of calibrated item

4 clear details of measured item

5 customer

6 date of measurement

7 internal document reference,

including date of issue and last

modified date

8 date of issue of calibration certificate

9 calibration certificate serial numbers

10 dakks registration number

11 year and month of calibration

12 authorized persons

the dakks calibration certificate from the national accreditation body for the federal Republic of Germany may only be issued by accredited partners. the hellma analytics calibration laboratory is the only such laboratory in Germany that is accredited to certify uV-Vis re-ference materials.

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1. IntroductIon

accuRate measuRement

Results GuaRanteed.

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1. IntroductIon

1 measurement value and smallest attributed measurement uncer-tainty that can be specified. this value only refers to hellma analytics measurements and applies solely to the company’s specific measu-rement conditions. in justified cases, calibration certificates may also show measurement results that do not fall within the calibration laboratory’s scope of accreditation. these must be clearly labeled as such on the calibration certificate.

2 notes on determining expanded measurement uncertainty.

3 notes on initial measurements of filters used to determine optical den-sity. initial measurements are not taken for filters used to determine wavelength accuracy.

4 calibration certificates shall not contain recommended recertification in-tervals (in accordance with dakks-dKd-5). exceptions may be made if re-quested by the customer or required by legislation.

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1.6 extended absorption range – from 0 to < 3.1 for uV/Vis reference materials

Underthepreviousaccreditation,thecompanywascertifiedto manufacture UV-Vis reference materials for wavelengthaccuracyandopticaldensityuptoanabsorbanceof2.05a.However, pharmaceutical applications, biotechnology andspectrophotometer manufacturers all increasingly requirereferencematerialswithacertifiedabsorbanceupto3a.

SinceNIST(NationalInstituteofStandardsandTechnology)is no longer producing the relevant SRM 2930 filter set,Hellmaanalyticshasmanufacturedanequivalentreferencematerial,whichhasbeenofficiallycertifiedbyNIST,and isavailableastheir666S300filterset.

aRticle no. 666S300

applicaton Filtersetfortestingthephotometricaccuracy

contentF390:glassfilter(0.04abs)F301:Neutraldensitiyglassfilter(2.5abs)F303:Neutraldensitiyglassfilter(3.0abs)

standaRd ceRtification Wavelengths: 440,465,546.1,590,635nmslit width:1nm

extended abs.range till < 3,1! neW

PREVIOUS ABSORBANCE RANGE: < to 2,05

190 300 400 500 600 700 800 8900

0,5

1

1,5

2

2,5

3

3,5Abs.

ERWEITERTE DAKKS-AKKREDITIERUNG FÜRZERTIFIZIERTE UV/VIS- REFERENZMATERIALIEN

nm

NEW ABSORBANCE RANGE: < to 3,1

extended dakks accreditation for certified uV-Vis reference materials

1312

230 430 530 6303300

1

Abso

rptio

n

Wavelength (nm)

2

279.3360.8 453.4

536.4637.5

270 470 570 8703700

1

Abso

rptio

nWavelength (nm)

4

329

2

3

770670

472

512

681

875

2.1 holmium oxide glass filter for checking wavelength accuracy

applicationTo measure wavelength accuracy, the filter absorbs thelightbeamof thespectrophotometer toagreaterextentatcertainwavelengthscreatingabsorbancepeaks.Ideally,anyreferencematerialsusedtodeterminewavelengthaccuracyshouldhavenarrow,well-definedpeaksatavarietyofwa-velengthsintheUVandvisiblerange.

pRoduct descRiptionHolmiumoxideglassfilter666-F1hasarangeofnarrow,well-definedpeaksintheUVandvisiblerange,makingholmiumanexcellentchoiceforcheckingthewavelengthscaleofspectro-photometers. In comparison to filters that use holmium oxi-de solutions, the holmium oxide glass filter has a somewhatweakerspectrumwithfewerpeaks.InthelowUVrangeinpar-ticular, theabsorbancebehaviorof theglassmatrix issuper-imposedontheholmiumpeaks.Themainadvantageofusingaglassfilteroveraliquidfilteristhatitismorerobust.

notesThepositionsofholmiumoxidepeaksmayvaryslightlydepen-dingontheglassbatchused.ThisiswhyHellmaanalyticscerti-fieseachholmiumoxideglassfilterindividually.

2.2 didymium glass filter for checking wavelength accuracy

applicationpleasesee2.1formoredetailsoftheapplication.

pRoduct descRiptionDidymiumglassfilter666-F7WismadefrommaterialspeciallymanufacturedbySchottag.Likeholmiumoxideglass,didymi-umglasshasavarietyofcharacteristicpeaksintheultravioletand visible rangeand is therefore typicallyused for checkingwavelengthaccuracy. Itspeaksarenotasnarrowasthoseofholmium oxide glass filters, however. The filter’s absorbancebehavior in theultraviolet rangealsomakes itsuitableasanabsorbancefilterforcheckingphotometricaccuracy(see2.3).

notesThe positions of didymium glass peaks may vary slight-ly depending on the glass batch used. This is why Hellmaanalyticscertifieseachdidymiumglassfilterindividually.

aRticle no. 666F1-339

application TestingthewavelengthaccuracyintheUVandVisrange(279nmto638nm)ataspectralbandwidthupto2nm.

content Holmiumoxideglassfilterwithmetalframe

standaRd ceRtification Wavelength accuracy at: 279;361;453;536;638nmslit width: 1nm

possible ceRtification additional possible wavelengths: 287;418;445;460nmslit width: allupto2nm

aRticle no. 666F7W-323or666F7-323

application TestingthewavelengthaccuracyintheUVandVisrange(379nmto875nm)ataspectralbandwidthupto2nm.

content Didymiumglassfilterwithmetalframe

standaRd ceRtification Wavelength accuracy at:379;472;512;681;875nm slit width:1nm

possible ceRtification additional possible wavelengths: 291;302;430;446;482;626nmslit width:allrecommendedupto2nm

Typicalspectrumforaholmiumoxideglassfilter Typicalspectrumforadidymiumglassfilter

2. Glass fIlters WaVelenGth accuRacy

Gla

ss f

ilte

Rs

Gla

ss f

ilte

Rs

1514

2.3 didymium glass filter for checking photometric accuracy

applicationTo measure photometric accuracy (absorbance), the filterreduces the light beam from the spectrophotometer. an ab-sorbancevalue(abs)canbededucedfromthelightextinctioncausedbythefilter.pRoduct descRiptionDidymiumglassfilter666-F7aismadefrommaterialspeciallymanufactured by Schott ag. The didymium glass filter’s ab-sorbancebehavior in theultravioletrangealsomakes itsui-table for use as an absorbance filter. Didymium glass filtersarethereforesuitableforcheckingwavelengthaccuracyintheUV-VisrangeaswellascheckingphotometricaccuracyintheUVrange.

absorbance behavior in the UV range can be checked at270nm,280nm,297nm,320nm,and340nm.Filtersarerou-tinelysetatathicknessthatproducesanominalopticaldensi-tyof0.5absat340nm.Thisresultsinincreasinglylargerab-sorbancestheshorterthewavelengthsbecome.

notesThese absorbance values vary greatly depending on theglass batch used, however, and can only be compared forfiltersderivedfromthesameglassmeltingprocess.Thisiswhyalldidymiumglassfiltersarecertifiedindividually.

Abso

rptio

n

0270 280 290 300 310 320 330 340 350 360 370

Wavelength (nm)

0.5

1.5

1

2

ca. 1.0 A

ca. 0.9 A ca. 0.8 A

ca. 0.6 A

ca. 0.5 A

aRticle no. 666F7a-323or666F7-323

application TestingthephotometricaccuracyintheUVrange(270nmto340nm)

content Didymiumglassfilterwithmetalframe

standaRd ceRtification photmetric accuracy: approx.0.5to1abs.at wavelengths: 270;280;297;320;340nmslit width: 1nm

possible ceRtification additional possible wavelengths:from270to290nmand310to320nm slit width:allpossibleupto5nm

Selectedspectrumforadidymiumglassfilterbetween270nmand370nm

400 650 700 750 800500 550 600450

Abso

rptio

n

Wavelength (nm)

0.2

0.4

0.6

0.8

1.0

1.2

1.4

1.6

1.8

2.0

2.2

2.4

0

666-F2

666-F203

666-F3

666-F4

666-F202

666-F3666-F201

2.4 neutral density glass filter for checking photometric accuracy

applicationTo measure photometric accuracy (absorbance), the filterreduces the light beam from the spectrophotometer. an ab-sorbancevalue(abs)canbededucedfromthelightextinctioncausedbythefilter.

pRoduct descRiptionHellmaanalyticsneutraldensityglassfiltersaremadefromfilter materials produced by Schott ag, which were selec-ted on account of their homogeneity and stability. Thankstoarelativelyconstanttransmittancewithinthewavelengthrangeof405nmto800nm, theyhavebeenused tocheckphotometricaccuracyandlinearityinthevisiblewavelengthrange(>405nm)fordecades.

notesIfyouhaveseveralneutraldensityglassfilterswithdifferentnominal absorbances, you can check the linearity of yourabsorbancescalebyplottingtheabsorbancevaluesmeasu-redforeachwavelengthagainstthemeasurementvaluesontheDakkScalibrationcertificateinadiagram.

Typicalspectrumforneutraldensityglassfilters,measuredwithaslitwidthof1nm

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of our products by writing to: [email protected]

2. Glass fIlters photometRic accuRacy

aRticle no. 666F2-39,666F3-38,666F4-37,666F201-39,666F202-36,666F203-36

application TestingthephotometricaccuracyintheVisrange(405nmto890nm)

contentf2: neutraldensityglassfilterNg11(0.25abs),f3:neutraldensityglassfilterNg5(0.5abs),f4: neutraldensityglassfilterNg4(1.0abs),f201:neutraldensityglassfilterNg11(0.3abs)f202: neutraldensityglassfilterNg3(1.5abs),f203:neutraldensityglassfilterNg3(2.0abs)

standaRd ceRtification Wavelengths:440;465;546.1;590;635nmslit width: 1nm

possible ceRtificationall wavelengths possible from 405 to 890 nm.alsopossibleabove890nm,withHellmaanalyticscalibrationcertificateslit width: allpossibleupto5nm

Gla

ss f

ilte

Rs

Gla

ss f

ilte

Rs

1716

2.5 Glass filter sets

Specificallycreatedtomeetcustomerrequirements,Hellmaanalyticsglassfiltersetsconsistofexistingindividualfilterssuitableforstandardorcustomvalidationprocedures.

Toensurethat filterscanbeeasily identified, thesetnum-ber isengravedoneachfilter frame.Theabsorbance/peakpositionvaluesmeasuredforeachfiltercanbefoundontheDakkScalibrationcertificateprovided.

2.6 General usage guidelines for glass filters

glass filtersaremadeofglassdopedwithmetal ions/rareearthmetals,whichareassembledstress-freeinblackan-odizedprecisionframesmadeofaluminum.Theyaredesi-gnedtofitintoallspectrophotometersequippedwithahol-derforstandardcuvetteswitha10-mmopticalpathlength.To ensure that filters can be easily identified, each filterframe is engraved with the filter type and serial number.Detailsoftheabsorbanceandpeakpositionvaluesmeasu-redforeachfiltercanbe foundontherespectivecalibrati-oncertificate.pleaseensurethatyoudonottouchtheglasssurfacesofthefilter.Dirt,dust,anddamagecansignificant-ly impair the accuracy of measurement results. anodizedaluminumholdersshouldnotcome intocontactwithacidsoralkalis.

stoRaGeafter use, we strongly recommend storing the filters atroom temperature, in their packaging, and in a dry, dust-freearea.

otheR factoRs that may influence measuRementsDirt(e.g.fingerprints)andduston,ordamage(scratches,cor-rosion)to,polishedopticalsurfacescansignificantlyimpairtheaccuracy of measurement results. always store the filters intheiroriginalpackagingandprotect theopticalwindows fromcontamination.Onlyhandlethefiltersbytheirframes.

cleaninGDirtoftenaccumulatesonopticalsurfacesasaresultofre-gularuse.This isbest removedusinga lint-freeclothandalcoholorHellmanexIIIcleaningsolution.

influence of tempeRatuRe on measuRementsTemperature has a very small influence on certified measu-rementvalues,andtemperaturesbetween20°Cund24°Cfallwithin themeasurementuncertaintystatedonthecalibrationcertificate. Measurements should therefore be taken in thisrange tokeepanypotential temperature influenceon there-sultstoaminimum.

aRticle no. 666S000 666S001

applicationCompleteglassFilterSetfortestingthephotometricaccuracyundthewavelengthaccuracyofthespectrophotometer

glassFilterSetfortestingthewavelengthaccuracyandthephotometricaccuracyofthespectrophotometer

content

F1:holmiumoxideglassfilterF2:neutraldensityglassfilterNg11(0.25abs)F3:neutraldensityglassfilterNg5(0.5abs)F4:neutraldensityglassfilterNg4(1.0abs)F0:filterframewithoutglass(referencefilter)

F3:neutraldensityglassfilterNg5(0.5abs)F4:neutraldensityglassfilterNg4(1.0abs)F7:didymiumglassfilter(0.5-1.0abs)

standaRd ceRtification

f1: holmium oxide glass filter:Wavelength accuracy at:279;361;453;536;638nmslit width: 1nm

f2, f3, f4: neutral density glass filter:wavelengths: 440;465;546.1;590;635nmslit width: 1nm

f3, f4: neutral density glass filter:wavelengths: 440;465;546.1;590;635nmslit width:1nm

f7: didymium glass filter:wavelength accuracy at: 329;472;512;681;875nmphotometric accuracy:from0.5to1absslit width: 1nm

possible ceRtification

Wavelength accuracy; possible wavelengths: 287;418;445;460nmslit width:upto2nmrecommended

photometric accuracy:wavelengths: allpossiblefrom405to890nmalsopossibleabove890nm,withHellmaanalyticscalibrationcertificateslit width: allpossibleupto5nm

f3, f4: neutral density glass filter:wavelength accuracy: allpossiblefrom405to890nm.alsopossibleabove890nm,withHellmaanalyticscalibrationcertificateslit width: allpossibleupto5nm

didymium glass filter:wavelength accuracy:possible wavelengths: 291;302;430;446;482;626nmslit width: upto2nmrecommendedphotometric accuracy:

possible wavelengths:from270to290nmandfrom310to320nmslit width: allpossibleupto5nm

for further sets, please see our product overview on pages 48 to 50.

hellmanex iii cleaning concentrate// the cleaning concentrate significantly lowers surface

tension and ensures optimum surface wetting even when used on complex shapes or on very large surfaces

// it is extremely easy to rinse off, leaving no trace of the cleaner on the glass surface

// it uses special surfactants that cause no absorption above 288 nm, and does not interfere with subsequent measurements in the uV/Vis range// it is extremely compatible with materials, reduces corrosion of glass and surfaces, and preserves optical surface quality

aRticle no. pRoduct

9-307-011-4-507 HellmanexIII,1l(1.3kg)

9-307-011-5-507 HellmanexIII,10l(14kg)

9-307-011-6-507 HellmanexIII,25l(35kg)

oRdeR hotline+49 7631 [email protected]

2. Glass fIlters sets

Gla

ss f

ilte

Rs

Gla

ss f

ilte

Rs

1918

steps to taKe befoRe peRfoRminG calibRation With Glass filteRs

1. Warmupthespectrophotometeruntilthecorrectoperatingtemperaturehasbeenreachedandremainsconstant(e.g.foronehour),takingcaretoobservethedevicemanufacturer’sguidelines.

2. Makesurethatyouuseastablecuvetteholderfor10mmstandardcuvettestomeasurethefilters,asthisistheonlywaytoguaranteethebestpositioningofthefiltersinthelightpath.Checkthattheholderissecureandstableinthesamplecompartment.

3. Tobeginwith,carryoutabaselinecorrectionwithanemptysamplecompartment.

4. CheckthatthefilteriscorrectlypositionedinthelightpathbyfirstplacingemptyfilterholderF0inthecuvetteholder.TheF0markingmustbevisiblefromabove.ensurethatallfilterframesarealwayspositionedinthesameway,i.e.withserialnumbersfacingthelightsource.

5. Checkthatthedevice’sdisplayhasnotchanged.Inspectrophotometerswithverylargebeams,themeasurementbeammaytouchthefilterframe(beamclipping).Ifthisisthecase,youwillnoticeachangeinthedevice’sdisplay.

» Ifnecessary,adjusttheheightofthecuvetteholderuntilthelightbeamshinesthroughtheaperture unimpeded.Tohelp,youcanswitchthedevice’smeasurementbeamtovisiblei.e.byadjustingthemonochromatorto

500nm.Theremaybeotherwaysofdoingthisdependingonthedevice. » Ifthelightbeamtouchesthesidesoftheaperture,adjustthehorizontalpositionofthecuvetteholderuntil thelightbeamshinesthroughthecenteroftheaperture.Thefilterframeiscorrectlypositionedifthedisplayvalues,

fromthezeroadjustmentperformedinstep3(baselinecorrection),donotchange.Inrarecaseswherethezero readingcannotberetainedafterinsertingtheemptyfilterframeandcarryingouttheaboveproceduresitispermissible tore-zerotheinstrumentwiththeemptyfilterframeinplaceandthencontinuewiththefiltermeasurements.

6. Carryoutthefiltermeasurementinaclosedsamplecompartmentascarefullyasyouwouldcarryoutasamplemeasurement

(opensamplecompartmentsproduceincorrectresults).

7. pleasenotethat,ifyouareusingadiodearrayspectrophotometerwithastand-alonecuvetteholderconnectedviaafiber-opticcable,extraneouslightandvibrations(e.g.movementoffiber-opticcables)mayalsoimpairtheaccuracyofmeasurementresults.

steps foR checKinG WaVelenGth accuRacy With a holmium oxide Glass oR didymium Glass filteR

1. Firstofall,followthe‘stepstotakebeforeperformingcalibrationwithglassfilters’.

2. Runthescanprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectascanningrangethatcoversallofthepeakslistedonthefilter’scalibrationcertificate.

3. Setyourspectrophotometertothemeasurementparametersthatappearonthecalibrationcertificateprovided.Selecttheslowestscanningspeedandasmalldatainterval.

4. Ifpossible,carryoutabaselinecorrection.

5. Measurementsaretakenusinganairblankwhichmeans,whichmeansthatthereferencecuvetteholderremainsemptyindoublebeamphotometers,whileareferencemeasurementistakenusingtheemptycuvetteholderinsinglebeamphotometers.

6. Inserttheholmiumoxideglassordidymiumglassfilterintothecuvetteholder.ensurethatthefilterisinsertedintotheholderasfarasitwillgo,andthatthefilterIDcanbeseenfromabove.Thefiltersmustalwaysbepositionedinthecuvetteholderinthesameway,i.e.withtheserialnumberfacingthelightsource.

7. Startthemeasurement.

8. Calculatethepositionsofthepeaksatthewavelengthsstatedonthecalibrationcertificate.(Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors).

9. Compareyourmeasurementvalueswiththecertifiedones.

measuRement paRameteRs foR checKinG WaVelenGth accuRacyensurethatyouhaveselectedthecorrectmeasurementpara-metersbeforeplottingtheabsorbancecurvetocalculatepeakpositions. Incorrect parameters may distort the absorbancecurveandthusshifttheactualpositionsofpeaks.pleaseusethesettingsstatedontheaccompanyingcalibrationcertificate.Itshouldbenotedthatchangingtheslitwidthofthespectro-photometercancausetheabsorptionmaximatoshiftslightly.Ignoreanyinfluencethatthespectralbandwidthfrom1nmto2nmhasonpeakpositions.peakheights,however,mayvarygreatlyfollowingchangestotheslitwidthduetotheirnarrownature. as a result, filters for checking wavelength accuracyareusuallyunsuitableforcheckingabsorbanceaccuracy.

»Generally speaking, filters can also be mea-sured using a slit width that differs from the in-formation provided on the calibration certificate. however, please note that large slit widths will prevent weaker peaks from being resolved. in ca-ses of doubt, it is therefore advisable to choose as small a slit width as possible. We recommend taking several measurements and then using the mean value to avoid errors during evaluation.«

thomas brenn,productManager

We look forward to hearing any comments,

criticism or suggestions you may have:

[email protected]

2. Glass fIlters2.7 calibration with glass filters

pRepaRations

Watch a video of theindividual steps here.



WaVelenGth accuRacy

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Therearemanydifferentmakes,modelsand

designsofspectrophotometer,thesestepsare

onlygeneralguidelines.pleaserefertoyour

manufacturer'sinformationtogetmoredetails.


designsofspectrometer,thesestepsareonly

generalguidelines.pleaserefertoyour


2120

steps foR checKinG photometRic accuRacy With neutRal density Glass oR didymium Glass filteRs

1. Firstofall,followthe‘stepstotakebeforeperformingcalibrationwithglassfilters’.

2. Runthewavelengthselectionprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectthewa-velengthsprovidedonthecalibrationcertificate.

3. Setyourspectrophotometertothemeasurementparametersthatappearonthecalibrationcertificateprovided.

4. adjusttozero.

5. Measurementsaretakenusinganairblankwhichmeansthatthereferencecuvetteholderremainsemptyindoublebeamspec-trophotometers,whileareferencemeasurementistakenusingtheemptycuvetteholderinsinglebeamspectrophotometers.

6. Inserttheneutraldensityglassordidymiumglassfilterintothecuvetteholder.ensurethatthefilterisinsertedintotheholderasfarasitwillgo,andthatthefilterIDcanbeseenfromabove.Thefiltersmustalwaysbepositionedinthecuvetteholderinthesameway,i.e.withtheserialnumberfacingthelightsource.

7. Starttheprogramformeasuringtheabsorbancevaluesatthewavelengthsstatedonthecalibrationcertificate.(Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors).


inteRpRetinG the measuRement Results of Glass filteRs foR checKinG photometRic and WaVelenGth accuRacy Themeasurementuncertaintiesthatappearonthecalibrationcertificate only refer to measurements conducted by Hellmaanalytics and apply solely to the measurement conditions atthe company (spectrophotometer used, environmental influ-encessuchas temperature,airhumidity,user influence,andreferencematerialsused).

The smallest possible measurement uncertainty can then bederivedbystatisticallycombiningthemeasurementuncertaintystatedonthecalibrationcertificateandalloftheuser’suncer-tainty contributions. These include the wavelength scale tole-rance of the spectrophotometer used and other influences onmeasurementaccuracy(environmentalfactorssuchastempe-rature,airhumidity,user influence,etc.).For further literatureoncorrectlycalculatingmeasurementuncertainty,pleaserefertochapter8ofthisusermanual.

see the faqs for more details.


photometRic accuRacy


inteRpRetinG measuRement Results


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»in practice it is easier to simply add up uncer-tainty contributions than to combine their stati-stics. however, the method used to determine measurement uncertainty depends on the speci-fications of your quality system and your measu-rement accuracy requirements.«

carola steinger,ChemicalLaboratory Technician





2322

3.1 holmium oxide liquid filter for checking wavelength accuracy

applicationTomeasurewavelengthaccuracy, the filterreduces the lightbeamofthespectrophotometertoagreaterextentatcertainwavelengths(peaks).Ideally,anystandardsusedtodeterminewavelengthaccuracyshouldhavenarrow,well-definedpeaksatavarietyofwavelengthsintheUVandvisiblerange.

pRoduct descRiptionTheholmiumoxideliquidfilterconsistsofasolutionofhol-miumoxidedissolvedinperchloricacid.Thisfilterisideallysuited tochecking thewavelengthaccuracyofspectropho-tometersintheUVandvisiblerange.Ithasaspectrumwitha variety of characteristic, very well-defined peaks in therangebetween240nmand650nm.

3.2 potassium dichromate liquid filter for checking photometric accuracy

applicationphotometricaccuracy (absorbance) ismeasuredbyshiningalight beam from the spectrophotometer through the insertedfilter.anabsorbancevalue(abs)canbededucedfromthelightextinctioncausedbythefilter.

pRoduct descRiptionpotassium dichromate in perchloric acid is very suitable forcheckingthephotometricaccuracyofspectrophotometers. Inthe UV range, the potassium dichromate spectrum has cha-racteristicmaximaat257nmand350nmandcharacteristicminimaat235nmand313nm.Thespectrumreachesapla-teauat430nm,whichisusedtodeterminephotometricaccu-racy in the visible range. Hellma analytics purchases the re-ferencematerial forthis filterdirectly fromNIST(SRM®935a

“potassiumDichromate”).Thefiltersolutionsaremanufactu-redinstrictcompliancewithNISTrequirementsandfilledun-der controlled conditions. The cuvettes are then immediatelyfusedtobecomeairtight.

notesas the filters are certified individually, measurement resultsare free fromsystematicerrorsmadewhenpreparingsoluti-onsandwithregardstotheopticalpathlengthofthecuvette.The measurement values of reference filter UV14 (perchloricacidmeasuredagainstanairblank)appearseparatelyontheDakkS calibration certificate. To check absorbance linearity,take measurements using potassium dichromate filters withdifferentconcentrations.plottheabsorbancevaluesmeasuredforeachfilterandwavelengthagainstthemeasurementvaluesthatappearontheDakkScalibrationcertificateinagraph.

230 430 530 6303300

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485.25

451.47

416.30

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287.20278.12

241.15

aRticle no. 667005

application TestingthewavelengthaccuracyaccordingtotheeuropeanpharmacopoeiaintheUV/Visrange

content Holmiumoxideinperchloricacid

standaRd ceRtification wavelengths:241;287;361;536;640nmslit width: 1nm

possible ceRtifcation additional wavelengths: 250;278;333;345;386;416;451;468;485nmslit width:allupto2nm,abovepeaksbecomeindistinct

aRticle no. 667020,667040,667060,667080,6670100,667600,667014(referencefilter)

application TestingthephotometricaccuracyintheUVrange(235to350nm)undVisrange(measurementwavelength430nm)ataspectralbandwidthof2nmorless

content

UV20,20mgpotassiumdichromateinHCL04(0.25abs)UV40,40mgpotassiumdichromateinHCL04(0.5abs)UV60,60mgpotassiumdichromateinHCL04(0.75abs),inaccordancewiththeeuropeanpharmacopoeiaUV80,80mgpotassiumdichromateinHCL04(1.0abs)UV0100,100mgpotassiumdichromateinHCL04(1.25abs)UV600,600mgpotassiumdichromateinHCL04(1.0abs)inaccordancewiththeeuropeanpharmacopoeiaUV14,perchloricacid(HCLO4),referencefilter

standaRd ceRtificationat uV20 - uV100 wavelengths: 235;257;313;350nmat uV600 wavelength: 430nmslit width:2nm

possible ceRtifcation wavelength: fixedslit width:allupto2nm

Typicalspectrumofholmiumoxidedissolvedinperchloricacid,measuredataslitwidthof1nm.

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Typicalspectrumofa0.006%potassiumdichromatesolution

Typicalspectrumofa0.06%potassiumdichromatesolution

We would be interested to hear about your

experiences with reference materials.

Write to us at [email protected]

3. lIquId fIlters WaVelenGth accuRacy

3. lIquId fIlters photometRic accuRacy

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3.3 liquid filters for checking for stray light

applicationIn a spectrophotometer, stray light is light that passes bythesampleand fallsdirectlyon thedetector.Thiscan lead toincorrect measurement results. Stray light may be caused byscatteringordiffraction,bypooropticalalignment,theuseofin-correctordamagedcuvettes,incorrectlyfittedsamplingacces-soriesordamagedsealsaroundalight-tightsamplechamber.Straylightisproblematic,asitreducestherangeofmeasurab-leabsorbanceandimpairsthelinearitybetweenconcentrationand absorbance. Cut-off filters (filters with a strictly definedspectrum)arerequiredtocheckthedeviceforstraylight.

pRoduct descRiptionHellma analytics stray light filters do not allow light topassthroughthembelowacertainwavelength(cut-offwa-velength).anytransmittancevaluesdisplayedinthecut-offwavelengthrangethereforerepresentstraylight.

Due to their strictly defined spectrum, potassium chloridefilters, sodium iodide filters, and sodium nitrite filters areideally suited to qualifying the stray light level of spectro-photometersincompliancewithpharmacopoeias.Thestepsarethesameforallstraylightfilters.

3.4 liquid filters for checking spectral resolution

applicationRegularly checking the spectral resolution of spectrophoto-meters ensures, for example, that neighbouring peaks areresolvedandnotsuperimposedonthepeaksofborderingwa-velengths.Thisalsopreventsabsorbanceerrors.

pRoduct descRiptionThetolueneinhexaneliquidfilterhasaprominentpointinitsspectrum,whichisexcellentfordeterminingthespectralreso-lutionand/oractualslitwidthofspectrophotometersincomp-liancewiththeeuropeanpharmacopoeia.

notesaspectrophotometer’sspectralresolutionisverycloselycon-nected to the correct slit width setting and characterized byitsabilitytoresolve(recognize)twoverycloselyrelatedpeaks.Thesmallertheslitandcorrespondingspectralbandwidth,thehighertheresolution.

aRticle no. 667001,667010,667011,667012(referencefilter)

application TestingforstraylightintheUVrange(UV1accordingtotheeuropeanpharmacopoeia)(atwavelengthsfrom198nmto370nm,dependingonthefilterselected)

content

UV1,potassiumchlorideinpurewaterUV10,sodiumiodideinpurewaterUV11,sodiumiodideinpurewaterUV12,purewater(referencefilter)


uV1:cut-offat200nmuV10:cut-offat259nmuV11: cut-offat385nmslit width: 2nm

possible ceRtifcation wavelengths: fixedpossible slit widths: allupto5nm

aRticle no. 667006,667009

application Testingtheresolutionaccodingtotheeuropeanpharmacopoeia

content UV6,tolueneinn-hexaneUV9,n-hexane(referencefilter)


wavelength: scanfrom265to270nmslit width: 0.5;1.0;2.0nmwithHellmaanalyticsCalibrationCertificate(noDakkSCalibrationCertificate)

possible ceRtifcation

wavelength: fixedpossible slit widths: from0.5to3nm

200 250 300 350 400 450 500

KCl(667-UV1)

NaI(667-UV10)

NaNO2(667-UV11)

H2O(667-UV12)

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slit 3.0

slit 2.0

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Typicalspectraoftheliquidfiltertoluene,measuredwithdifferentslitwidths

3. lIquId fIlters stRay liGht

3. lIquId fIlters spectRal Resolution liquid filteRs

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»as a rule of thumb, the slit width should be no more than 10% of the peak width at half maximum in order to be able to determine its absorbance with an accuracy of 99.5%. two peaks are deemed to be resolved separately if the minimum absorbance between them amounts to less than 80% of the maximum peak. impairments to the spectrophoto-meter’s spectral resolution will cause two different peaks to be shown as a combined peak, leading to inaccurate measurement results.«

timo Rapp,ChemicalLaboratoryTechnician

2726

3.5 liquid filter set compliant with the european pharmacopoeia

The complete liquid filter set 667003 was compiled on thebasis of european pharmacopoeia requirements and con-tainsallfiltersrequiredtocarryoutacompletespectropho-tometercheck.

checking photometric accuracy (uV60, uV600, uV14)

checking wavelength accuracy (uV5) checking for stray light (uV1 and uV12)

checking spectral resolution (uV6 and uV9)

all liquid filters consist of reference materials that arefilled into precision Hellma cuvettes made of quartz glass(SUpRaSIL®). These cuvettes are permanently sealed, andthecompletesetisdeliveredinahigh-qualitystoragebox.Toensureeasyidentification,eachfilterisengravedwithitsserialnumber.ThecalibrationvaluesmeasuredforeachfiltercanbefoundontheDakkSandHellmaanalyticscalibrationcertifica-tesprovided.

3.6 liquid filter set compliant with the united states pharmacopeia

3.7 General usage guidelines for liquid filters

Liquidfiltersbearamarkingononesideshowingthechemi-calformulaofthesubstancecontainedinthecuvette. Ifafil-terbreaks,pleaseobservethecodesofconductandsafetyin-structionsthatapplytothissubstance.Thisinformationcanbefoundinthesafetyinstructions.Up-to-datesafetyinstructionsforallsubstancesusedtomanufactureliquidfiltersareavai-lableatwww.hellma-analytics.com/download.

stoRaGeafter use, we strongly recommend storing the filters atroom temperature, in their packaging, and in a dry, dust-freearea.Liquidfiltersmustnotbeexposedtotemperatu-resbelow4°Corabove40°C.Thisalsoapplieswhentrans-portinganddeliveringliquidfiltersforrecertification.

otheR factoRs that may influence measuRementsDirt (e.g. fingerprints) and dust on, or damage (scratches,corrosion)to,polishedsurfacescansignificantly impairtheaccuracy of measurement results. always store the filtersin theiroriginalpackagingandprotect theopticalwindowsfromcontamination.Onlyhandlethefiltersbytheircapsormattsurfaces.

cleaninGDirtoftenaccumulatesonopticalsurfacesasaresultofregularuse.Thisisbestremovedusingalint-freeclothandalcoholorHellmanexIIIcleaningsolution.

influence of tempeRatuRe on measuRementsTemperaturehasaverysmall influenceoncertifiedmeasu-rementvalues,andtemperaturesofbetween20°Cund24°Cfall within the measurement uncertainty stated on the cali-brationcertificate.Measurementsshould thereforebe takeninthisrangetokeepanypotentialtemperatureinfluenceontheresultstoaminimum.

Want other products or services?

Write to us at

[email protected]

for further information, please see page 17

aRticle no. 667003

application Completefiltersetfortestingthespectrophotometeraccordingtotheeuropeanpharmacopoeiaphotometricaccuracy,wavelengthaccuracy,straylightandresolution

content

UV1,potassiumchlorideinpurewaterUV12,purewater(referencefilter)UV5,holmiumoxideinHClO4UV6,tolueneinn-hexaneUV9,n-hexane(referencefilter)UV60,60mgpotassiumdichromateinHClO4UV600,600mgpotassiumdichromateinHClO4UV14,perchloricacid(referencefilter)


uV1/uV12: wavelength: 200nm(cut-off)uV5: wavelenght:241;287;361;536;640nm;slit width:1nmuV6/uV9: wavelength: scanfrom265to270nm; slit width: 0.5;1.0;2.0nmuV60: approx.0.75abs; wavelengths:235;257;313;350nm; slith width: 2nmuV600: approx.1.0abs; wavelength: 430nm; slit width: 2nm


uV1/uV12: wavelength: fixed; slit width:allupto5nmuV5: wavelenght: 250,278,333,345,386,416,451,468,485nm; slit width: allupto2nm;above,peaksbecomeindistinctuV6/uV9: wavelength: fixed;slit width:0.5to3nmuV60: wavelength: fixed; slit width:allupto2nmuV600: wavelength: fixed; slit width:allupto2nm

aRticle no. 667004

application FiltersetfortestingthespectrophotometeraccordingtoUSp851,photometricaccuracyandwavelengthaccuracy

content

F2:neutraldensityglassfilter(0.25abs)F3:neutraldensityglassfilter(0.5abs)F4:neutraldensityglassfilter(1.0abs)F0:filterframewithoutglass(referencefilter)UV60:60mgpotassiumdichromateinHClO4UV14:perchloricacid(HClO4,referencefilter)UV5:holmiumoxideinHClO4


f2, f3 and f4: wavelengths: 440,465,546.1,590,635; slit width:1nmuV60: wavelengths:235,257,313,350; slit width: 2nmuV5: wavelengths: 241,250,278,287,333,345,361,385,416,452,468,485,536,640nm;slit width: 1 nm


f2, f3, f4: wavelengths: allpossiblefrom405to890nmalsopossibleabove890nmwithHellmaanalyticsCalibrationCertificateslit width:allpossibleupto5nmuV60: wavelength:fixed; slit width: upto2nmuV5: wavelength: fixed, slit width: allupto2nm;above,peaksbecomeindistinct

always take great care when placing li-quid filters in the sample holder of your spectrophotometer. Wherever possible, only touch filters by their caps or matt sides. take care not to touch the polis-hed surfaces. the filters are fragile and should be handled with the utmost care.

3. lIquId fIlters sets

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steps to taKe befoRe peRfoRminG calibRation With liquid filteRs

1. Warmupthespectrophotometeruntilthecorrectoperatingtemperaturehasbeenreachedandremainsconstant(e.g.foronehour),takingcaretoobservethedevicemanufacturer’sguidelines.

2. Makesurethatyouuseastablecuvetteholderfor10mmstandardcuvettestomeasuretheliquidfilters,asthisistheonlywaytoguaranteethebestpositioningofthefiltersinthelightpath.Checkthattheholderissecureandstableinthesamplecompartment.

3. Thefiltersshouldalwaysbepositionedinthecuvetteholdersinthesameway,i.e.withtheHellmaletteringfacingthelightsource.Thelightbeammustpassthroughthepartofthefilterfilledwithliquid(solution).

4. Carryoutthefiltermeasurementinaclosedsamplecompartmentascarefullyasyouwouldcarryoutasamplemeasure-ment(opensamplecompartmentsproduceincorrectresults).

5. pleasenotethat,ifyouareusingadiodearrayspectrophotometerwithastand-alonecuvetteholderconnectedviaafiber-opticcable,extraneouslight,extraneouslightandvibrations(e.g.movementoffiber-opticcables)mayalsoimpairtheaccuracyofmeasurementresults.

measuRement paRameteRs foR checKinG WaVelenGth accuRacyensurethatyouhaveselectedthecorrectmeasurementpa-rameters before plotting the absorbance curve to calcula-te peak positions. Incorrect parameters may distort the ab-sorbancecurveandthusshift theactualpositionsofpeaks.pleaseusethesettingsstatedontheaccompanyingcalibra-tioncertificate.Itshouldbenotedthatchangingtheslitwidthof thespectrophotometercancause theabsorptionmaximatoshiftslightly. Ignoreany influencethat thespectralband-widthfrom1nmto2nmhasonpeakpositions.peakheights,however,mayvarygreatlyfollowingchangestotheslitwidthdue to their narrow nature. as a result, filters for checkingwavelengthaccuracyareusuallyunsuitableforcheckingab-sorbanceaccuracy.

share your experience of our

products by writing to:

[email protected]


steps foR checKinG WaVelenGth accuRacy With a holmium oxide liquid filteR

1. Firstofall,followthe‘stepstotakebeforeperformingcalibrationwithliquidfilters’.

2. Runthescanprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectascanningrangethatcoversallofthepeakslistedonthefilter’scalibrationcertificate.

3. Setyourspectrophotometertothemeasurementparametersthatappearonthecalibrationcertificateprovided.Selecttheslowestscanningspeedandasmalldatainterval.


5. MeasurementsaretakenagainsttheperchloricacidblankUV14,whichmeansthatthereferencecuvetteholderremainsemptyindoublebeamphotometers,whileareferencemeasurementistakenusingtheemptycuvetteholderinsinglebeamphotometers.

6. Inserttheholmiumoxideliquidfilterintothecuvetteholder,observingthegeneralusageguidelinesforliquidfilters.Thefiltersshouldalwaysbepositionedinthecuvetteholdersinthesameway,i.e.withtheHellmaletteringfacingthelightsource.

7. Startthemeasurement.

8. Calculatethepositionsofthepeaksatthewavelengthsstatedonthecalibrationcertificate.(Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors).


3. lIquId fIlters3.8 calibration with liquid filters

pRepaRations


WaVelenGth accuRacy

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»Generally speaking, filters can also be mea-sured using a slit width that differs from the in-formation provided on the calibration certificate. however, please note that large slit widths will prevent weaker peaks from being resolved. in cases of doubt, it is therefore advisable to choose as small a slit width as possible. We recommend taking several measurements and then using the mean value to avoid errors during evaluation.«

carola steinger,ChemicalLaboratory Technician






designsofspectrometer,thesestepsare



3130

steps foR checKinG photometRic accuRacy With a potassium dichRomate liquid filteR


2. Runthewavelengthselectionprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectthewavelengthsprovidedonthecalibrationcertificate.

3. Setyourspectrophotometertothemeasurementparametersthatappearonthecalibrationcertificateprovided.

4. adjusttozero.

5. asageneralrule,measurementsaretakenusingareferencefilterfilledwithperchloricacidobservingthegeneralusageguidelinesforliquidfilters.Thefiltersshouldalwaysbepositionedinthecuvetteholdersinthesameway,i.e.withtheHellmaanalyticsletteringfacingthelightsource.

6. Measurementsinasinglebeamspectrophotometer:Carefullyinserttheperchloricacidreferencefilterprovidedintothecu-vetteholder.Startthemeasurement.Next,measurethecertifiedreferencematerial,whichcontainspotassiumdichromatedissolvedinperchloricacid.Thensubtractthereferencemeasurementvaluesfromthemeasurementvaluesofthecertifiedreferencematerial.Orsetzero/blanktheinstrumentwiththeperchloricacidreferencefilterUV14fitted,thenreplacewiththecertfiedreferencematerialandreadthevaluedirectlyfromthedisplay.

7. Measurementsinadoublebeamspectrophotometer:Carefullyinsertthecertifiedreferencematerial,whichcontainspotassiumdichromatedissolvedinperchloricacid,intothesampleholder,andtheperchloricacidreferencefilterintothereferencesampleholder.

8. Starttheprogramformeasuringtheabsorbancevaluesatthewavelengthsstatedonthecalibrationcertificate.(Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors).


measuRement paRameteRs foR checKinG photometRic accuRacyasthedifferencebetweenthemaximaandminimaintheab-sorbancespectrumisrelativelylarge,thepotassiumdichro-mateliquidfiltersmayalsobemeasuredwithaslitwidththatdiffers from the one on the calibration certificate. However,pleasenote thatusing largeslitwidths (>2nm)mayresultinslightdeviationsfromthevaluesstatedonthecalibrationcertificate.Incasesofdoubt,itisthereforeadvisabletochoo-setheslitwidthquotedonthecalibrationcertificate.Were-commend takingseveralmeasurementsand thenusing themeanvaluetoavoiderrorsduringevaluation.

inteRpRetinG the measuRement Results of liquid filteRs foR checKinG photometRic and WaVelenGth accuRacy The measurement uncertainties that appear on the calibrati-oncertificateonlyrefertomeasurementsconductedbyHellmaanalyticsandapplysolelytothemeasurementconditionsatthecompany (spectrophotometer used, environmental influencessuchastemperature,airhumidity,userinfluence,andreferencematerialsused).

The smallest possible measurement uncertainty that canbeachievedbytheusercanthenbederivedbystatistical-ly combining the measurement uncertainty stated on thecalibration certificate with all the user’s uncertainty con-tributions, such as the wavelength scale tolerance of thespectrophotometerusedandother influencesonmeasu-rementaccuracy(environmentalfactorssuchastempera-ture, air humidity, user influence, etc.). For further lite-rature on correctly calculating measurement uncertainty,pleaserefertochapter8ofthisusermanual.


birgit Kehl,HeadHellmaanalyticscalibrationlaboratory



[email protected]





inteRpRetinG measuRement Results

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measuRement paRameteRs foR checKinG stRay liGht leVelFor a realistic calculation of the stray light level, choose afilter with a cut-off wavelength as close above the requiredwavelength as possible. This enables the stray light test tobecarriedoutat thewavelengthatwhichthestray light fil-ter can fully absorb light. The remaining transmittancedisplayed by the device at the measurement wavelengthrepresents the stray light level. Since this value differs de-pending on the properties of the measuring system, filterscan only be certified with regard to their suitability for useas a stray light filter. Certification therefore demonstra-tes that filters have virtually full absorbance in the measu-ring range and steep peaks at high transmittance values.

You can check the lower absorbance range of your spectro-photometerusingreferencefilter667-UV12,whichisfilledwithultrapure water. The filter’s absorbance characteristics from200 nm to NIR are practically only determined by the reflec-tion losseson the twoair/glasssurfaces. Youcancheck yourdevice’sdisplayatverylowabsorbancevaluesagainstthecer-tifiedvaluesat198nm,200nm,300nmand400nm.Ifyourre-sultsdiffersignificantlyfromthecertifiedvalues,particularlyifthemeasuredvaluesaresmallerthan0.02a,youshouldcon-tactthedevicemanufacturer.

inteRpRetinG measuRement Results When checKinG foR stRay liGhtToestimatethesamplemeasurementerrorduetostraylight,comparethecalculatedstraylightleveltothesignalstrengthfromthesamplemeasurement.Forexample,astraylightva-lueof0.1%transmittanceandasamplewithanabsorbanceofaround1abswouldequatetoameasurementerrorduetostraylightofaround0.4%.Ifyouhavecalculatedastraylightlevel that is considerably higher than the level stated in thedevice specifications, check whether extraneous light couldhaveinterferedwiththisresult.Ifyoucanruleoutextraneouslight,pleasecontactaservicetechnician.

please note that the reference for the measuring system to be tested is not provided by the entire transmittance characteristic of the stray light filter, but solely by the transmittance value measured in the range of virtually full absorbance.

steps foR checKinG the stRay liGht leVel


2. Runthescanprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectascanningrangethatcoversallofthevalueslistedonthefilter’scalibrationcertificate.

3. Setyourspectrophotometertothemeasurementparametersquotedonthecalibrationcertificateprovided

4. Setthespectrophotometertoawavelengthofapprox.20nmabovethecut-offwavelengthforthestraylightfilterused(forpotassi-umchloride(UV1),forexample,startat220nm)andscanuptothewavelengthforwhichyouwishtodeterminethestraylightlevel.


6. asageneralrule,takethemeasurementsusingareferencefilterfilledwithwater(UV12)observingthegeneralusagegui-delinesforliquidfilters.Thefiltersshouldalwaysbepositionedinthecuvetteholdersinthesameway,i.e.withtheHellmaanalyticsletteringfacingthelightsource.

7. Measurementsinasinglebeamspectrophotometer:Carefullyinsertthereferencefilterprovidedintothecuvetteholder.Startthemeasurement.Next,measurethecertifiedreferencematerial.Thensubtractthereferencemeasurementvaluesfromthemeasurementvaluesofthecertifiedreferencematerial.

8. Measurementsinadoublebeamspectrophotometer:Carefullyinsertthecertifiedreferencematerialintothesampleholderandthereferencefilterintothereferencesampleholder.Startthemeasurement.

9. Scanuptothewavelengthforwhichyouwishtodeterminethestraylightlevel.

Thelightlevel(remainingtransmittancevalue)measuredbelowthecut-offwavelengthrepresentsstraylight.10.


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measuRement paRameteRs When checKinG spectRal ResolutionWhen measuring spectral resolution, the liquid filter ab-sorbs the light beam from the spectrophotometer to si-gnificantly different extents in a narrow wavelength range(5nm).Thefilterwillshowaclearmaximumandminimumwithinthenarrowrange.afterplacingtheliquidfilterinthespectrophotometer, run the scan program in the definedwavelengthrangeanddividethemaximumpeakmeasuredatλmax=269nmbytheminimumpeakmeasuredatλmin266nm.Theresultingratiorepresentstheabsorbanceratio,whichisdirectlylinkedtotheslitwidth.Iftheratioisconsi-derablylower(e.g.15%),pleasecontactthedevicemanufac-turer.pleasenote,however,thattheresultalsodependsonthe measurement conditions. Therefore, please make surethatyouselectasufficiently long integration time,particu-larlyifusingasmallslitwidth.

inteRpRetinG measuRement Results When checKinG spectRal ResolutionRegulatory codes or internal applications and measuringproceduresmayplacerequirementsontheratiosthatmustbe achieved. In addition, comparing calculated ratios withcertified valuesmayprovidean indicationof theactual slitwidthofthedeviceused.

steps foR checKinG spectRal Resolution


2. Runthescanprogramonyourspectrophotometer,observingtheguidelinesintheusermanual.Selectascanningrangethatcoversbothoftherequiredpeaks.

3. Setyourspectrophotometertothemeasurementparametersstatedonthecalibrationcertificateprovided.


5. Takethemeasurementusingareferencefilterfilledwithhexane–ifthespectrumiscorrectedtozeroat300nm,measu-rementscanalsobetakenusingair–observingthegeneralusageguidelinesforliquidfilters.Thefiltersshouldalwaysbepositionedinthecuvetteholdersinthesameway,i.e.withtheHellmaanalyticsletteringfacingthelightsource.

6. Measurementsinasinglebeamspectrophotometer:Carefullyinsertthehexanereferencefilterprovidedintothecuvetteholder.Startthemeasurement.Next,measurethecertifiedreferencematerial,whichcontainstolueneinhexane.Thensubtractthereferencemeasurementvaluesfromthemeasurementvaluesofthecertifiedreferencematerial.

7. Measurementsinadoublebeamspectrophotometer:Carefullyinsertthetolueneinhexaneliquidfilterintothesampleholderandthehexanereferencefilterintothereferencesampleholder.Startthemeasurement.

8. Measuretheactualminimumoftheabsorbancevaluesat266nmandtheactualmaximumat269nm.(Takeseveralmeasure-mentsandthenusethemeanofthemeasuredvaluestoavoiderrors).

9. Ifpossible,carryoutabaselinecorrectioneachtimeyouchangetheslitwidth.

Determinetheratiousingbothofthemeasuredvalues,asstatedonthecalibrationcertificateprovided.10.

please note that the filter set for de-termining spectral resolution does not fall within our scope of accredit-ation, and therefore cannot be issued with a dakks calibration certificate or calibration mark.

slit Width absoRbance Ratio

0,25 2,3

0,5 2,2

1,0 2,0

2,0 1,4

3,0 1,1

4,0 1,0

absoRbance Ratio of maximum/minimum peaK in Relation to slit Width

(seeStandardsandBestpracticeinabsorptionSpectrometry,editedbyC.Burgess&T.Frost)



[email protected]


spectRal Resolution


inteRpRetinG the measuRement Results

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4. certIfIed reference plates photometRic accuRacy

4. certIfIed reference plates photometRic and WaVelenGth accuRacy

4.1 Reference plates for checking photometric accuracy

applicationTheHellmaanalyticsreferenceplate666R013canbeusedtocheckthephotometricaccuracyofmicrotitreplatereaders.

pRoduct descRiptionThe reference plate dimensions are suitable for a 96-wellmicrotitre plate with a diameter of 6.6 mm per window (H14.5mmxD125mmxL85.5mm).eachofthefiveneutraldensityglassfilters(fieldsS1–S5)inthereferenceplatecanmeasuretheabsorbancevaluefor16windows.Theother16windowsdonotcontainglass(S0)andserveasreferences.

notesThereferenceplatehasfiveneutraldensityglassfilterswithdifferentnominalabsorbancevalues,allowingyoutocheckthe linearity of your absorbance scale by plotting the ab-sorbancevaluesmeasuredforeachwavelengthagainstthemeasurementvaluesontheDakkScalibrationcertificateinadiagram.

4.2 Reference plate for checking photometric and wavelength accuracy

applicationTheHellmaanalyticsreferenceplate666R113canbeusedtocheckthephotometricandwavelengthaccuracyofmicrotitreplatereaders.

pRoduct descRiptionThe reference plate dimensions are suitable for a 96-wellmicrotest plate with a diameter of 6.6 mm per window (H14.5mmxD125mmxL85.5mm).eachofthefourneutraldensityglassfiltersused(fieldsS1–S4)canmeasuretheab-sorbancevaluefor16windows.Holmiumoxideglass(S5)isusedtotestthewavelengthaccuracyin16windowswhileafurther 16 windows (S0) do not contain glass and serve asreferences.

notesThe reference plate has four neutral density glass filterswith different nominal absorbance values, allowing you tocheckthelinearityofyourabsorbancescalebyplottingtheabsorbance values measured for each wavelength againstthemeasurementvaluesontheDakkScalibrationcertifica-teinadiagram.

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aRticle no. 666R013

application Referenceplateformicroplatereadersfortestingthephotometricaccuracy

content

neutraldensityglassfilterNg11(0.25abs)neutraldensityglassfilterNg5(0.5abs)neutraldensityglassfilterNg4(1.0abs)neutraldensityglassfilterNg3(1.5abs)neutraldensityglassfilterNg3(2.5abs)framewithoutglass(referencefilter)

standaRd ceRtification photometric accuracy certified at wavelengths:405;450;490;650nmat 8 points in a row

possible ceRtificationwavelengths: allpossiblefrom405to890nmalsopossibleabove890nm,butonlywithaHellmaanalyticsCalibrationCertificateslit widths: allpossibleupto5nm

aRticle no. 666R113

application Referenceplateformicroplatereadersfortestingthewavelengthaccuracyandthephotometricaccuracy

content

neutraldensityglassfilterNg5(0.5abs)neutraldensityglassfilterNg4(1.0abs)neutraldensityglassfilterNg3(1.5abs)neutraldensityglassfilterNg3(2.0abs)holmiumoxideglassfilterframewithoutglass(referencefilter)

standaRd ceRtificationphotometric accuracy certified at 8 points in row at wavelengths:405;450;490;650nmwavelength accuracy certified at: 279;361;453;536;638nmslit width: 1nm


photometric accuracy:additionalwavelengths:allpossiblefrom405to890nmalsopossibleabove890nm,butonlywithaHellmaanalyticsCalibrationCertificateslit widths: allpossibleupto5nmwavelength accuracy:additional possible wavelengths: 287;418;445;460nmslit widths: allupto2nmrecommended

Want other products or services?

Write to us at

[email protected]

3938

4.3 General usage guidelines for reference plates

Referenceplatesaremadeofglassdopedwithmetal ionsorrareearthmetals,whichisannealedandassembledinblackanodized precision frames made of aluminum. They are de-signed to fit into all microtitre plate readers. To ensure easyidentification,each referenceplate isengravedwith the refe-renceplatetypeandserialnumber.Detailsoftheabsorbanceandpeakpositionvaluesmeasuredforeachfiltercanbefoundontherespectivecalibrationcertificate.pleaseensurethatyoudonottouchtheglasssurfacesofthefilter.Dirt,dust,andda-mage can significantly impair the accuracy of measurementresults.anodizedaluminumframesshouldnotcomeintocon-tactwithacidsoralkalis.

stoRaGeafter use, we recommend storing reference plates at roomtemperature,intheirpackaging,andinadry,dust-freearea.

otheR factoRs that may influence measuRementsDirt(e.g.fingerprints)andduston,ordamage(scratches,cor-rosion)to,polishedsurfacescansignificantly impairtheaccu-racyofmeasurementresults.alwaysstorereferenceplatesintheiroriginalpackagingandprotect theopticalwindows fromcontamination.Onlyhandlereferenceplatesbytheirframes.

cleaninGDirtoftenaccumulatesonopticalsurfacesasaresultofre-gularuse.This isbest removedusinga lint-freeclothandalcohol.

influence of tempeRatuRe on measuRementsTemperaturehasaverysmallinfluenceoncertifiedmeasure-ment values.Measurements takenat temperaturesbetween20°Cund24°Cfallwithinthemeasurementuncertaintystatedonthecalibrationcertificate.Measurementsshouldthereforebetakeninthisrangetokeepanypotentialtemperatureinflu-enceontheresultstoaminimum.

4. certIfIed reference plates for mIcrotItre plate readers

steps to taKe befoRe peRfoRminG calibRation With RefeRence plates

1. Warmupthemicrotitreplatereaderuntilthecorrectoperatingtemperaturehasbeenreachedandremainsconstant(e.g.foronehour),takingcaretoobservethedevicemanufacturer’sguidelines.

2. Tobeginwith,carryoutabaselinecorrectionwithanemptysamplecompartment.

3. Checkthatthereferenceplateiscorrectlypositionedinthelightpathbyfirstmeasuringthewindowswithoutglass(usuallyrows1and2).Thelabelshowingthereferenceplatetypemustbevisiblefromabove.

4. Checkthatthedevice’sdisplayhasnotchanged.Inmicrotitreplateswithverylargebeams,themeasurementbeammaytouchthewindowframe.Ifthisisthecase,youwillnoticeachangeinthedevice’sdisplay.

» Ifnecessary,adjustthepositionofthereferenceplateholderuntilthelightbeamshines throughtheemptywindowunimpeded. » Thereferenceplateiscorrectlypositionedifthedisplayvaluesfromthezeroadjustmentperformedinstep2

(baselinecorrection)donotchange.

5. Carryoutthefiltermeasurementinaclosedsamplecompartmentascarefullyasyouwouldcarryoutasamplemeasure-ment(opensamplecompartmentsproduceincorrectresults).

ideas, criticism, experiences?

We want to hear your thoughts!

[email protected]

4. certIfIed reference plates4.4 calibration with reference plates

pRepaRations Therearemanydifferentmakes,models

anddesignsofspectrometer,thesestepsare



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inteRpRetinG measuRement Results With RefeRence plates foR checKinG photometRic and WaVelenGth accuRacy The measurement uncertainties stated on the calibrationcertificate only refer to measurements conducted by Hellmaanalytics and apply solely to the measurement conditions atthe company (spectrophotometer used, environmental influ-encessuchas temperature,airhumidity,user influence,andreferencematerialsused).

The smallest possible measurement uncertainty that canbeachievedbytheusercanthenbederivedbystatisticallycombiningthemeasurementuncertaintystatedonthecali-brationcertificatewithalltheuser’suncertaintycontributi-ons,suchasthewavelengthscaletoleranceofthemicrotestplate reader used and other influences on measurementaccuracy (environmental factors such as temperature, airhumidity,userinfluence,etc.).Forfurtherliteratureoncor-rectlycalculatingmeasurementuncertainty,pleaserefertochapter8ofthisusermanual.

measuRement paRameteRs foR checKinG photometRic accuRacygenerally speaking, reference plates can also be measuredusing a slit width that differs from the information providedonthecalibrationcertificate.However,pleasenotethatusinglargeslitwidthsmayresult inslightdeviations fromtheva-luesstatedonthecalibrationcertificate.Incasesofdoubt,itisthereforeadvisabletochooseassmallaslitwidthaspos-sible.Werecommendtakingseveralmeasurementsandthenusingthemeanvaluetoavoiderrorsduringevaluation.

measuRement paRameteRs foR checKinG WaVelenGth accuRacyensure that you have selected the correct measurementparameters before plotting the absorbance curve to cal-culate peak positions. Incorrect parameters may distorttheabsorbancecurveandthusshift theactualpositionsofpeaks.pleaseusethesettingsstatedontheaccompanyingcalibrationcertificate. Itshouldbenoted thatchanging theslit width of the microtitre plate reader can cause the ab-sorbance maxima to shift slightly. Ignore any influence

thatthespectralbandwidthfrom1nmto2nmhasonpeakpositions.peakheights,however,mayvarygreatlyfollowingchanges to the slit width due to their narrow nature. as aresult, filters forcheckingwavelengthaccuracyareusuallyunsuitableforcheckingabsorbanceaccuracy.

steps foR checKinG WaVelenGth accuRacy With RefeRence plates

1. Firstofall,followthe‘stepstotakebeforeperformingcalibrationwithreferenceplates’.

2. Runthescanprogramonyourmicrotitreplatereader,observingtheguidelinesintheusermanual.Selectascanningran-gethatcoversallofthepeakslistedonthefilter’scalibrationcertificate.

3. Setyourmicroplatereadertothemeasurementparametersquotedonthecalibrationcertificateprovided.Selecttheslowestscanningspeedandasmalldatainterval.


5. placethereferenceplateintheplateholder.ensurethatthefilterIDisvisiblefromabove.Referenceplatesmustalwaysbepositionedintheplateholdersinthesameway.

6. Startthemeasurementforthepositionswhereholmiumoxideglassfiltersareinserted(usuallyS5).

7. Calculatethepositionsofthepeaksatthewavelengthsstatedonthecalibrationcertificate.

8. Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors.




birgit Kehl,HeadHellmaanalyticscalibrationlaboratory




WaVelenGth accuRacy

steps foR checKinG photometRic accuRacy With RefeRence plates

1. Firstofall,followthe‘stepstotakebeforeperformingcalibrationwithreferenceplates’.

2. Runthewavelengthselectionprogramonyourmicrotitreplatereader,observingtheguidelinesintheusermanual.Selectthewavelengthsstatedonthecalibrationcertificate.

3. Setyourmicrotestplatereadertothemeasurementparametersquotedonthecalibrationcertificateprovided.

4. adjusttozero.

5. placethereferenceplateintheplateholder.ensurethatthereferenceplateIDisvisiblefromabove.Referenceplatesmustalwaysbepositionedintheplateholdersinthesameway.

6. Starttheprogramformeasuringabsorbancevaluesatthewavelengthsstatedonthecalibrationcertificate–thepositionsmeasuredarethosewhereneutraldensityglassfiltersareinserted.

7. Takeseveralmeasurementsandthenusethemeanofthemeasuredvaluestoavoiderrors.







designsofspectrometer,thesestepsare



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5. recertIfIcatIon

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ReceRtification inteRVals foR RefeRence mateRials

as is the case for all measuring devices, the referencematerials used to verify spectrophotometers must alsobecheckedand recertifiedat regular intervals (see forex-ampleISO9001:2008“ControlofMonitoringandMeasuringequipment”).Thisallowsyoutoensurethatyouconsistent-lyfulfillyourin-housequalityrequirementsandguaranteeshighlevelsofaccuracyandreliabilityinyourmeasurements.

The length of intervals between the recertification of re-ference materials depends on how frequently materialsare used, the wear associated with this, accuracy require-ments, and the requirements of a company’s internal au-diting. Therefore, only clients themselves can determinethese. In general, a recertification interval of 12 months isrecommendedforcheckingandrecertifyingglassfiltersdu-ringthefirsttwoyearsofuse,withanintervalof24monthsthereafter. We recommend verifying and recertifying liquidfilterswithinamaximumof12months.IntervalsshouldbespecifiedindividuallyinaccordancewithyourQasystem.

RetuRninG youR RefeRence mateRials foR ReceRtification

30-yeaR WaRRanty

allofourreferencematerialscomewitha30-yearwarranty,providedthattheyareregularlyrecertified(atleasteverytwo years) at the Hellma analytics calibration laboratory.Certifiedreferencematerials(filters)sentforrecertificationarecarefullycleanedandrecertifiedbeforebeingsentbackwith a new DakkS calibration certificate and calibrationmark. Damaged filters and filters that deviate significantlyfrom nominal values are usually replaced in consultationwiththecustomer.

1. yeaR 2. yeaR 3. yeaR 4. yeaR 5. yeaR 6. yeaR 7. yeaR 8. yeaR

liquid filteRs

ReCeRTIFYINgeVeRY12 months

Glass filteRs

1. yeaR 2. yeaR 3. yeaR 4. yeaR 5. yeaR 6. yeaR 7. yeaR 8. yeaR

ReCeRTIFYINgeVeRY24 months

01.Completethereturnshipmentforminfull.Whenreturningseveralfiltersorfiltersets,pleaseuseoneformforeach.

04.FiltersarecleanedandrecertifiedintheHellmaanalyticsca-librationlaboratory.Ifnecessary,filterswillalsoberepairedorexchangedfollowingaconsultation.

05.you will receive your filter with a new dakks calibration certificate or a hellma analytics calibration certificate in the case of filters for verifying spectral resolution.

03.SendyourfiltertotheHellmaanalyticscalibrationlabora-toryviayourlocalHellmaoffice.Usetheaddresslabelprintedontherearofthereturnship-mentformtodoso.

02.encloseacopyofthecurrentcalibrationcertificate.

12 MONTHS12 MONTHS

12 MONTHS

24 MONTHS 24 MONTHS 24 MONTHS

COmPAny:

nAmeOfCOntACtPeRSOn:

StReet/StReetnumBeR:

POStAlCODe/CIty:

COuntRy:

emAIl:

PHOne:

tHISADDReSSIS:SHIPPInGADDReSS BIllInGADDReSS

ARtIClenumBeR/fIlteRtyPe*:

Set/SeRIAlnumBeR*:

yOuRORDeRnumBeR:


RetuRnfORm

PleASenOte!manyqualitysystemsstipulatearegular

inspectionofthereferencematerialsfor

anidealmeasuringcertainty.Itisrecom-

mendtorecertifyglassfilterservery24

monthsandliquidfiltersevery12months.

RecertificationintheaccreditedHellmaAnalyticsCalibrationlaboratory

I’dliketoreceivethefollowingrecertification

DAkkSCalibrationcertificate (according to the already existent calibration certificate; please enclose a copy of it)

IndividualrecertificationwithDAkkSCalibrationcertificate Please specify:

SlItWIDtH/S:

WAVelenGtH/S:

OtHeR:

Additionaloptions I’dliketoreceiveadocumentationofthemeasureddatauponreceipt(priortocleaning)

measurementreportoftheabsorptionvalues for neutral density glass filters (at 1 nm slit width and the following wavelengths 440, 465, 546.1, 590, 635) (charge 18.90 € – depending on the country and exchange rate) Individual documentation of the measureddatauponreceiptcombinedinameasurementreport (charges depending on the type of filter and extent of measurement) SlItWIDtH/S:

WAVelenGtH/S:

Documentation of the measureddatauponreceiptwithaDAkkSCalibrationcertificate for all measurement parameters, according to the recertification stated above (charges depending on the type of filter and extent of measurement)

thankyouverymuchfortrustingtheHellmaAnalyticsCalibrationlaboratory!

Pleasecompletethisformandencloseitwiththereturnshipment.

*You’ll find this information on your calibration certificate

Returnformavailablefor

download:

www.hellma.com/return

4544

6. faqs

6.1 Why do holmium oxide glass filters become cloudy? Will this interfere with the measurement?The glass material used for this filter is somewhat hygro-scopic,whichmeans that the filtersbecomecoatedwithakindofwaterfilm.Thefilmdoesnotinterferewithmeasure-mentsorchangethecharacteristicpeakpositionsofholmi-umoxide.Thefiltercanbeeasilywipeddownusingalcoholand a soft cloth. The filter should generally be stored in adryplace.6.2 how long can a calibration standard be used for in total?Depending on the conditions in which they are used andstored,aswellashowtheyaremaintained,filtersusuallylastformanyyears.Werecommendhavingfiltersregularlyrecer-tifiedsothatanysignsofdeteriorationcanberecognizedatanearlystage.6.3 how often should filters be recertified?Certifiedreferencematerialsshouldberecertifiedat regu-lar intervalstocheckthatthevaluesstatedonthecalibra-tion certificate are still valid. It is up to the user to decideon the regularityof these intervals,whichshould take intoaccount theuse,storageandusageconditionsof the filterinthelaboratory.Toestablishastatisticaldatabaseforde-termining recertification intervals, we recommend havingallreferencematerialsrecertifiedat leastevery12monthsduringtheirfirsttwoyearsofuse,andthenselectingasui-tablerecertificationintervalbasedonthevaluesmeasured.(pleaseseechapter9).6.4 What do the tolerances on the calibration certificate tell us and how can they be correctly interpreted?The measurement uncertainties that appear on calibrati-on certificates only refer to measurements conducted byHellmaanalyticsandapplysolelytothemeasurementcon-ditions at the company (spectrophotometer used, environ-mental influences such as temperature, air humidity, userinfluence,referencematerialsused,etc.).Consequently,themeasurementuncertaintiesoftheNISTreferencematerialsusedtoensuretraceabilityhavebeenmathematicallycom-binedwith themeasurementuncertaintystatisticscalcula-tedbyHellmaanalytics.Thevalueprovided is thereforeanexpandedmeasurementuncertainty(doublestandarddevia-tion,coveragefactork=2).Thismeansthattheactualvalueis95%certaintofallwithinthisrange.Tocorrectlycalcula-tethemeasurementuncertaintiesvalidfortheirmeasuringsystem, reference material users should follow the samesteps,mathematically/statistically combining themeasure-ment uncertainties provided with the measurement uncer-tainty statistics they have calculated themselves for a par-ticularspectrophotometerandrelevantconditions(seeISO/IeCguide98-3:2008‘guidetotheexpressionofUncertaintyinMeasurement’).

6.5 What is a baseline correction?Baseline corrections are carried out with an empty cuvetteholder tocompensate for the lamps.Since lampsemit lightat different strengths at various wavelengths, baseline cor-rections(alsoknownasautozero)arecarriedouttodetermi-neazerovalue.Baselinecorrectionsareusuallyperformedautomaticallywhenthespectrophotometerisstartedup,butcanalsobecarriedoutmanually.6.6 What is background correction?Backgroundcorrectioniscarriedouttoeliminateanyinflu-encesthatextendbeyondthesample’sproperties.Indoub-le beam photometers, background correction is performedbysimultaneouslymeasuringthecomparisoncuvetteinthereferencebeampath.Thiscomparisoncuvetteusuallycon-tainspuresolvent.Insinglebeamphotometers,backgroundcorrectioniscarriedoutbeforetheactualsamplemeasure-mentistakenbymeasuringthecomparisoncuvette.Theva-luesobtainedforthecomparisoncuvettearethendeductedfromthevaluesofthesamplemeasurement.6.7 Why does the calibration certificate for the filter set used to determine spectral resolution look different to other calibration certificates? Determiningspectralresolutiondoesnotfallwithinourscopeofaccreditation.The filterset fordeterminingspectral resolutionthereforecannotbeissuedwithaDakkScalibrationcertificateorcalibrationmark.Thatiswhythiscalibrationcertificatelooksdif-ferentfromothercalibrationcertificatesforfiltersets.6.8 Why does hellma analytics no longer offer potassium dichromate filters for checking photometric accuracy with sulfuric acid as a solvent, as described in the european pharmacopoeia?Inthepast,certifiedreferencematerialsforcheckingphoto-metricaccuracy intheUVrangecontainedasolutionofpo-tassiumdichromate insulfuricacidandweremanufacturedin strict compliance with european pharmacopoeia require-ments. Over a number of years, Hellma analytics noticeda continuous decrease in the absorbance values of the ‘po-tassium dichromate dissolved in sulfuric acid’ filter duringdaily calibrations. We do not have a sufficient explanationforwhythishappens,butweassumethatthecomparativelyhigh ionic strength of sulfuric acid causes mixed chromium(VI)complexestoform.Tocompensateforthisbehavior,overwhichwehavenocontrol,filterswouldneedtoberecertifiedmuchmoreregularly.anotherpossibilitywouldbepreparingnewsolutionseverytimethespectrophotometer ischecked.asasimplealternative,weofferaliquidfilterthatuses‘po-tassiumdichromatedissolvedinperchloricacid’.Thistypeofliquidfilterforcheckingphotometricaccuracyhasprovenit-selfasareliableandverystablestandardformanyyears.Nochangesinabsorbancepropertiescomparabletothoseofthe

sulfuricacidmodelareknownforthisfilter.Hellmaanalyticscuvettesarepermanentlysealed,eliminatingconcernsaboutthe toxicity of perchloric acid. Furthermore, the europeanpharmacopoeiastates that “suitablecertified referencema-terials”mayalsobeused,whichundoubtedlyapplies toourperchloricacidsolventmodel.Thismodelalsocontainsafor-mulationdescribedbyNIST.6.9 Why does the weight of potassium dichromate filters seem to change after every recertification?Due to measurement uncertainties, measurement valuesmay fall within a specific range. This leads to an apparentchange in weight from qualification to qualification, as theinitial weight is calculated directly from the measured ab-sorbance values. earlier versions of regulatory codes stipu-lated that filters for checking photometric accuracy had tocontain60.06mg/lpotassiumdichromate,andallowedato-lerance of 0.01 abs. More current versions of the europeanpharmacopoeiahavereplaced this verystrictprovision,nowacceptingweightsbetween57.0mg/land63.0mg/l.Thespe-cific absorbance calculated (see european pharmacopoeia,chapter2.2.25)isnowstatedwithamarginoftolerance.6.10 Why are these peaks measured for certifying holmium oxide glass and didymium glass filters?Measurement errors are low in medium to high transmit-tanceranges.asaresult,peaks in therange from0abs to1.0abs(correspondsto100%-Tto10%T)arepreferredforcertification.6.11 how do i calculate my measurement uncertainty?The measurement uncertainties stated on the calibration cer-tificate only refer to measurements conducted by Hellmaanalyticsandapplysolelytothemeasurementconditionsatthecompany (spectrophotometer used, environmental influencessuch as temperature, air humidity, user influence, referencematerialsused,etc.).Thesmallestpossiblemeasurementun-certaintythatcanbeachievedbytheusercanthenbederivedbystatistically combining the measurementuncertainty stated onthecalibrationcertificatewithalltheuser’suncertaintycontribu-tions,suchasthewavelengthscaletoleranceofthespectropho-tometer used and other influences on measurement accuracy(environmentalfactorssuchastemperature,airhumidity,userinfluence,etc.).

example of calculatinG standaRd measuRement unceRtainty foR a neutRal density Glass filteR (hiGhly simplified): the calibRation ceRtificate sta-tes the folloWinG measuRement Values and measu-Rement unceRtainties:

here, a wavelength of 440 nm produces the following parameters:target measurement value (xs) : 0,2542 absexpanded measurement uncertainty: +/- 0,0024 abs (coverage factor k=2)standard measurement uncertainty (xa): +/- 0,0012 abs

Next, you must calculate the measuring error specific to yourspectrophotometer(xb)–refertotheoperatinginstructionsformoredetails–anddefineavalueforthemeasuringerrorduetoenvironmentalinfluencesatyourcompany(xu)(suchastempe-ratureandairhumidity).

example of measuring error parameters:spectrophotometer (xb): +/- 0,01 absenvironmental influences (xu): +/- 0,001 abscalculating standard measurement uncertainty (mu):mu = √xa² + xb² + xu² = 0,0101expanded measurement uncertainty is calculated by multip-lying this value by coverage factor k.

asshownhere,inpracticeitisofteneasiertosimplyaddupun-certaintycontributionsthantocombinetheirstatistics.However,the method used to determine measurement uncertainty de-pends on the specifications of your quality system and yourmeasurement accuracy requirements. For further literatureoncorrectlycalculatingmeasurementuncertainty,pleaserefertotherecommendationsforfurtherreadinginchapter8ofthisusermanual.

seRial numbeR 3524

optische dichte (abs)optical density (abs)

440nm 465nm 546.1nm 590nm 635nm

measuRed Value 666-f2 0.2542

±0.00240.2254±0.0024

0.2254±0.0024

0.2415±0.0024

0.2416±0.0024

4746

7. Glossary 8. recommendatIons for further readInG

abbreviations:

a: absorbanceastm: americanSocietyforTestingandMaterialsbG: SpecifictermforSchottglassdab: Deutschesarzneibuch(germanpharmacopoeia)dakks: Deutscheakkreditierungsstelle(Nationalaccreditation bodyfortheFederalRepublicofgermany)daR: Deutscherakkreditierungsrat (germanaccreditationbody)dKd: DeutscherKalibrierdienst(germancalibrationbody)ph. eur.:europeanpharmacopoeiafaqs: FrequentlyaskedquestionsGlp: goodlaboratorypracticeGmp: goodmanufacturingpracticei: Intensityi0: Originalintensity0: Originalk: Coveragefactorformeasurementuncertaintyλmax: Maximumpeakatdefinedwavelengthλmin: MinimumpeakatdefinedwavelengthniR: Near-infrarednist: NationalInstituteofStandardsandTechnologyptb: physikalisch-TechnischeBundesanstalt (germany’snationalmetrologyinstitute)sRm®: StandardReferenceMaterial (registeredtrademarkofNIST)usp: UnitedStatespharmacopeiauV: UltravioletVis: Visible(visiblewavelengthrange)

absorbance (abs): Whenlightfallsonorpassesthroughasample,thequantityofabsorbed light is equal to the difference between the originalintensityI0andtheintensityIafterinteractionwiththesample.Thisisbecausepartoftheirradiatedlightistransferredtothemolecules,causingthebeamtohaveasmalleroutputwhenitexitsthesample.Theextenttowhichlightisabsorbedisdeter-minedbytheprinciplesoftheBeer-Lambertlaw.Theamountofabsorbedlightcanbeexpressedastransmittance(seedefiniti-on)orabsorbance.absorbanceisdefinedasabs=-logT.accordingtotherelevantstandard,thisparameterisreferredtoasspectralopticaldensityontransmittance(“opticaldensity”).

optical density: seeabsorbance

Visible range: partof theoptical spectrum thatstretches from380nmto780nmofthewavelengthrangeofelectromagneticra-diation.Thisrangeisgenerallyreferredtoaslight.Thisistheonlyrangeinwhichthehumaneyecan‘see’electromagneticradiation.

spectral resolution: Thisreferstoameasuringsystem’sabilitytoseparateindividualwavelengthranges.

spectral bandwidth: Wavelengthrangethatappearswithacontinuum at the exit slit when the monochromator is ex-posed to irradiation. Spectral bandwidth is determined bythebandwidthofemittedradiationwherethelighthasrea-chedhalfthemaximumintensity.

spectral optical density on transmittance:seeabsorbance

transmittance (t): When light falls on or passes through asample,thequantityofabsorbedlightisequaltothedifferencebetweentheoriginal intensity I0and the intensity Iafter inter-action with the sample. This is because part of the irradiatedlightistransferredtothemolecules,causingthebeamtohaveasmalleroutputwhenitexitsthesample.Theextenttowhichlight is absorbed is determined by the principles of the Beer-Lambertlaw.Theamountofabsorbedlightcanbeexpressedastransmittance (see definition) or absorbance. Transmittance isnormallyexpressedasafractionof1orasapercentage,andisdefinedasfollows:T=I/I0or%T=(I/I0)*100.

ultraviolet range (uV range):alsoknownasUVradiation,thisistheshort-wavepartoftheopticalradiationspectrum.UVradiati-onhasawavelengthrangeof100nmto380nm.

Wavelength: Wavelength isthedistancebetweentwo identical,adjacentcorrespondingpointsofthesamewavephaseatacer-tainpointintime.

standards and best practice in absorption spectrometry;edited by C. Burgess and T. Frost UVSg, ISBN 0-632-05313-5BlackwellService

qualitätssicherung in der analytischen chemie;WernerFunk,VeraDammann,gerhildDonnevert;ISBN-10:3-527-31112-2;Verlag:WILeY-VCH

ISO/IeC guide 98-3:2008; evaluation of measurement data – Guide to the expression of uncertainty in measurement NISTSpecialpublication260-54StandardReferenceMaterials:certification and use of acidic potassium dichromate solutions as an ultraviolet absorbance standard – sRm 935

NIST Special publication 260-116 Standard ReferenceMaterials: Glass filters as a standard Reference material for spectrophotometry – selection, preparation, certification, and use of sRm 930 and sRm 1930

NIST Special publication 260-102: Standard ReferenceMaterials:holmium oxide solution Wavelength standard from 240 to 640 nm – sRm 2034

european pharmacopoeia (ph.eur.)

dKd3


products by writing to: [email protected]

type mateRial WaVelenGth nm aRticle-no.

Glass filter for testing the wavelength accuracy

666-f1 HolmiumOxideglassFilterF1 279;361;453;536;638 666f1-339

666-f7W DidymiumglassFilterF7W 329;472;512;681;875 666f7W-323

Glass filter for testing the photometric accuracy

666-f2 NeutralDensityglassFilterF2(Nominalvalueoftheabsorption0.25)

440;465;546,1;590;635 666f2-39


440;465;546,1;590;635 666f201-39


440;465;546,1;590;635 666f3-38


440;465;546,1;590;635 666f4-37


440;465;546,1;590;635 666f202-36


440;465;546,1;590;635 666f203-36

666-f7a NeutralDensityglassFilterF7a(Nominalvalueoftheabsorptionapprox.0.5–1.0)

270;280;297;320;340 666f7a-323

Glass filter for testing the wavelength accuracy and the photometric accuracy

666-f7 DidymiumglassFilterF7 a:270;280;297;320;340W:329;472;512;681;875

666f7-323

empty filter mount

666-f0 aluminumframe 666f0-71

type consistinG of WaVelenGth nm aRticle-no.

sets for testing the wavelength accuracy and the photometric accuracy

666-s000 CompleteglassFilterSet:F1,F2,F3,F4,F0

a:440;465;546,1;590;635W:279;361;453;536;638

666s000

666-s001 glassFilterSet:F3,F4,F7

a:270;280;297;320;340;440;465;546,1;590;635W:329;472;512;681;875

666s001

666-s002 glassFilterSet:F2,F3,F4 a:440;465;546,1;590;635 666s002

666-s004 glassFilterSet:F201,F202,F203 a:440;465;546,1;590;635 666s004

666-s300 glassFilterSet:F301,F303,F390(abs:0.04;2.5;3.0)

a:440;465;546,1;590;635 666s300

666-R113666-R013

5048

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Glass filters with dakks calibration certificates liquid filters with dakks calibration certificates liquid filters with dakks calibration certificates

667-UV 1

667-UV100

667-UV 60

667-UV305

667-UV 6

667-UV200

667-UV 5667-UV 12 667-UV 14667-UV 600667-UV 9

Complete Set 667-UV003

typ inhalt WellenlänGe nm aRtiKel-nR.

flüssigfilter zur Überprüfung der fotometrischen Genauigkeit

667-uV20 20mgKaliumdichromatinHClO4(0,25abs) 235;257;313;350 667020





667-uV600 600mgKaliumdichromatinHClO4(1,0abs) 430 667600

667-uV14 perchlorsäure(Referenzfilter) 235;257;313;350 667014

667-uV301 Filter-SetfürUV-Bereich:UV60,UV14 235;257;313;350 667301

667-uV304 Filter-SetfürVis-Bereich:UV600,UV14 430 667304

667-uV305 Filter-SetfürUV/Vis-Bereich:UV60,UV600,UV14 235;257;313;350;430 667305

flüssigfilter-set zur Überprüfung der linearität der absorption

667-uV307 Filter-Set:UV20,UV40,UV60,UV80,UV100,UV14 235;257;313;350 667307

flüssigfilter zur Überprüfung der Wellenlängengenauigkeit

667-uV5 Holmiumoxidinperchlorsäure 241;287;361;536;640 667005

667-uV400 Filter-Set:UV05,UV14 241;287;361;536;640 667400

flüssigfilter zur Überprüfung von streulicht

667-uV1 KaliumchloridinReinstwasser 200(cut-off) 667001

667-uV10 NatriumiodidinReinstwasser 259(cut-off) 667010

667-uV11 NatriumnitritinReinstwasser 385(cut-off) 667011

667-uV12 Reinstwasser(Referenzfilter) 198;200;300;400 667012

667-uV100 Filter-Set:UV1,UV12 200(cut-off) 667100



667-uV103 Filter-Set:UV1,UV10,UV11,UV12 200;259;385(cut-off) 667103

a:WavelengthsforabsorbanceW:WavelengthsforwavelengthaccuracyS:WavelengthsforstraylightR:Wavelengthsforspectralresolution *withHellmaanalyticscalibrationcertificate

Reference plates for qualifying microplate readers with dakks calibration certificates

666-F0 666-F1 666-F2 666-F3 666-F4 666-F203666-F202666-F201 666-F7

Complete Set 666-S000 SingleFilters

a:WavelengthsforabsorbanceW:Wavelengthsforwavelengthaccuracy

type content WaVelenGth nm aRticle-no.

liquid filter for testing the photometric accuracy

667-uV20 20mgpotassiumdichromateinHClO4(0.25abs) 235;257;313;350 667020





667-uV600 600mgpotassiumdichromateinHClO4(1.0abs) 430 667600

667-uV14 perchloricacid(HClO4),referencefilter 235;257;313;350 667014

667-uV301 FilterSetforUVrange:UV60,UV14 235;257;313;350 667301

667-uV304 FilterSetforVisrange:UV600,UV14 430 667304

667-uV305 FilterSetforUV/Visrange:UV60,UV600,UV14 235;257;313;350;430 667305

liquid filter set for testing the linearity of the absorption

667-uV307 Filter-Set:UV20,UV40,UV60,UV80,UV100,UV14 235;257;313;350 667307

liquid filter for testing the wavelength accuracy

667-uV5 Holmiumoxideinperchloricacid 241;287;361;536;640 667005

667-uV400 FilterSet:UV05,UV14 241;287;361;536;640 667400

liquid filter for testing to stray light

667-uV1 potassiumchlorideinpurewater 200(cut-off) 667001

667-uV10 Sodiumiodideinpurewater 259(cut-off) 667010

667-uV11 Sodiumnitriteinpurewater 385(cut-off) 667011

667-uV12 purewater(referencefilter) 198;200;300;400 667012

667-uV100 FilterSetUV-100:UV1,UV12 200(cut-off) 667100



667-uV103 FilterSetUV-103:UV1,UV10,UV11,UV12 200;259;385(cut-off) 667103

667-uV104 FilterSetUV-104:UV10,UV11,UV12 259;385(cut-off) 667104

liquid filter for testing the resolution

667-uV6* Tolueneinn-hexane Scan:265–270 667006

667-uV9* n-hexane(referencefilter) Scan:265–270 667009

667-uV200* FilterSetUV-200:UV6,UV9 Scan:265–270 667200

a:WavelengthsforabsorbanceW:WavelengthsforwavelengthaccuracyS:WavelengthsforstraylightR:Wavelengthsforspectralresolution *withHellmaanalyticscalibrationcertificate

type content WaVelenGth nm aRticle-no.

liquid filter – set according to usp 851

667-uV004 F0:aluminumframe,F2:NeutralDensityglassfilter(0.25abs),F3:NeutralDensityglassfilter(0.5abs),F4:NeutralDensitiyglassfilter(1.0abs),UV60:60mgpotassiumdichromateinHClO4UV14:perchloricacid(HClO4,referencefilter,

UV5:Holmiumoxideinperchloricacid

a:440;465;546.1;590;635a:440;465;546.1;590;635a:440;465;546.1;590;635a:235;257;313;350a:235;257;313;350W:241;250;278;287;333;345;361;385;416;452;468;485;536;640

667004

complete filter set for testing the photometer according to ph.eur.

667-uV003 CompleteFilterSet:UV1,UV12,UV6,UV9,UV60,UV600,UV14,UV5

a:235;257;313;350;430W:241;287;361;536;640S:200(cut-off)R:Scan265–270

667003

type usaGe mateRialNominalvalueofabsorption

WaVelenGth nm

aRticle-no.

666-R013 tocheckphotometricaccuracy NeutralDensityglassFilterNg11(0.25),Ng5(0.5),Ng4(1.0),Ng3(1.5),(2.5)

405,450,490,650 666R013

666-R113 tocheckphotometricaccuracyandwavelengthaccuracy

NeutralDensityglassFilterNg5(0.5),Ng4(1.0),Ng3(1.5),(2.0)HolmiumOxideglassfilter

405;450;490;650279;361;453;536;638

666R113

new

Subject to change without notice 064-501-PB05-725//GB//09//2015 Printed in Germany


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notes

certified uv/vis reference materials - dialab · certified uv/vis reference materials guidelines //...

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