colu

mns A c c l a i m C 3 0 C o l u m n

Passion. Power. Productivity.

Acclaim® C30 columns provide unique selectivity and superior separations of hydrophobic, structurally related analytes.

• Highshapeselectivity

• Uniqueselectivitycomplementarytootherreversed-phasecolumns

• Compatibilitywithhighlyaqueousmobilephases

• High-qualitycolumnswithlowcolumnbleed,highefficiency,andruggedpacking

Columns for Separating Hydrophobic Structurally Related Isomers

TheAcclaimC30isdesignedtoprovidehighshapeselectivityforseparatinghydrophobicstructuralrelatedisomersanduniqueselectivitycomplementarytootherreversed-phasecolumns(e.g.,C18).Itisbasedoncova-lentmodificationofC30alkylsilaneonhigh-purity,spherical,poroussilicagel.ThecombinationofadvancedsurfacemodificationtechnologyandcarefulmatchingofC30alkylchainwiththeporesizeofthesilicasubstrateprovidesthefollowingbenefits:

10 12 14 16 18 20 22 240

100

pA

MinutesCorona® ultra™ detector

Standards in Isopropanol

Saponified Chicken Fat

1 2

3 4

5

6 7

8

9 10

11

12

13

14 Peaks: 1. SDA 2. EPA 3. ALA 4. GLA 5. DHA 6. Arach. 7. LLA 8. DPA 9. 9E, 14Z-CLA 10. ETA 11. Adrenic 12. Oleic 13. EDA 14. Erucic

28381

2

High Shape Selectivity TheAcclaimC30columnprovides

significantlygreatershapeselectiv-itycomparedtoC18columns,makingitsuitedtoseparatinghydrophobicstructuralisomers,suchascarotenoids,tocopherol,etc.Shapeselectivitycanbecharacterizedbyelutionorderofthreepolycyclicaromatichydrocar-bons(PAHs)inacetonitrile:benzo[a]pyrene(BaP),1,2:3,4:5,6:7,8-tetra-benzonaphthalene(TBN,alternatename,dibenzo[g,p]chrysene),andphenanthro[3,4-c]phenanthrene(PhPh)describedinNISTSRM869a.Asmallerα(TBN/BaP)(=kʹTBN/kʹBaP)valueindicates

highershapeselectivity.AsshowninFigure1,theAcclaimC30columnexhibitsthehighestshapeselectivityofallAcclaimreversed-phasecolumns.

Unique Selectivity Complementary to Other Reversed-Phase Columns

Selectivityisthekeyforsuccess-fulseparation.WhileC18columnsaremostcommonlyusedforsmallmoleculeseparations,otherreversed-phasecolumnswithdifferentselectivityareoftenneededforoptimalresult.Comparedtoothertypesofreversed-phasecolumns—includingC18,polar-embedded,andphenylphases—theAcclaimC30demonstratesdifferentselectivityandprovidesacomple-mentarytoolformethoddevelopment(Figure2).

100% Aqueous CompatibilityToseparatehydrophilicanalytes,

suchasorganicacidsandwater-solublevitaminsonareversed-phasecolumn,ahighlyaqueousmobilephaseisoftenrequiredtoachieveadequateretention.However,mostC18columnswilldewetundersuchcondition,resultinginunreliableresults.DuetoarigidandhighlyorderedC30alkylchain,alongwiththeuseofawiderporesilicasubstrate,AcclaimC30columnsexhibit100%aqueouscompatibility—nolossofretentionwasobservedafter50stop-flowcycles(seeFigure3).

Figure 1. Shape selectivity comparison. Note: lower (TBN/BaP) suggests higher shape selectivity.

Figure 2. The unique selectivity of the Acclaim column family.

28371

0.0

0.5

1.0

1.5

2.0

2.5

3.0

AcclaimC30

Acclaim120 C18

AcclaimPA

AcclaimPA2

AcclaimPhenyl-1

0.99 2.10 1.79 1.94 1.85

α (T

BN/B

aP)

Column Name

Columns: (4.6 × 150 mm)Mobile Phase: Acetonitrile/water 85:15 v/vTemperature: 25 °CFlow Rate: 2 mL/minInj. Volume: 5 µLDetection: UV (254 nm)Sample: NIST SRM 869a standard mix 1. Benzo[a]pyrene (BaP)

2. Phenanthro[3,4-c]phenanthrene (PhPh)

3. Dibenzo[g,p]chrysene (TBN)

α(T

BN/B

aP) =

kT́B

N/ kB́a

P

28372

1 2 3

Acclaim Phenyl

Acclaim Polar Advantage

Acclaim Polar Advantage II

Acclaim 120 C18

Minutes0 6 12 9 3

AU

0

12

3

12

3

12 3

Acclaim C30 2

3, 1 Columns: (4.6 × 150 mm)Mobile Phase: Acetonitrile/water 85:15 v/vTemperature: 25 °CFlow Rate: 2 mL/minInj. Volume: 5 µLDetection: UV (254 nm)Sample: NIST SRM 869a standard mix 1. Benzo[a]pyrene (BaP)

2. Phenanthro[3,4-c]phenanthrene (PhPh)

3. Dibenzo[g,p]chrysene (TBN)

28373

0

2

4

6

8

10

12

0 10 20 30 40 50

Retn

etion

of T

hym

ine (

min

)

Number of Stop-Flow Cycle

De-wetting Test Column: Acclaim C30, 5 µm (4.6 × 150 mm)Mobile Phase: 25 mM Ammonium acetate, pH 5Temperature: 30 °CFlow Rate: 1 mL/minInj. Volume: 5 µLDetection: UV (254 nm)Sample: 0.1 mg/mLProbe: Thymine

Stop-Flow Protocol:1. Test the column under above condition2. Stop the pump for 5 min (pump pressure drops to 0)3. Resume the flow and condition the column under above condition for 10 min4. Inject the sample and run the test under the same conditions.5. Repeat 2 through 4

Figure 3. Aqueous compatibility of the Acclaim C30 column.

3

High Quality of Column ManufacturingLowcolumnbleedisessentialfor

gooddetectionlimits,especiallyformassspectrometer(MS)andchargedaerosoldetectors(CAD).Thepropri-etarybondingprocessensurestheC30bondedphasemaintainsgoodhydrolyticstabilitywithextremelylowcolumnbleed.Ourinnovativepackingmethodprovideshighcolumnefficiencyandro-bustcolumnpackingformostchalleng-ingapplications.EachAcclaimcolumnismanufacturedtostrictspecificationstoensurecolumn-to-columnreproduc-ibility.Inaddition,eachcolumnisindividuallytestedandshippedwithaqualificationassurancereport.

Wide Range of ApplicationsTheAcclaimC30columnisa

reversed-phaseHPLCcolumn.LikeC18columns,theAcclaimC30canbeusedinabroadrangeofapplicationsincludingfoodandbeverage,chemical,environmental,pharmaceutical,academia,etc.TheAcclaim30alsooffersseveraluniquefeaturesthatsetitapartfromC18columns.First,itexhibitshighershapeselectivitysuitedtoseparationofhydrophobic,long-chain,structuralisomers(e.g.,carotenoids,steroids,etc).Second,itisfullycompatiblewithvariousaqueousbuffers,resultinginabroaderapplica-tionrange(e.g.,water-solublevitamins,organicacids,etc)andmoreflexibilityinmethoddevelopment.Third,itcanbeaviablealternativetonormal-phasecolumnsforlipidanalysis.Therefore,theAcclaimC30beusedasageneral-purposecolumnthatcomplementsC18columns,andalsoserveasaspecialtycolumnwhenaC18columnfailstoprovidesatisfactoryresults.

CarotenoidsCarotenoidsareoccurnaturally

inthechloroplastsandchromoplastsofplants,andsomefungiandbacte-ria.Theyservetwokeyrolesinplantsandalgae:1)theyabsorblightenergyforuseinphotosynthesis,and2)theyprotectchlorophyllfromphotodamage.Inhumans,fourcarotenoids(beta-car-

Figure 5. Glucocorticosteroids.

Figure 4. Carotenoids in vegetables.

otene,alpha-carotene,gamma-carotene,andbeta-cryptoxanthin)havevitaminAactivity,andcanalsoactasantioxidants.AsshowninFigure4,sixcommoncarotenoidsandchlorophyllaresepa-ratedontheAcclaimC30columnwithexcellentselectivityandresolution.

GlucocorticosteriodsGlucocorticosteroidsareagroupof

naturallyoccurringandsynthetichor-monesthatmoderateinflammationandotherstressresponses.Thesecompounds

areincludedontheWorldAnti-DopingAgency’slistofsubstancesprohibitedincompetition.Separationofthesesubstancesischallengingduetotheirstructuralsimilarity.ThemethodofMcWhinneyiscommonlyusedinclini-callaboratoriesformonitoringofthesesubstances.WhiletheoriginalmethodusesaC18column,theAcclaimC30offersimprovedresolutionandthrough-putwiththesameelutionorderusedtheliteraturemethod.Figure5showsbaselineseparationofeightglucocortico-steroidsusingtheAcclaimC30column.

28374

0 5 10 15 20 25

0

80

mAU

Minutes

C

B

A 1

2

3

4 5

6

7

8

260 800

452.4476.6

276.3

nm

Column: Acclaim C30, 5 µm (4.6 × 150 mm)LC System: UltiMate® 3000 RSMobile Phases: A. Acetonitrile B. Methanol/ethyl acetate 1:1 (v/v) C. 200 mM Acetic acid in waterGradient: Time (min) %A %B %C -5 85.0 14.5 0.5 0 85.0 14.5 0.5 2 85.0 14.5 0.5 15 65.0 34.5 0.5 25 65.0 34.5 0.5Flow: 1.50 mL/minTemperature: 30 °CInj. Volume: 8 µLDetector: Diode array; VIS 450 nm, spectra 260–800 nmSamples: A. Maize extract in acetone B. Spinach extract in acetone C. Carrot extract in acetone

Peaks: 1. Lutein 2. Zeaxanthin 3. Chlorophyll-b 4. alpha-Cryptoxanthin 5. beta-Cryptoxanthin 6. Chlorophyll-a 7. alpha-Carotene 8. beta-Carotene

Reference: “HarvetsPlus Handbook for Carotenoid Analysis,” D.B. Rodriguez-Amaya and M. Kimura, International Food Policy Research Institute, 2004.

β-Carotene

28375

0 2 4 6 8 10 12

0

100

mAU

Minutes

1 2

3 4

5

6

7

8

Column: Acclaim C30, 3 µm (3.0 × 150 mm)LC System: UltiMate 3000 RSMobile Phase: Methanol/tetrahydrofuran/water 3:25:72 (v/v)Flow: 0.50 mL/minTemperature: 30 °CInj. Volume: 2 µLDetector: Diode array; UV 240 nm

Peaks: 1. Prednisone 50 µg/mL 2. Cortisone 3. Prednisolone 4. Hydrocortisone 5. Dexamethasone 6. 6-Methylprednisolone 7. Corticosterone 8. 11-Deoxyhydrocortisone

Reference: McWhinney B C, Ward G, Hickman P E, Clin. Chem, 1996, 42, 979–981.

4

Figure 7. Analysis of cooking oil: Acclaim C30 vs. C18.

Water-soluble VitaminsWater-solublevitaminsareessential

nutrients.Theyarefoundnaturallyinfoods,andtheyarealsoaddedtofood,andformulatedsupplements.Chemi-cally,theyareaheterogeneousgroupofanions,cations,zwitterions,andneutrals.Duetothevarietyandcom-plexityofmatriceswherevitaminsarefound,theiranalysisisoftenchalleng-ing.TheAcclaimC30columndemon-stratesbothhighhydrophobicretentionandaqueouscompatibility;thisdesir-ablecombinationoffeaturesenablessimplereversed-phaseseparationofthesecomplexanalytes(Figure6).

Triglycerides in Cooking OilsCookingoilsarepurifiedlipids

fromplants,andtheyaretypicallyliquidatroomtemperature.Thesecompoundscontaintriglyceridesasmajorcomponents,andsmallquantitiesoffreefattyacids,mono-,anddiglyc-erides.Thecompositionofcookingoilsishighlycomplexduetothewidevarietyofalkylchainlength,degreeofunsaturation,origin,etc.Whilenormal-phasechromatographyisoftenusedtocharacterizeoilsbytheirhydrophilic-ity,reversed-phasechromatographyprovideshighresolutionforanalyzingmajorandminorcomponents,andobtainingadetailedfingerprint.Duetoitshighshapeselectivity,theAcclaimC30provideshigherresolutionthantheC18columnforoilanalysis(Figure7).SeparationsofseveralcookingoilsareillustratedinFigure8.

Figure 6. Water-soluble vitamins.

28376

0 5 10 12

0

160

Minutes

mAU

200 400

283.6

nm

Folic acid

1

2

3

4

6

5 7

Column: Acclaim C30, 5 µm (4.6 × 150 mm)LC System: UltiMate 3000 RSMobile Phases: A. Acetonitrile B. 7 mM Phosphoric acid + 40 mM sodium sulfate (pH 2.65)Gradient: Time (min) %A %B –3.0 0 100 0.0 0 100 2.0 0 100 10.0 25 75 12.0 25 75Flow Rate: 1.50 mL/minTemperature: 30 °CInj. Volume: 10 µLDetection: Diode array; UV 210 nm, spectra 260–400 nmSamples: Vitamin standards in mobile phase

Peaks: 1. Thiamine 25 µg/mL 2. Ascorbic acid 62 3. Niacinamide 25 4. Pyridoxine 25 5.Pantothenic acid 25 6. Folic acid 6.2 7. Riboflavin 5.0

28377

30 46 62 54 38 70 Minutes

pA

0

150

A. Acclaim C30

B. Acclaim 120 C18

Columns: A. Acclaim C30, 5 µm (4.6 × 150 mm) B. Acclaim 120 C18, 5 µm (4.6 × 150 mm)Mobile Phase: Acetonitrile (CH3CN)/Iso-propanol (IPA)/ Ammonium acetate buffer (0.1 M, pH 5.0)Gradient: Time (min) CH3CN IPA Buffer -15 90 5 5 0 90 5 5 0.1 90 5 5 60 0 95 5 70 0 95 5Temperature: 40 °CFlow Rate: 1.0 mL/minInj. Volume: 2 µLDetection: Corona ultra (gain = 100 pA; filter = medium; neb. temp. = 25 °C)Sample: Peanut oil (5 mg/mL in iso-propanol)

Figure 8. Analysis of cooking oils.

28378

0 32 724816 80Minutes

pA

0

200

A

B

C

Column: Acclaim C30, 5 µm (4.6 × 150 mm)Mobile Phase: Acetonitrile (CH3CN)/iso-propanol (IPA)/ ammonium acetate buffer (0.1 M, pH 5.0)Gradient: Time (min) CH3CN IPA Buffer –15 90 5 5 0 90 5 5 0.1 90 5 5 60 0 95 5 80 0 95 5Temperature: 40 °CFlow Rate: 1.0 mL/minInj. Volume: 2 µLDetection: Corona ultra (gain = 100 pA; filter = medium; neb. temp. = 25 °C)Samples: A. Peanut oil B. Olive oil C. Wheat germ oil (5 mg/mL in iso-propanol)

5

Figure 9. Profile of egg lecithin.

Figure 10. Phospholipids in egg lecithin.

Phospholipids in Egg LecithinThisclassoflipidsaremajor

componentsofallcellmembranesastheycanformlipidbilayers.Mostphospholipidscontainadiglyceride—aphosphategroupconnectedtoahydro-philicorganicgroupsuchascholine.Thefirstphospholipididentifiedinbiologicaltissueswaslecithinintheeggyolk.Analysisofphospholipidsischallengingduetoitscomplexity(hydrocarbonlength,degreeofsatura-tionofdiglyceridemoiety,andtypeofthehydrophilicorganicgroup).Figure9showstheprofileoflecithinfromeggyolkandsoybeanobtainedusinganAcclaimC30columnwithaCoronaultra™CAD®detector.Itindicatesthatlecithinfromeggyolkcontainsbothphospholipidsandtriglycerides,withminorimpurities.Toobtaindetailedinformationonphospholipidcomposi-tion,ahigher-resolutionmethodcanbeused(Figure10).

Omega Fatty AcidsOmega-3fattyacidsareafamily

ofunsaturatedfattyacidsthatshareafinalcarbon–carbondoublebondinthen-3position.Omega-3fattyacids,suchasα-linolenicacid(18:3,n-3;ALA),eicosapentaenoicacid(20:5,n-3;EPA),anddocosahexaenoicacid(22:6,n-3;DHA),areimportantinhumannutri-tion.ThebiologicaleffectsofOmega-3arelargelymediatedbytheirinterac-tionswiththeOmega-6fattyacids.SomemedicalresearchsuggeststhatexcessivelevelsofOmega-6relativetoOmega-3fattyacidsmayincreasetheprobabilityofanumberofdiseasesanddepression.AsshowninFigure11,theAcclaimC30columnexhibitsexcellentresolutionpowerforasuiteofcloselyrelatedOmegafattyacids.

28379

0 24 483612 60Minutes

pA

0

60 Triglycerides

Phospholipids

Degradants

Column: Acclaim C30, 5 µm (4.6 × 150 mm)Mobile Phase: Acetonitrile (CH3CN)/iso-propanol (IPA)/ ammonium acetate buffer (0.1 M, pH 5.0)Gradient: Time (min) CH3CN IPA Buffer -15 70 0 30 0 70 0 30 0.1 70 0 30 10 90 0 10 35 10 80 10 50 0 95 5 60 0 95 5Temperature: 40 °CFlow Rate: 1.0 mL/minInj. Volume: 2 µLDetection: Corona ultra (gain = 100 pA; filter = medium; neb. temp. = 25 °C)Sample: Egg lecithin – PL30S (5 mg/mL in iso-propanol)

Figure 11. Omega fatty acids.

28380

0 60 1209030 150Minutes

pA

0

50

Phospholipids

Degradants

Column: Acclaim C30, 5 µm (4.6 × 150 mm)Mobile Phase: A. Acetonitrile (CH3CN) B. Iso-propanol (IPA) C. Ammonium acetate buffer (0.1 M, pH 5.0)Gradient: Time (min) CH3CN IPA Buffer –15 60 20 20 0 60 20 20 0.1 60 20 20 150 38 50 12 151 5 90 5 160 5 90 5Temperature: 40 °CFlow Rate: 1.0 mL/minInj. Volume: 5 µLDetection: Corona ultra (gain = 100 pA; filter = medium; neb. temp. = 25 °C)Sample: Egg lecithin – PL100M (5 mg/mL in iso-propanol)

28381

10 12 14 16 18 20 22 240

100

pA

Minutes

A

B

1 2

3 4

5 6 7

8 9

10

11

12

13

14 0

6000

ng

0 9Area

LLA

Column: Acclaim C30, 5 µm (4.6 × 150 mm)LC System: UltiMate 3000 RS, Dual GradientMobile Phases: A. Water:formic acid:mobile phase B 900:3.6:100 (v/v) B. Acetone:acetonitrile:THF:formic acid 675:225:100:4(v/v)Gradient: Time (min) %A %B 0 100 0 1 40 60 13 30 70 22 5 95 24 5 95 29 100 0 32 100 0Flow Rate: 1.00 mL/minTemperature: 30 °CInj. Volume: 2 µLDetection: Corona ultra, nebulizer 15 °C, filter highSamples: A. Standards in isopropanol B. Saponified chicken fat

Peaks: 1. SDA 2. EPA 3. ALA 4. GLA 5. DHA 6. Arach. 7. LLA

8. DPA 9. 9E, 14Z-CLA 10. ETA 11. Adrenic 12. Oleic 13. EDA 14. Erucic

ORDERING INFORMATION

IntheU.S.,call(800)346-6390orcontacttheDionexRegionalOfficenearestyou.OutsidetheU.S.,orderthroughyourlocalDionexofficeordistributor.Referthefollowingpartnumbers:

Acclaim C30 Analytical and Guard Columns Part Number

AcclaimC30,5µmAnalytical,4.6×250mm.......................................................... 075718

AcclaimC30,5µmAnalytical,4.6×150mm.......................................................... 075719

AcclaimC30,3µmAnalytical,4.6×150mm.......................................................... 075723

AcclaimC30,3µmAnalytical,3.0×250mm.......................................................... 075726

AcclaimC30,3µmAnalytical,3.0×150mm.......................................................... 075724

AcclaimC30,3µmAnalytical,2.1×150mm.......................................................... 075725

AcclaimC30,5µm,Guard,4.6×10mm................................................................. 075720

AcclaimC30,5µm,Guard,3.0×10mm................................................................. 075721

AcclaimC30,5µm,Guard,2.1×10mm................................................................. 075722

Accessories Part Number

GuardHolder(V-2).................................................................................................... 069580

GuardHolderkitV-2(HolderV-2andCoupler)....................................................... 069707

GuardCoupler............................................................................................................ 074188

SPECIFICATIONS

Column Chemistry: C

30alkyl

Silica Substrate: Spherical,high-purity Particlesizes: 3and5µm Surfacearea: 200m2/g Poresize: 200Å

Operating pH range: pH2–8

Operating Temperature: Upto60°C

Aqueous compatibility: 0to100%

Solvent compatibility: 0to100%

Operating pressure and flow rate: Seetabletopright

Corona,Acclaim,CAD,andUltiMateareregisteredtrademarks,andultraisatrademarkofDionexCorporation.

North America

U.S./Canada (847) 295-7500 South America

Brazil (55) 11 3731 5140

Europe

Austria (43) 1 616 51 25 Benelux (31) 20 683 9768 (32) 3 353 4294 Denmark (45) 36 36 90 90 France (33) 1 39 30 01 10 Germany (49) 6126 991 0 Ireland (353) 1 644 0064 Italy (39) 02 51 62 1267 Sweden (46) 8 473 3380 Switzerland (41) 62 205 9966 United Kingdom (44) 1276 691722

Asia Pacific

Australia (61) 2 9420 5233 China (852) 2428 3282 India (91) 22 2764 2735 Japan (81) 6 6885 1213 Korea (82) 2 2653 2580 Singapore (65) 6289 1190Taiwan (886) 2 8751 6655

Dionex Corporation

1228 Titan Way P.O. Box 3603 Sunnyvale, CA 94088-3603 (408) 737-0700 www.dionex.com

LPN 2776 PDF 3/11©2011 Dionex Corporation

Column Dimensions

Particle Size

Part Number

Maximum Pressure

(psi)

Typical Flow Rate (Recommended)

(mL/min)

Maximum Flow Rate (mL/min)

4.6×250mm 5µm 075718 9,000 0.8–1.5 2.0

4.6×150mm 5µm 075719 8,000 0.8–1.5 2.0

4.6×150mm 3µm 075723 8,500 0.8–1.5 1.0

3.0×250mm 3µm 075726 12,000 0.4–0.6 1.0

3.0×150mm 3µm 075724 8,500 0.4–0.6 1.0

2.1×150mm 3µm 075722 8,800 0.2–0.3 0.5

OPERATING PRESSuRE AND FlOw RATE SPECIFICATIONS