csh technology: acquity csh columns applications notebook€¦ · list of compounds analyzed using...

CSH TeCHnology: ACQUITY CSH Columns Applications Notebook

List of compounds anaLyzed using csh coLumns

Lit Code Title Page

WA64123 Acetaminophen Impurities 2

WA64122 Acids, Bases, and Neutrals 3

WA64133 Aromatic Acids 4

WA64124 Basic Drugs 5

WA64125 Beta Blockers 6

WA64134 Buproprion Hydrochloride (Wellbutrin) in Extended Release Tablets 7

WA64135 Clozapine Impurities 8

WA64136 Food Additives 9

WA64126 Formic and Acetic Acid 10

WA64138 Mirtazapine Impurities 11

WA64127 Nonsteroidal Anti-Inflammatory Drugs (NSAIDS) 12

WA64139 Organic Acids 13

WA64128 Paroxetine Impurities 14

WA64129 Paroxetine Isomers 15

WA64130 Pharmaceutical Test Mix 16

WA64137 Statin Drugs 17

WA64131 Sulfa Drugs 18

WA64132 Tricyclic Antidepressants 19

WA64140 Veterinary Drugs 20

2

ACQU

ITY

CSH

WA6

4123

©2010 Waters Corporation. Waters, The Science of What’s Possible, ACQUITY CSH and ACQUITY UPLC are trademarks of Waters Corporation.

LC CONDITIONS

Column: ACQUITY CSH™ Fluoro-Phenyl, 2.1 x 50 mm, 1.7 μm

Part Number: 186005351

Mobile Phase A: 0.1% acetic acid in water

Mobile Phase B: acetonitrile

Gradien:

Injection Volume: 2 μL

Sample Diluent: water

Column Temperature: 40 °C

Weak Needle Wash: 95/5 water/acetonitrile

Strong Needle Wash: 95/5 acetonitrile/water

Detection: UV @ 270 nm

Sampling Rate: 20 points/sec

Filter Time Constant: 0.1 sec

System: ACQUITY UPLC® System with

ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.6 98 2 Initial2.00 0.6 50 50 62.50 0.6 50 50 62.51 0.6 98 2 63.50 0.6 98 2 6

STRUCTURES

-0.005

0.000

0.005

0.010

0.015

0.020

0.025

0.030

0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.0 min

1

2

3 4

Compounds

1. 4-Aminophenol (0.002 mg/mL)

2. Acetaminophen (0.2 mg/mL)

3. 4-Nitrophenol (0.002 mg/mL)

4. 4-Chloroacetanilide (0.002 mg/mL)

HO

NH

O

OH

NO2Cl

HN

O

4-Aminophenol Acetaminophen

4-Nitrophenol 4-Chloroacetanilide

OH

NH2

anaLysis of acetaminophen impurities using acQuit y csh coLumns

3


ACQU

ITY

CSH

WA6

4122

LC CONDITIONS

Columns: ACQUITY CSH™ C18, 2.1 x 50 mm, 1.7 μm


ACQUITY CSH Phenyl-Hexyl, 2.1 x 50 mm, 1.7 μm


ACQUITY CSH Fluoro-Phenyl, 2.1 x 50 mm, 1.7 μm


Mobile Phase A: 0.1% formic acid in water (pH ~ 2.7)


Gradient Table:

Injection Volume: 2 μL (PLNO)

Sample Diluent: 25/75 methanol/water







System: ACQUITY UPLC® System with column manager

and ACQUITY UPLC PDA detector

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.5 95 5 Initial5.00 0.5 5 95 65.01 0.5 95 5 65.50 0.5 95 5 6

STRUCTURES

ACQUITY CSH C18

AU

0.00

0.15

0.30

0.45

AU

0.00

0.15

0.30

0.45 ACQUITY CSH Phenyl-Hexyl

1

5

6 7

2

3

1

2

3

4

4

5

6 7

AU

0.00

0.15

0.30

0.45

0.00 1.20 2.40 3.60 4.80 min

ACQUITY CSH Fluoro-Phenyl

1

2

3

4

5

6 7

Compounds

1. Imipramine (100 µg/mL)

2. Amitriptyline (100 µg/mL)

3. 1-Pyrenesulfonic acid (50 µg/mL)

4. Flavone (30 µg/mL)

5. Fenoprofen (100 µg/mL)

6. Diclofenac (100 µg/mL)

7. Octanophenone (100 µg/mL)

N

N N

SO3H

O

O

OCOOH

COOHHN

Cl

Cl

O

Imipramine Amitriptyline

1-Pyrenesulfonic acid

Flavone

Fenoprofen

DiclofenacOctanophenone

anaLysis of acids, Bases, and neutraLs using acQuit y csh coLumns

4

ACQU

ITY

CSH

WA6

4133


LC CONDITIONS

Column: ACQUITY CSH™ C18, 2.1 x 50 mm, 1.7 μm




Gradient:


Sample Concentration: 10 µg/mL each









ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.6 98 2 Initial5.00 0.6 40 60 65.50 0.6 40 60 65.51 0.6 98 2 66.00 0.6 98 2 6

STRUCTURES

0.00

0.04

0.08

0.12

0.16

0.20

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 min

AU

1

2

3

4

5

67

COmPOUNDS

1. Protocatechuic acid

2. 3,4-Dihydroxyphenylacetic acid (DOPAC)

3. 4-Hydroxybenzoic acid

4. Vanillic acid

5. Syringic acid

6. Salicylic acid

7. 2,4-Dichlorophenoxyacetic acid (Weed B Gon)

Protocatechuic acid

COOH

OHOCH3

OCH3

HO

H3CO COOH COOHOH

4-Hydroxybenzoic acid

Vanillic acid

Syringic acid

Salicylic acid

2,4-Dichlorophenoxyacetic acid (Weed B Gon)

3,4-Dihydroxyphenylacetic acid (DOPAC)

Cl Cl

O COOH

COOH

OH

COOH

OHOH

HO

COOH

OH

anaLysis of aromatic acids using acQuit y csh coLumns

5


ACQU

ITY

CSH

WA6

4124

LC CONDITIONS

Column: ACQUITY CSH™ Phenyl-Hexyl, 2.1 x 50 mm, 1.7 μm


Mobile Phase A: 10 mM ammonium formate, pH 3


Gradient:









System: ACQUITY UPLC® System with ACQUITY

UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.4 95 5 Initial3.00 0.4 45 55 63.50 0.4 45 55 63.51 0.4 95 5 64.50 0.4 95 5 6

STRUCTURES

0.00

0.04

0.08

0.12

0.16

0.20

0.0 0.5 1.0 1.5 2.0 2.5 3.0 min

AU

1

2

3

4

5

6

7

8

9

Compounds

1. 2-aminopyrazine (25 μg/mL)

2. Pindolol (10 μg/mL)

3. Quinine (10 μg/mL)

4. Quinine impurity

5. Labetalol (60 μg/mL)

6. Diltiazem impurity

7. Diltiazem (25 μg/mL)

8. Verapamil (50 μg/mL)

9. Amitriptyline (25 μg/mL)

N

S

NO

O

O

OCH3

HO

HN

H2N

O OH

OHN

OH

HN

N

HO

H3CO

N

H

H

H3CO

H3CO

CNN OCH3

OCH3

N

Pindolol2-aminopyrazine

Quinine Diltiazem

Labetalol Amitriptyline

Verapamil

N

N NH2

anaLysis of Basic drugs using acQuit y csh coLumns

6

ACQU

ITY

CSH

WA6

4125


LC CONDITIONS



Mobile Phase A: 10 mM ammonium formate, pH 3


Gradient:











ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.4 95 5 Initial3.00 0.4 15 85 63.50 0.4 15 85 63.51 0.4 95 5 64.50 0.4 95 5 6

STRUCTURES

0.00

0.02

0.04

0.06

0.08

0.10

0.0 0.5 1.0 1.5 2.0 2.5 3.0 min

AU

1

2

3

4

5

6 Compounds

1. Atenolol

2. Pindolol

3. Metoprolol

4. Oxprenolol

5. Labetalol

6. Propranolol

Atenolol

NH2

OON

H OH

HO

HN

H2N

O OH

OHN

OH

H3CO

OHN

O

OH

OHN

OH

HN

OOH

NH

Labetalol

Metoprolol

Oxprenolol

Pindolol

Propranolol

anaLysis of Beta BLockers using acQuit y csh coLumns

7


ACQU

ITY

CSH

WA6

4134

LC CONDITIONS



Mobile Phase A: water


Mobile Phase C: 2% formic acid in water

Gradient:


Sample Diluent: H2O

Sample Concentration: 20 µg/mL


Wash Solvent: 50/50 water/acetonitrile

Purge Solvent: 50/50 water/acetonitrile

System: ACQUITY UPLC® H-Class with

ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.9 90 5 5 Initial3.00 0.9 0 95 5 63.50 0.9 0 95 5 63.51 0.9 90 5 5 65.00 0.9 90 5 5 6

SamPLE PREPaRaTION PROCEDURE

Ten extended release tablets (each containing 100 mg bupropion

hydrochloride) were homogenized in 50 mL of 50/50 methanol/

water to give 20 mg/mL of bupropion hydrochloride. The sample was

then centrifuged at 4000 rpm for 10 minutes. The supernatant was

then diluted to 0.02 mg/mL in water and filtered through a 0.45 µm

PVDF syringe filter prior to injection.

STRUCTURES

AU

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.35

0.40

0.0 0.5 1.0 1.5 2.0 2.5 3.0 min

Peak width (4.4%) = 1.3 secondsUSP tailing factor = 1.27

1 COmPOUND

1. Bupropion (pKa = 7.9)

ClHN

O

Bupropion

anaLysis of Bupropion hydrochLoride (WeLLButrin) in extended reLease taBLetsusing acQuit y csh coLumns

8

ACQU

ITY

CSH

W

A641

35


LC CONDITIONS






Gradient:



Sample Concentration: 400 µg/mL (impurities present at ~0.2%)




System: ACQUITY UPLC® H-Class with

ACQUITY UPLC PDA and SQD

mS CONDITIONS

Capillary: 3.0 kV (ES+)

Cone: 20 V

Extractor: 3 V

Desolvation Temperature: 350 °C

Source Temperature: 150 °C

Desolvation Gas: 800 L/hr

Scan Range: 200 to 650 m/z

Scan Rate: 1000 Da/sec

STRUCTURES

AU

-0.005

0.000

0.005

0.010

0.015

0 1 2 3 4 5 min

UV 254 nm

1

2

3 4 5

Inte

nsity

Inte

nsity

Inte

nsity

Inte

nsity

0 1 2 3 4 5 min

Extracted Ion Chromatogramat m/z 313.1

Extracted Ion Chromatogramat m/z 345.1

Extracted IonChromatogramat m/z 539.2

Extracted IonChromatogramat m/z 245.0

0.0

5.0 x 105

1.0 x 106

0

1 x 106

0

2.0 x 105

4.0 x 105

0

2.0 x 105

4.0 x 105

5.0 x 105

COmPOUNDS

1. 8-Chloro-11-(piperazin-1-yl)-5H-dibenzo[b,e][1,4]-diazepine

2. Clozapine

3. 11-[2-[(2-Amino-4-chlorophenyl)amino]benzoyl]- 4-methylpiperazine

4. 11,11'-(Piperazine-1,4-diyl)bis(8-chloro-5H-dibenzo[b,e] [1,4]diazepine)

5. 8-Chloro-5,10-dihydro-11H-dibenzo[b,e][1,4]diazepin-11-one

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 90 5 5 Initial5.0 0.6 45 50 5 66.0 0.6 0 95 5 66.1 0.6 90 5 5 68.0 0.6 90 5 5 6

N

NH

N

NH

Cl

8-Chloro-11-(piperazin-1-yl)-5H-dibenzo[b,e][1,4]-diazepine

N

NH

N

N

Cl

Clozapine

HN

Cl NH2

NON

1-[2-[(2-Amino-4-chlorophenyl)amino]benzoyl]-4-methylpiperazine

N

N

N

NH

Cl

N

HN

Cl

11,11'-(Piperazine-1,4-diyl)bis(8-chloro-5H-dibenzo

[b,e][1,4]diazepine)

HN

NH

O

Cl

8-Chloro-5,10-dihydro-11H-dibenzo[b,e][1,4]

diazepin-11-one

anaL aysis of cLozapine impurities using acQuit y csh coLumns

9


ACQU

ITY

CSH

WA6

4136

LC CONDITIONS

Column: ACQUITY CSH Fluoro-Phenyl, 2.1 x 50 mm, 1.7 μm


Mobile Phase A: 0.1% phosphoric acid in water


Gradient:










ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.6 98 2 65.00 0.6 2 98 65.50 0.6 2 98 65.51 0.6 98 2 66.00 0.6 98 2 6

STRUCTURES

0.00

0.10

0.20

0.30

0.40

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 min

1

2

3 4

5

COmPOUNDS

1. p-Hydroxybenzoic acid (10 µg/mL)

2. Sorbic acid (10 µg/mL)

3. Dehydroacetic acid (100 µg/mL)

4. Methyl paraben (25 µg/mL)

5. Saccharin (100 µg/mL)

HO

OH

O

COOH

HO

O

O

SNH

O

OO

p-Hydroxybenzoic acid

Sorbic acid

Methyl paraben

Saccharin

O

O O

ODehydroacetic acid

anaLysis of food additives using acQuit y csh coLumns

10

ACQU

ITY

CSH

WA6

4126


LC CONDITIONS



Isocratic Mobile Phase: 0.1% phosphoric acid in water

Flow Rate: 0.6 mL/min


Sample Concentration: 0.02%









ACQUITY UPLC PDA

STRUCTURES

0.00

0.01

0.02

0.03

0.04

0.05

0.06

0.07

0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 min

AU

1

2

COmPOUNDS

1. Formic acid

2. Acetic acid

Formic acid

OH

O

Acetic acid

O

OHH

anaLysis of formic and acetic acids using acQuit y csh coLumns

11


ACQU

ITY

CSH

WA6

4138

AU

0.000

0.008

0.016

0.0 1.5 3.0 4.5 min

1

2

3

4

5

6

7

8

COmPOUNDS

1. Mirtazapine Impurity A (0.15%)

2. Mirtazapine Impurity B (0.05%)

3. Mirtazapine Impurity C or F (0.09%)

4. Mirtazapine Impurity D (0.55%)

5. Mirtazapine (98.81%)

6. Unknown impurity (0.08%)

7. Mirtazapine Impurity E (0.13%)

8. Mirtazapine Impurity C or F (0.15%)

LC CONDITIONS



Mobile Phase A: 0.1% ammonium hydroxide in water


Gradient:


Sample Concentration: 1 mg/mL of mirtazapine USP resolution standard

Sample Diluent: 50/50 acetonitrile/water

Column Temperature 45 °C







ACQUITY UPLC PDA

STRUCTURES

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.7 95 5 Initial6.5 0.7 22 78 67.5 0.7 22 78 67.6 0.7 95 5 68.0 0.7 95 5 6

NN

N

OH

NN

N

NN

N O

Mirtazapine Impurity A

Mirtazapine Impurity B

NN

NH

Mirtazapine Impurity D Mirtazapine Impurity E

NN

NO

Mirtazapine Impurity C

NN

N

Mirtazapine

NN

N

O

Mirtazapine Impurity F

anaLysis of mirtazapine impurities using acQuit y csh coLumns

12

ACQU

ITY

CSH

WA6

4127


LC CONDITIONS






Gradient:









System: ACQUITY UPLC® H-Class System with

ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 70 25 5 Initial5.0 0.6 15 80 5 66.0 0.6 0 95 5 66.1 0.6 70 25 5 610.0 0.6 70 25 5 6

STRUCTURESAU

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

0 1 2 3 4 5 min

1

2

3

4

5

6

COmPOUNDS

1. Tolmetin (10 µg/mL)

2. Naproxen (10 µg/mL)


4. Ibuprofen (100 µg/mL)

5. Indomethacin (10 µg/mL)


NO

COOH

Tolmetin

OCOOH

OH

O

H3CO

COOHCOOH

HN

Cl

Cl

N

O Cl

COOHH3CO

Fenoprofen Ibuprofen

Naproxen Diclofenac

Indomethacin

anaLysis of nonsteroidaL anti-infL ammatory drugs (nsaids) using acQuit y csh coLumns

13


ACQU

ITY

CSH

WA6

4139

COmPOUNDS

1. Tolmetin (10 µg/mL)

2. Naproxen (10 µg/mL)


4. Ibuprofen (100 µg/mL)

5. Indomethacin (10 µg/mL)


LC CONDITIONS



Mobile Phase: 0.1% phosphoric acid in water

Isocratic Flow Rate: 0.2 mL/min

Injection Volume: 1 to 3 μL

Sample Concentration: 125 µg/mL (250 µg/mL for lactic acid)

(250 µg/mL for lactic acid)









ACQUITY UPLC PDA

Sample Preparation Procedure: Samples were filtered through a 0.45 μm

PTFE syringe filter and injected directly.

STRUCTURES

0.00

0.02

0.04

0.06

0.00

0.02

0.04

0.06

0.08

0.10

-0.02

0.00

0.02

0.04

0.06

0.08

10 2 3 4 5 6 7 8 9 10 min

1

2 3 4

5

6

6

6

3

Standard mix

Lemon-lime sports drink

Vitamin-fortified water (grape flavor)

COmPOUNDS

1. DL-Tartaric acid

2. L-Malic acid

3. Lactic Acid

4. Acetic Acid

5. Succinic Acid

6. Citric Acid

HOOH

O

OH

OH

O

HOOH

O

O

DL-Tartaric acid

OH

HO

OOH

O

L-Malic acid

OH

O

Acetic Acid

Succinic Acid

OHOH

O

Lactic Acid

HO COOH

COOH

COOH

Citric Acid

anaLysis of organic acids using acQuit y csh coLumns

14

ACQU

ITY

CSH

WA6

4128


LC CONDITIONS




Mobile Phase B: methanol

Mobile Phase C: 2% acetic acid in water

Gradient:

Pre-Injector volume: 200 µL



Sample Concentration: paroxetine at 0.2 mg/mL; impurities at

0.002 mg/mL each



Purge solvent: 50/50 water/acetonitrile





ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 85 10 5 Initial5.0 0.6 75 20 5 66.0 0.6 75 20 5 66.1 0.6 85 10 5 612.0 0.6 85 10 5 6

STRUCTURES

AU

-0.003

-0.002

-0.001

0.000

0.001

0.002

0.003

0.004

0 1 2 3 4 5 6 min

1

2

3

4

COmPOUNDS

1. trans-4-phenyl-3-[(3,4-methylenedioxy) phenoxymethyl]-piperidine hydrochloride

2. cis-Paroxetine

3. Paroxetine hydrochloride

4. (3S-trans)-1-Methyl-3-[(1,3-benzodioxol-5- yloxy)methyl]-4-(4-fluorophenyl)piperidine

HN

O

O

O

trans-4-Phenyl-3-[[3,4-(methylenedioxy)phenoxy]methyl]piperidine

HN

O

O

O

F cis-Paroxetine

HN

F

O

O

O

Paroxetine

N

O

O

O

F(3S-trans)-1-Methyl-3-[(1,3-benzodioxol-5-yloxy)

methyl]-4-(4-fluorophenyl)piperidine

anaLysis of paroxetine impurities using acQuit y csh coLumns

ACQU

ITY

CHS

Colu

mns

15


ACQU

ITY

CSH

WA6

4129

COmPOUNDS

1. trans-4-phenyl-3-[(3,4-methylenedioxy) phenoxymethyl]-piperidine hydrochloride

2. cis-Paroxetine

3. Paroxetine hydrochloride

4. (3S-trans)-1-Methyl-3-[(1,3-benzodioxol-5- yloxy)methyl]-4-(4-fluorophenyl)piperidine

LC CONDITIONS




Mobile Phase B: methanol

Mobile Phase C: 2% acetic acid in water

Gradient:

Pre-Injector volume: 200 µL








SQD STRUCTURES

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 85 10 5 Initial5.0 0.6 75 20 5 66.0 0.6 75 20 5 66.1 0.6 85 10 5 612.0 0.6 85 10 5 6

mS CONDITIONS


Cone: 30 V

Extractor: 3 V






STRUCTURES

Inte

nsity

0

1x106

2x106

3x106

4x106

5x106

6x106

7x106

8x106

1 2 3 4 5 6 7 8 min 0

1

2

Extracted ion chromatogram at m/z 330.2

COmPOUNDS

1. cis-Paroxetine

2. (+)-trans-Paroxetine

(+)-trans-Paroxetinecis-Paroxetine

HN

O

O

O

F

HN

F

O

O

O

anaLysis of paroxetine isomers using acQuit y csh coLumns

16

ACQU

ITY

CSH

WA6

4130


LC CONDITIONS

Columns: ACQUITY CSH™ C18, 2.1 x 50 mm, 1.7 μm


ACQUITY CSH Phenyl-Hexyl, 2.1 x 50 mm, 1.7 μm


ACQUITY CSH Fluoro-Phenyl, 2.1 x 50 mm, 1.7 μm





Gradient:










ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 90 5 5 Initial5.0 0.6 0 95 5 66.0 0.6 0 95 5 66.1 0.6 90 5 5 610.0 0.6 90 5 5 6

STRUCTURES

AU

0.00

0.05

0.10

0.15

AU

0.00

0.05

0.10

0.15

AU

0.00

0.05

0.10

0.15

0 1 2 3 4 5 6 min

18

ACQUITY CSH C18

ACQUITY CSH Phenyl-Hexyl

ACQUITY CSH Fluoro-Phenyl

1

2 3

4

5

6

7 8

9

10

1 2 3

4

5

6

7 8

9

10

1 2

3

4

5

6

7 8

9

10

COmPOUNDS

1. 8-Bromoguanosine (6.25 µg/mL)

2. Acetanilide (3.75 µg/mL)

3. Triamcinolone (6.25 µg/mL)

4. Hydrocortisone (2.5 µg/mL)

5. 2-Amino-7-chloro-5-oxo-5H-[1]benzopyrano[ 2,3-b]pyridine-3-carbonitrile (2.5 µg/mL)

6. 3-Aminofluoranthene (6.25 µg/mL)

7. 6α-Methyl-17α-hydroxyprogesterone (5 µg/mL)

8. 2-Bromofluorene (3 µg/mL)

9. Perylene (2.5 µg/mL)

10. Naphtho[2,3-a]pyrene (2.5 µg/mL)

8-Bromoguanosine

N

HN

N

N

O

H2N

O

HO

OHOH

Br

Acetanilide

NH

O

Triamcinolone

O

O

HOH

HF

OHOH

OH

Hydrocortisone

O

O

HOH

HH

OH

OH

2-Amino-7-chloro-5-oxo-5H-[1]benzopyrano[2,3-b]

pyridine-3-carbonitrile

NO

NO

Cl

NH2

3-Aminofluoranthene

NH2

6α-Methyl-17α-hydroxyprogesterone

OH

H

H

O

OH

2-Bromofluorene

Br

Perylene

anaLysis of pharmaceuticaL test mix on acQuit y csh coLumns

Naphtho[2,3-a]pyrene

ACQU

ITY

CHS

Colu

mns

17


ACQU

ITY

CSH

WA6

4137

LC CONDITIONS



Mobile Phase A: 0.1% formic acid in water


Gradient:











ACQUITY UPLC PDA

STRUCTURES

0.00

0.04

0.08

0.12

0.16

0 1 2 3 4 5 min

AU

1

2

3

4

COmPOUNDS

1. Pravastatin sodium (Pravachol)

2. Atorvastatin (Lipitor)

3. Lovastatin (Mevacor)

4. Simvastatin (Zocor)

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.6 95 5 Initial5.00 0.6 30 70 65.50 0.6 5 95 65.51 0.6 95 5 66.00 0.6 95 5 6

HO

HO

O

–OOCHO

OH

H

Na+

NCOOH

OH OH

NH

O

F

HO

OH

H

O

OHO

Pravastatin sodium

Atorvastatin

Simvastatin

HO

OH

H

O

OHO

Lovastatin

anaLysis of statin drugs using acQut y csh coLumns

18

ACQU

ITY

CSH

WA6

4131


LC CONDITIONS





Gradient:











ACQUITY UPLC PDA

Time (min)

Flow(mL/min)

%A %B Curve

Initial 0.5 95 5 Initial5.00 0.5 40 60 65.50 0.5 40 60 65.51 0.5 95 5 66.00 0.5 95 5 6

STRUCTURES

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 min

AU

1

2

3

4

5

6

7

8

COmPOUNDS

1. Sulfanilamide

2. Sulfathiazole

3. Sulfamerazine

4. Sulfamethazine

5. Sulfamethizole

6. Succinylsulfathiazole

7. Sulfamethoxazole

8. Sulfasoxazole

Sulfanilamide

H2N

SNH

O O

N

N

H2N

SNH

O O

N

N

H2N

SNH

O O

NN

S

NH

SNH

O O

N

S

HOOC

O

H2N

SNH

O O ON

Sulfamerazine Sulfamethazine

Sulfamethizole

Succinylsulfathiazole

Sulfamethoxazole

H2N

SNH2

O O

Sulfathiazole

H2N

SNH

O O

N

S

SNH

NOO O

H2NSulfasoxazole

anaLysis of suLfa drugs using acQuit y csh coLumns

ACQU

ITY

CHS

Colu

mns

19


COmPOUNDS

1. Sulfanilamide

2. Sulfathiazole

3. Sulfamerazine

4. Sulfamethazine

5. Sulfamethizole

6. Succinylsulfathiazole

7. Sulfamethoxazole

8. Sulfasoxazole

ACQU

ITY

CSH

WA6

4132

LC CONDITIONS






Gradient:











ACQUITY UPLC PDA and SQD

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.6 70 25 5 Initial2.0 0.6 60 35 5 63.0 0.6 0 95 5 63.1 0.6 70 25 5 65.0 0.6 70 25 5 6

mS CONDITIONS


Cone: 30 V

Extractor: 3 V






STRUCTURES

Inte

nsity

0.5 1.0 1.5 2.0 min 0.0

AU

1.0x107

2.0x107

3.0x107

4.0x107

0.000

0.004

0.008

0.012

UV

MS

1

2

3 4

5 6

0.5 1.0 1.5 2.0 min 0.0

COmPOUNDS

1. Doxepin

2. Desipramine

3. Imipramine

4. Nortriptyline

5. Amitriptyline

6. Trimipramine

O

N

Doxepin

NH

N

N NH

N

N

N

Desipramine

Imipramine

Nortriptyline

Amitriptyline

Trimipramine

anaLysis of tricycLic antidepressants using acQuit y csh coLumns

20


anaLysis of veterinary drugs using acQuit y csh coLumns

LC CONDITIONSColumns: ACQUITY CSH™ C18, 2.1 x 100 mm, 1.7 μm Part Number: 186005297Mobile Phase A: waterMobile Phase B: acetonitrile Mobile Phase C: 2.0% formic acid in water (pH ~ 2.7)Gradient:

Injection Volume: 5 μL Sample Diluent: 10/90 methanol/waterColumn Temperature: 30 °CWash: 1% formic acid in waterSystem: ACQUITY UPLC® H-Class with TQD (MassLynx™ v. 4.1)

TQD CONDITIONSCapillary: 3.00 kVExtractor: 3.00 VRF: 0.10 VSource Temperature: 150 °CDesolvation Temperature: 350 °CCone Gas Flow: 30 L/HrDesolvation Gas Flow: 800 L/Hr

Collision Gas Flow: 0.10 mL/min

1

7

8

3

6

4

5 9

2

100

%

00 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 min

COmPOUNDS

1. Ciprofloxacin (4 µg/mL)

2. Enrofloxacin (2 µg/mL)

3. Erythromycin (2 µg/mL)

4. Oxacillin (4 µg/mL)

5. Oxytetracycline (4 µg/mL)

6. Penicillin G (2 µg/mL)

7. Sulfamerazine (2 µg/mL)

8. Sulfamethazine (0.5 µg/mL)

9. Tetracycline (4 µg/mL)

Time (min)

Flow(mL/min)

%A %B %C Curve

Initial 0.3 80 15 5 Initial2.5 0.3 55 40 5 63.9 0.3 0 95 5 64.9 0.3 0 95 5 65.0 0.4 80 15 5 66.0 0.4 80 15 5 66.1 0.3 80 15 5 67.0 0.3 80 15 5 6

Compound MRM transition (m/z) Cone (V) CID (eV)

Ciprofloxacin332.00>288.21332.00>314.32

20 20

2525

Enrofloxacin360.10>316.38360.10>342.39

15 15

2525

Erythromycin734.68>158.18734.68>576.49

30 30

20 15

Oxacillin402.40>160.25402.40>243.24

25 25

15 15

Oxytetracycline461.10>426.43461.10>443.67

25 25

35 30

Penicillin-G335.17>160.16335.17>176.03

30 30

30 25

Sulfamerazine265.18>92.10

265.18>156.2225 25

20 30

Sulfamethazine279.00>92.21

279.00>124.3130 30

30 25

Tetracycline445.20>410.26445.20>427.80

30 30

35 30

N

F

HNN

O

OH

O

Ciprofloxacin

N

F

NN

O

OH

O

Enrofloxacin

HH

OH O OH O

NH2

OOH

NHOOH

OH

Oxytetracycline

NH N

S

COOH

HH

O

O

Penicillin G

NO

N

SHN

O

OCOOH

HH

Oxacillin

H2N

SNH

O O

N

N

Sulfamerazine

H2N

SNH

O O

N

N

Sulfamethazine

HH

OH O OH O

NH2

OOH

NHOOH

Tetracycline

O

O

OHHO OH

O

O

O

O

NHO

OCH3

OH

Erythromycin

ACQU

ITY

CSH

WA6

4140

21

ACQU

ITY

CSH

NOTE

S

22

ACQU

ITY

CSH

NOTE

S

23

ACQU

ITY

CSH

NOTE

S

Austria and European Export (Central South Eastern Europe, CIS and Middle East) 43 1 877 18 07

Australia 61 2 9933 1777

Belgium 32 2 726 1000

Brazil 55 11 5094-3788

Canada 1 800 252 4752 x2205

China 86 21 6879 5888

CIS/Russia +497 727 4490/290 9737

Czech Republic 420 2 617 1 1384

Denmark 45 46 59 8080

Finland 09 5659 6288

France 33 1 30 48 72 00

Germany 49 6196 400600

Hong Kong 852 2964 1800

Hungary 36 1 350 5086

India and India Subcontinent 91 80 2837 1900

Ireland 353 1 448 1500

Italy 39 02 265 0983

Japan 81 3 3471 7191

Korea 82 2 6300 4800

Mexico 52 55 5524 7636

The Netherlands 31 76 508 7200

Norway 47 6 384 60 50

Poland 48 22 6393000

Puerto Rico 1 787 747 8445

Singapore 65 6593 7100

Spain 34 936 009 300

Sweden 46 8 555 11 500

Switzerland 41 56 676 70 00

Taiwan 886 2 2543 1898

United Kingdom 44 208 238 6100

All other countries: Waters Corporation U.S.A. 1 508 478 2000

1 800 252 4752

www.waters.com

Sales Offices

©2010 Waters Corporation. Waters, CSH, ACQUITY UPLC,

ACQUITY CSH, MassLynx and The Science of What’s Possible

are trademarks of Waters Corporation.

June 2010 720003537EN KK-PDF

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