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Novel immobilised polysaccharide-derived CSPs

to enhance success rate in the resolution of enantiomers

P. Franco, D. Nguyen, T. Zhang

Screening strategy for the resolution of enantiomers

Higher success rates can be achieved by: • Screening only LC or SFC with 7 (or more) columns • Screening LC and SFC with a limited number of columns

What is the best choice?

LC SFC

Can we achieve this with a minimum number of columns and efficient screening strategies?

• Resolution of the enantiomeric pair (or isomeric mixture)

• Resolution from impurities

• Short analysis times

• Suitable elution order

• Compatibility of the sample media with mobile phase and column

• Low LOD/LOQ

• Reproducibility and robustness of the method

• Stability of the sample under the analytical conditions

Some of the requirements / challenges

• Samples are screened systematically in LC and SFC with a FAST screening

• Systematic screening

• LC on IA, IB and IC (H/IPA, H/EtOH, H/THF, H/DCM, H/AcOEt, ACN and alcohol)

• SFC on IA, IB and IC (MeOH, IPA and EtOH)

How were we handling in the last years?

RO

O

OR

ROO n RO

O

OR

RO

O

n

Amylose-based

carbamates Cellulose-based

carbamates

CHIRALPAK IA CHIRALPAK IB CHIRALPAK IC NH

O CH3

CH3

NH

O CH3

CH3

NH

O Cl

Cl

• Free choice of solvents in the mobile phase to match solubility

• Greater options for chemical stability

• New selectivities (and elution orders)

• Favouring elution kinetics with certain solvents

• Choice in the sample solvent dictated by sample solubility

• Column and method robustness

In which aspects solvent versatility was helping?

T. Zhang et al., J. Chromatogr. A 1191 (2008) 214

New selector – June 2011 CHIRALPAK ID

O

O

O

O O

n

NH

Cl

OONH

NH

O Cl

Cl

CHIRALPAK ID

Based on

amylose tris-(3-chlorophenylcarbamate)

A new selector for the immobilised series!!

CHIRALPAK IA CHIRALPAK IB

N

CH3

H

O CH3

Different substituents New enantiorecognition

N

Cl

H

O Cl

CHIRALPAK IC N

H

O Cl

CHIRALPAK ID

Looking for complementary recognition patterns

HPLC method development

CHIRALPAK ID Complementary properties

Hexane/2-PrOH/DEA 80/20/0.1 (250 x 4.6 mm, 5 µm); 1.0 mL/min; 25 C

min 0 1 2 3 4 5 6 7 min 0 1 2 3 4 5 6 7

min 0 1 2 3 4 5 6 7

CHIRALPAK IA CHIRALPAK IB

CHIRALPAK IC

min 0 1 2 3 4 5 6 7 8 9

CHIRALPAK ID

Isradipine

O

O

O

O

NH

N

O

N

Ex

am

ple

s in

H

PLC

Column dimensions: 250 x 4.6 mm, 5 µm; 1.0 mL/min; 25 C

Mobile phase selection in HPLC Analytical methods

CHIRALPAK IA

min 0 1 2 3 4 5 6 7

ACN/IPA/TFA 95/5

N

O

OH

O

CHIRALPAK IB

Hex/IPA/TFA 80/20/0.1

min 0 1 2 3 4 5 6 7 8 9

O NH

O

OH

O

CHIRALPAK IC

Hex/THF/DEA 85/15/0.1

min 0 2 4 6 8 10

NH

O

O

NO2

O

OOMe

CHIRALPAK ID

MtBE/MeOH/TFA 90/10/0.1

min 2 4 6 8 10 12 14 16 18

OOH

OH O

OH

OMe

HP

LC

Comparison of success rates between immobilized type columns and representative coated type columns for 123 compounds under Hexane/2-PrOH

Example of complementarity CHIRALPAK ID

Success rate with a single mobile phase combination!

Success rate

n-Hexane / 2-PrOH

81.3% 82.1%

89.4%

70%

75%

80%

85%

90%

95%

100%

ADH/ODH /ASH/OJH

IA/IB/IC IA/IB/IC/ID

N

COOC2H5H3COOC

H

NO2

N

H3COOC

O

H

NO2

ON

N

COOC2H5H3COOC

H

ONH2

Cl

n-Hex-MTBE-EtOH-DEA 50:50:1:0.1

0 5 10 15 20 25 30

Minutes

0 5 10 15 20 25 30

Minutes

0 4 8 12 16 20

Minutes

0 4 8 12 16 20

Minutes

n-Hex-THF-DEA 90:10:0.1

n-Hex-THF-DEA 90:10:0.1

n-Hex-IPA-Isopropylamine 80:20:0.1

Nitrendipine Benidipine

Nicardipine Amlodipine

N

H3COOC

O

H

NO2

ON

CHIRALPAK ID (250 x 4.6 mm, 5 µm); 1.0 mL/min; 254 nm; 25 C

Dihydropyridine-type drugs on CHIRALPAK ID

CHIRALPAK ID (250 x 4.6 mm, 5 µm); 1.0 mL/min; 254 nm; 25 C

Phenothiazine-type drugs on CHIRALPAK ID

n-Hex-IPA-DEA 80:20:0.1

0 5 10 15 20 25 30 Minutes

Dimetotiazine

N

S

N

NS

OO

0 2 4 6 8 10 12 14 16 18 20

Minutes

n-Hex-EtOH-DEA 100:0.1:0.1 (0.5mL/min.)

Profenamine

N

S

N

n-Hex-CH2Cl2-EtOH-DEA 95:5:1:0.1

0 2 4 6 8 10 14 18 12 16 20

Minutes

Thioridazine

N

S

N

S

Recognition in RP-mode

0 5 10 15 20 25 30 35 40

Minutes

HCOOH aq.(pH=2.0)/ACN=85/15

O CO2H

2-Phenoxypropionic acid

0 5 10 15 20 25 30

Minutes

Aminoglutethimide

20mM NH4HCO3aq. (pH=9)/ACN=30/70

NH

O ONH2

CHIRALPAK ID (250 x 4.6 mm, 5 µm); 0.5 mL/min; 25 C

Different elution profile and method optimisation

CHIRALPAK IA (4.6 x 150mm)

SFC CO2/ MeOH (0.5% DEA) 80/20

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 min 0 1 2 3 4 5 6 7

SFC CO2/ MeOH (0.5% DEA) 80/20

CHIRALPAK IC (4.6 x 150mm)

HPLC MeOH/DEA 100/0.1

min 0 1 2 3 4 5 6 7 8 9

CHIRALPAK IA (4.6 x 250mm)

min 2 4 6 8 10 12 14

HPLC MeOH/DEA 100/0.1

CHIRALPAK IC (4.6 x 250mm)

Analysis time: 12min

N

N

MeO

MeO

OMe

OMe

Tofisopam • Same enantioselectivity • Reduced column length • Reduced analysis time • No loss in resolution degree

CHIRALPAK IC-3 (4.6 x 150mm)

min 0 1 2 3 4 5 6 7 8

HPLC MeOH/DEA 100/0.1

Analysis time: 8min

Column transfer from 5µm to 3µm

SFC method development

Screening in SFC

CHIRALPAK IA

CHIRALPAK IB

CHIRALPAK IC

CHIRALPAK ID

Mobile phase system

EtOH, MeOH, 2-PrOH

Acetonitrile

Dichloromethane

Ethyl acetate

THF

MtBE

Primary Screen

CHIRALPAK AD-H

CHIRALPAK AS-H

CHIRALPAK AY-H

CHIRALPAK AZ-H

CHIRALCEL OD-H

CHIRALCEL OJ-H

CHIRALCEL OZ-H

CHIRALCEL OX-H

Co-solvents

EtOH, MeOH, 2-PrOH

Acetonitrile

Secondary Screen

Column dimensions: 150 x 4.6 mm, 5 µm; 3.0 mL/min; 25 C

Mobile phase selection in SFC Examples

SFC

CO2 / THF (0.3% TFA) 80/20

min 0 0.5 1 1.5 2 2.5 3

O

COOH

CHIRALPAK IA

CO2 / MeOH (1.0% DEA) 90/10

min 0 0.5 1 1.5 2 2.5 3 3.5

N

N

CHIRALPAK IB

CO2 / ACN (0.3% TFA) 80/20

min 0 1 2 3 4 5

O

OH

COOH

CHIRALPAK IC

CO2 / 2-PrOH (1% DEA) 90/10

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

N

O N

CHIRALPAK ID

N

O

OH

O

Indoprofen

CHIRALPAK ID Complementary properties

CHIRALPAK IA

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

CHIRALPAK IB

0 min 0.5 1 1.5 2 2.5 3 3.5 4 4.5

CHIRALPAK IC

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

CHIRALPAK ID

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

Ex

am

ple

s

in S

FC

30% MeOH (+0.2% TFA) in CO2 (150 x 4.6 mm, 5 µm); 3.0 mL/min; 150 bar

Isocratic or gradient?

0

5

10

15

20

25

30

35

40

45

0 2 4 6 8 10 12

Time (min)

*M

eO

H %

(by v

olu

me) MeOH gradient program

* Containing 0.3% DEA for screening of basic compounds

min 0 2 4 6 8 10 12

Cl

Cl

N

N

NOH

Diniconazole

CHIRALPAK IA

Back pressure: 150bar, Flow rate: 3.0ml/min, Temperature: 35°C

CHIRALPAK ID

min 0 2 4 6 8 10 12

N

O

OH

O

Ketorolac

CTE examples in SFC

Back pressure: 150 bar, Flow rate: 3.0ml/min, Temperature: 35

C

Containing 0.2-0.3% DEA for basic compounds

Main co-solvents and their relative eluting strength

MeOH > EtOH > 2-PrOH

MeOH > ACN THF

Time (min)

Co

-so

lve

nt

%

(by v

olu

me

)

0

10

20

30

40

50

60

0 2 4 6 8 10 12

0

10

20

30

40

50

60

0 2 4 6 8 10 12

Time (min)

MeOH-CO2 gradient ACN- or THF-CO2 gradient

Method development - SFC gradient programs

SFC-From gradient to isocratic

CHIRALPAK IB

min 0 0.5 1 1.5 2 2.5 3 3.5

min 0 2 4 6 8 10 12

MeOH-CO2 gradient

CO2 / MeOH (1.0% DEA)

90/10

N

N

Trimipramine

CHIRALPAK IC

min 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5

min 0 2 4 6 8 10 12

MeOH-CO2 gradient

CO2 / MeOH (0.3% DEA)

70/30

N

O

OH

N

Carazolol

CHIRALPAK ID

N

N

Bifonazole

THF-CO2 gradient

CO2 / THF (0.3% DEA)

70/30

min 0 2 4 6 8 10 12

min 0 1 2 3 4 5

CHIRALPAK IA

O

2-Ethyl-1-indanone

ACN-CO2 gradient

CO2 / ACN

95/5

min 0 0.5 1 1.5 2 2.5 3 3.5

min 0 2 4 6 8 10 12

SFC-From gradient to isocratic

Conclusions

Screening strategies by LC and SFC

• First objective is fast screen with high success rate

• Different derivatives introduce additional recognition options

• Solvent versatility is necessary to match resolution, solubility and chemical stability

Acknowledgements Team work

All other colleagues at

CHIRAL TECHNOLOGIES and DAICEL

DAICEL Chemical Industries, Ltd.

Tatsushi MURAKAMI

Atsushi OHNISHI

Chiral Technologies Europe

Dr. Tong ZHANG Dung NGUYEN