Crystallization strategies

www.approcess.com© APC Strictly Confidential

Crystallization strategies

the quest for increased yield and chiral purity

Andrew Byrne

APC Ltd.

Technobis Crystallisation Systems

4th February 2021


General Overview

➢ Brief Background

• Stereoisomers, chirality and Active Pharmaceutical Ingredients

• Ternary Phase Diagrams – Why are they relevant to Enantioselective Crystallization?

➢ Case Study 1: Ternary Diagrams as a tool for Process Definition

• Problem Statement

• Methodology & Development

• Outcomes, Conclusions & Recommendations

➢ Case Study 2: Enhanced Process Understanding & Yield Improvement

• Problem Statement

• Methodology & Development

• Outcomes, Conclusions & Recommendations

➢ Conclusions, Learnings & Acknowledgements

2


APC and Our Partners

3

Material to Medicine Process Research

▪ Engineering

▪ Formulation

▪ Scale-up

▪ Containment

▪ Analytical

▪ Chemistry

▪ Bioprocessing

▪ Continuous

140+Ph.D. Chemical

Engineers & Scientists

200+Medicines

8/10Big pharma

5/10Big biotech


Brief Background – Chirality & Pharmaceutical APIs

• Altered Chirality of Active Pharmaceutical Ingredients –> different physiological & pharmaceutical effects

4

• Revenue Commerical Chiral Small Molecule APIs 2019:

1. Lipitor (Atorvastatin) – Sales: ~ $ 1.9 billion

2. Plavix (Clopidogrel) – Sales: ~ $ 2.1 billion

3. Seretide (Fluticasone) – Sales: ~ $ 5.3 billion

4. Crestor (Rosuvastatin) – Sales: ~ $ 5.1 billion

• Separation & Purification

• Ideal Scenario: 100 % enantio. pure synthesis!

• Reality:

• Classical synthesis – Racemic mixtures – can be optimised

• Chiral resolution strategies

• Prochiral substrates + chiral catalysts -> improved ee%

• Chiral Chromatography – High cost

• Resolving Agents

• Crystallization – Key pharmaceutical Unit Operation

**

*

**

Mirror Plane


Brief Background – Chirality & Pharmaceutical APIs

• Glossary of Terms (Refresher) –

• Chirality:

Geometric property of molecules. A chiral molecule is non superimposable with its mirror image

• Achiral:

The opposite of chiral is achiral. Achiral objects are superimposable with their mirror images.

• Enantiomer:

Chiral compounds that are mirror images of each other and non-superimposable.

• Diastereomer:

Chiral compounds that are not mirror images of each other and non-superimposable.

• Eutomer:

The chiral enantiomer having the desiredpharmacological activity

• Distomer:

The chiral enantiomer having the undesiredpharmacological activity

5

Enantiomers

Enantiomers are stereoisomers that are non-superimposable mirror

images

Are always in pairs

Have identical physical properties except the ability rotate plan-

polarized light

Shape of the molecules is similar

Diastereomers

Diastereomers are stereoisomers that are non-superimposable and

are not mirror images

There can be several molecules

Have distinct physical properties

Have different molecular shapes

Vs

Lactic Acid Threonine2 pairs of Enantiomers

Or2 pairs of Diasteromers


Brief Background – Characterisation of the Solid Phase

➢Crystallization of Chiral Compounds

6

• Solubility measurement - determine the solubility of pure enantiomers (R or S) and their mixtures

• Eutectic Point – Highest solubility point obtained, Typically Symmetrical for Enantiomers!

Diastereomer Enantiomer


Case Study 1: Ternary Diagrams as a tool for Process Definition


• Title: Development of an optimized enantioselective crystallization process

• Processing Issues:

• Long process with Inconsistent key process output achieved post workup. Enantiomeric purity (AP%)),

yield, assay purity (wt%), variable heavy metals content (Rhodium, Iron and Zinc 1-200 ppm)

• Project Objectives:

a) To measure API solubility (conglomerate) and determine the effect of heavy metals on solubility (if any)

b) To develop an optimized enantioselective crystallization process for the purification of crude API (desired enantiomer)

c) To demonstrate this process on 10 g & 100 g scale

7



8

➢Key Findings During Tech Transfer (Part 1)

Sample Description

HPLC

Assay

HPLC

Chirality

ICP-MS

Rh

ICP-MS

Zn

ICP-MS

Fe

wt%AP% Desired

Enantiomerppm ppm ppm

Hydrogenation: crude reaction material

(Evaporated to Dryness)60.4

91.7

Range: 87 -92%1900 31000 1700

Charcoal treatment #1 – dry cake 73.5 92.1 1200 29000 1300

Post water slurry # 1 – dry cake 86.1 94.7 800 1400 1900

Post Isopropanol reslurry # 1 – dry cake 84.8 95.9 39.9 36.8 71.8

Charcoal treatment #2 – dry cake 96.1 96.2 1.5 28.1 49.6

Post Isopropanol reslurry # 2 – dry cake 97.2 97.5 1.0 38.4 35.4

➢Unit Operations: Solid phase analysis (Assay, Chirality & Heavy Metals)

With Each Unit Operation:

• Assay (wt%) increases

• Chirality (AP) increases

• Heavy Metals content…

Early indications:

Charcoal Treatments



9

➢Key Findings During Tech Transfer (Part 2)

➢Unit Operations: Liquid phase analysis – Loss to mother liquor

Process Run Sample DescriptionDesired

Enantiomer

# - mg/g

1 Post water slurry # 1 11.3






1 Post Isopropanol reslurry # 1 3.8






Reverse Phase - AssayDesired

EnantiomerUndesired

Enantiomer %Desired

mg/g mg/g

5.4 5.9 48.1

5.4 5.6 49.0

5.1 5.9 46.3

4.9 5.4 47.8

4.9 5.3 47.9

6.9 5.6 55.3

1.8 2.1 46.1

1.5 1.8 46.2

2.0 2.1 49.5

1.7 1.7 50.6

1.6 1.3 54.3

1.4 1.8 43.3

Normal Phase - Chirality HPLC method - Key

0

40

80

120

160

200

10 20 30 40 50 60

Solu

bili

ty (

mg/

g)

Temperature (°C)

Normal Phase - Chirality

Reverse Phase - Assay


Crystallization Workflow – Overview Early Phase Solvent Selection

10

Commercial Benefits: accelerated screening of conditions leading to time & cost savings



11

➢Preliminary Computational Solvent Screening

• Computational Quantum Calculations using both COSMO & Dynochem Software packages

• Relative Solubility calculated based on electron density mapping & Chemical Potential

• 27 Solvents screened – Key Outputs guided Screening Process

0

1

2

3

4

5

6

7

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

Rel

ativ

e So

lub

ility

25 degrees Celcius

-2.0

-1.5

-1.0

-0.5

0.0

0.5

1.0

Hex

(So

lvat

e P

rop

ensi

ty)

High Solvate Propensity

Low Solvate Propensity

PotentialPoor Solvents

Good Anti-Solvents

PotentialGood Solvents

Poor Anti-Solvents

www.approcess.com© APC Strictly Confidential 12


➢ Solubility Design Space: the Ternary Diagram (Method 1)

• Classic Isothermal Method (Both Enantiomers & Mixtures)

• Synthesis of the Undesired Enantiomer was required

• Excess solids into known volume / mass of solvent mixture

• Allowed to equilibrate over extended hold period at fixed temperature

• Slurry was allowed to settle and filtered supernatant Liquor Concentration was measured by HPLC

• Concentration collected as mg/g and converted to wt%

• Data collected for a number of pure and binary solvent systems

• Remember – HPLC method selection here is critical

Note:

• Crystal 16 – Clear point determination was also used.

020406080

100120

0 10 20 30 40 50 60 70 80 90 100

Des

ired

So

lub

ility

(m

g/g)

Isopropanol (Volume %)

10 °C 20 °C 40 °C 60 °C

Simple EutecticComposition 50:50


Method 2: Ternary Diagram Construction


• Two fold objective: a) Characterise the solid phase (binary & Ternary)

b) to generate solubility design space

• Methodology

• Crystal 16 – Clear point determination

• 100% Transmission corresponds to solubility of a given solids mixture

• Different masses of solid are typically charged to different vials to obtain the clearpoint temperatures of each vial and build up the solubility relationships.

• For our system solid wt% mixtures of Desired Diastereomer : Undesired Diastereomer

• Range of solid wt% mixtures: 100:0 Desired: Undesired -> 0:100 Desired: Undesired

• Generation of traditional solubility curves for different solids mixtures was therefore

possible at varied solvent compositions (0 to 100 % Desired)

• Convert data to wt%, isothermal cross-sections via data interpolation

• Spot checking by HPLC – For both concentration & Chirality

13

Slurry

undissolved solids

Solution

Dissolved solids

Clearpoint – 100% Transmissivity


0

20

40

60

80

100

120

140

0 10 20 30 40 50 60 70 80

AP

I s

olu

bili

ty (

mg/

mL

solv

en

t)

Temperature (oC)


14

80:20

90:10

70:30

60:40

20:80

50:50

Solubility Design Space A:B MixturesIsothermal Cross Sections • Isotherm 60 oC

• Isotherm 40 oC• Isotherm 15 oC

Interpolate

Solubility

&

Convert to wt%

Simple EutecticConglomerate system

A:B

Method 2: Ternary Diagram Construction

Enantiomer A

Solvent Enantiomer B



➢Simple Eutectic Verification Experiments (HPLC tracking)

Solvent DescriptionDesired

Enantiomer

Undesired

EnantiomerSolvent

Desired

Enantiomer

Undesired

Enantiomer

Composition Solids Charged mg / g mg / g wt% wt% wt%

Slurry - Liquor Composition (in presence of excess solids)

Vial 1: 100 vol% MeOH Charge 1: Desired Enantiomer 21.34 1.31 97.73 0.13 2.13

Vial 1: 100 vol% MeOH Charge 2: Undesired Enantiomer 21.95 22.60 95.55 2.26 2.19

Vial 2: 100 vol% MeOH Charge 1: Undesired Enantiomer 0.25 21.39 97.84 2.14 0.02

Vial 2: 100 vol% MeOH Charge 2: Excess Desired Enantiomer 21.53 21.97 95.65 2.20 2.15

Vial 3: 100 vol% MeOHCharge 2:

Excess Desired Enantiomer & Excess Undesired Enantiomer20.26 21.82 95.79 2.18 2.03

20 oC isotherm

Pure materials

20 oC isotherm

&

Eutectic Investigation

Experiments



➢Does residual metal content effect API solubility?

• Historical Batch Data – Highlighted this as a significant Issue

• Using Normal Phase HPLC method – Accounts for Chirality

Solids Charged

ICP-MS

Rh

ICP-MS

Zn

ICP-MS

FeTemp.

Desired

Enantiomer

Undesired

Enantiomer

ppm ppm ppm °C mg / g mg / g

Isolated

Pure Material0.45 9.13 5.33

10 20.7 0

20 21.0 0

40 46.8 0

Water Slurry #1

Process

Midpoint

Dry cake

800 1400 1900

10 16.2 1.9

20 21.9 2.1

40 41.8 2.4

Crude reaction

Material

(dry cake)

1900 31000 1700

10 20.1 8.9

20 26.2 14.6

40 53.2 13.6



➢Proof of concept Enantioselective Crystallization runs (Part 1)

• Starting point: Liquor 70:30 (by Mass) Desired: Undesired in selected solvent system 50:50 IPA: Water

14.2

23.1

37.2

61.683.2

14.924.2

39.637.637.1

0.72

0.78

0.940.90

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

0

10

20

30

40

50

60

70

80

90

5101520253035404550556065

Frac

tio

n o

f D

esir

ed in

So

lid P

has

e

Co

nce

ntr

atio

n o

f En

anti

om

er in

liq

uid

p

has

e (

mg/

g)

Process Temperature (°C)

➢In-Situ PAT: Understanding the Crystallization ➢Off Line HPLC Sampling (Crystallization)

• Sampling of Solution & Solid phase – Chiral content

• Desired Enantiomer – Solution phase Concentration

• Undesired Enantiomer – Solution phase Concentration

• Desired Enantiomer – Solid phase Fraction

5 61 4

1

2

3

4

5

6

Internal Reactor Temperature (Tr)

In-Situ FBRM: Counts No Wt. 1-1000

In- Situ FTIR: Peak Height at 1439 cm-1



➢Proof of concept Enantioselective Crystallization runs (Part 2)

• Starting point: Liquor 70:30 (by Mass) Desired: Undesired in selected solvent system 50:50 IPA: Water

➢In-Situ PAT: Understanding the Crystallization ➢Off Line HPLC Sampling (Crystallization)

• Sampling of Solution – Chiral content

• Expressed as wt% & plotted on ternary1

2

3

4

5

6

Internal Reactor Temperature (Tr)

In-Situ FBRM: Counts No Wt. 1-1000

In- Situ FTIR: Peak Height at 1439 cm-1


➢ Ternary Diagrams as a tool for Process Definition

• Tech Transferred Process Flow:

• Modified Process Flow:

• Advantages:

• IPC for Range of operation 87 AP% to 92 AP% Desired from reaction stream

• Shorter Duration & Scalable Process Developed - Removes Facile operations

Carbon Treatments x 2, Drying x 1, Filtrations x 2 & Distillations x 2

• Higher Assay & Enantio. purity >99%, Yield % increase, Heavy Metals Purge <20 ppm

Case Study 1: Conclusions

19

OperationAssay(wt%)

Chirality(AP%)

ICP-MSRh

(ppm)

ICP-MSZn

(ppm)

ICP-MSFe

(ppm)

Crystallisation 99.3 99.2 29 250 50

Recrystallisation 99.9 99.9 3.7 3.4 19


Case Study 2: Enhanced Process Understanding for Yield Improvement


• Title: Enhanced Process Understanding for Yield Improvement

• Processing Issues:

• Isolated yield of telescoped reaction & crystallization step 81 % at plant scale. Incomplete mass balance for

active pharmaceutical ingredient and hence fate of desired API yield was unknown.

• Tightly defined limits in Global filing

• Project Objectives:

• Enhanced process understanding to Investigate process yield improvement opportunities

• Reduce process wastes and increase productivity with initial target of 4% yield improvement (Current Yield: 81%)

20


Data Rich Experimentation – Process Assessment

21

00

Event 1: Methanesulfonic acid addition; temperature Increase; dissolution of starting material Event 2: Effective Seeding of batch

Event 3: Desaturation during 3 h isothermal hold

Event 4: Desaturation during cooling profile

Event 5: End

Figure 1: Standard process followed by in situ PAT: PVM and FBRM


Mass Balance – Standard Process Conditions

API mg/mL

concentration

Volume

of Liquor (mL)Description

Mass Lost

grammes% of total Yield

16.3 90 Mother Liquor 1.470 12.41

4.6 26 Wash 1 0.119 1.01

1.9 13 Wash 2 0.025 0.21

N/A N/A Isolated Solids 10.120 85.47

Total 99.10 %

• Typical Process Yield: 75 – 87 % (APC Yield 83 – 85 %)

• Mass balance: 99 – 96 % (Loss to ML – 10.8 to 12.4 % of batch)

• Proven Acceptable Ranges defined by FilingUnderstanding the Crystallisation Roadmap

• Seeded cooling crystallization – end point liquor = starting liquor composition

• C16 solubility by clear point for Standard crystallization mother liquor

• Please Note: -10 oC solubility by HPLC

Data Rich Experimentation – Process Assessment


Solubility Design Space – Proof of Concept (Artificial Solvent Systems)

23

#

Acetone

(vol%)

Water

(vol%)

70%

Acid

(vol%)

Ratio

Acetone/Wa

ter

100%

Acid

(vol%)

100%

Water

(vol%)

1 80.9 17.0 2.1 4.59 1.47 17.63

2 78.4 17.8 3.8 4.14 2.66 18.94

3 77.8 17.7 4.5 4.08 3.15 19.05

4 71.3 17.1 11.6 3.46 8.12 20.58

5 82.7 17.3 0 4.78 0 17.30

6 80.6 19.4 0 4.15 0 19.40

• The Global filing indicated ranges of operation for the process solvents – variation of these ratios (volumes) within the filing could yield

altered solubility. This possibility was investigated.

• The solubility of the target product in these solvent mixtures was studied using clearpoint by Crystal 16.

• HPLC was utilised for the solubility at -10°C .

87.7

11.0

82.3

9.4

82.0

8.5

56.6

4.5

85.3

36.2

85.3

37.1

0.0

10.0

20.0

30.0

40.0

50.0

60.0

70.0

80.0

90.0

100.0

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65A

PI s

olu

bili

ty (

mg/

g)

Temperature (°C)

API (Artificial Solvent Systems)

Solvent Mix 1

Solvent Mix 2

Solvent Mix 3

Solvent Mix 4

Solvent Mix 5

Solvent Mix 6

• Variation of the solvent ratios within accepted PAR leads to altered solubility in artificial solvent systems.


Solubility Design Space – Proof of Concept (True Process)

24

Process Liquor Composition 1

35.3

104.1

11.3

54.7

19.0

92.5

0.0

10.0

20.0

30.0

40.0

50.0

60.0

70.0

80.0

90.0

100.0

110.0

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65

AP

I so

lub

ility

(m

g/g)

Temperature (°C)

API solubility: Varied liquor compositions within PAR• Working within PAR defined by Filing

• C16 solubility by clear point for Standard crystallization mother liquor. • Please Note: -10 oC solubility by HPLC

• Variation of the solvent ratios within accepted PAR leads to altered solubility

• Can be harnessed to increase process yield!




Solubility Design Space – Proof of Concept (True Process)

25


35.3

104.1

11.3

54.7

19.0

92.5

0.010.020.030.040.050.060.070.080.090.0

100.0110.0

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65

AP

I so

lub

ility

(m

g/g)

Temperature (°C)

API solubility: Varied liquor compositions within PAR• Working within PAR defined by Filing



Process Run IDIsolated

Yield

Loss

Mother Liquor

Loss

Cake wash 1

Loss

Cake wash 2

Mass

Balance

– (%) (%) (%) (%) (%)

Process Liquor Composition 1 85.4 12.4 1.0 0.2 99.1




Process Development – Defining the Future State (in-situ FBRM & FT-IR)

26

Isolated yield: 90.8%

Mass Balance – Modified Process (High acid/water; low acetone; median MS Acid)

mg/mL concentration

Volumeof Liquor (mL)

DescriptionMass Lostgrammes

% of total Yield

8.58 565 Mother Liquor 4.84 5.84

2.89 158 Wash 1 0.45 0.55

1.05 138 Wash 2 0.14 0.18

N/A N/A Isolated Solids 75.36 90.76

Total 97.32

• Crystal 16 solubility by clear point for standard, modified process & PAR limits crystallization mother liquors.

• New setpoint established within PAR limits

• From process development work, the concentration of the API lost to the mother liquor has been decreased from:

16.33 mg/mL → 8.58 mg/mL

• Results in a ~7% isolated yield increase.equivalent physical and chemical properties (purity, form & PSD)

In- Situ FTIR


Conclusion – How it performed on plant?

27

~7% isolated yield increase.


Overall Conclusions

28


• Stereoisomers, chirality and Active Pharmaceutical Ingredients

• Ternary Phase Diagrams & Enantioselective Crystallization

➢ Case Study 1: Ternary Diagrams as a tool for Process Definition

• Methodology & Development: End-to-end process definition & Enantioselective Crystallization

• Development of solubility design space (‘Roadmap’)

• Shorter Duration & Scalable Process Developed - Removed Facile operations

• More Robust & Cleaner - Higher Assay & Enantio. purity >99%, Yield increase, Heavy Metals Purge <20 ppm

➢ Case Study 2: Enhanced Process Understanding - Yield Improvement

• Methodology & Development: data rich experimentation

• Development of solubility design space (‘Roadmap’) & models to describe the unit operations

• Enhanced process knowledge leading to increased yield


Documents

Crystallization strategies