Pharmaceutical Polymorphism Studies by DSC

good morning everyone I'm sorry that we

are starting just about eight minutes

later than planned there are a few

technical errors in the internet

connection on my side I hope this will

not prevent us from continuing my name

is John Ernie

I'll be presenting today's webinar on

polymorphs and pseudo polymorphism by

means of differential scanning

calorimetry just as an introduction I

wanted to let everyone know that you can

use your GoToWebinar tool panel to ask

questions I would very much look forward

to receiving questions from you the

beginning of this presentation the first

45 minutes will be presentation

component and then we'll follow that

with a question and answer session with

that being said and because I know we

were beginning just a few minutes late I

want to go ahead and just get right into

this so here at net we the niche family

comprises three different groups of

three different companies the analyzing

and testing portion is the group that I

work for this includes thermal analysis

instrumentation and determination of

thermo physical properties so we're

definitely an old old company we've been

around since the 1870s

we're mid-sized company and apart from

the analyzing and testing groups

especially for pharmaceuticals it's good

to know that we also are on the process

side of things with grinding and

dispersing as well as pumps and systems

for pharmaceutical grade applications

very small a bit about your host today

my name is again John Ernie I am a

regional Technical Manager and regional

technical sales associate for neck

covering here in the u.s. Texas

Louisiana Oklahoma as well as Mexico my

background is in chemistry I've worked

in the polymers oil and gas and

pharmaceuticals industry primarily from

an analytical chemistry standpoint I've

been with net since 2017 in my current

role here based in Houston Texas and

covering pharma for the US

but most accounts here within within my

local region so let's get right into it

what we're doing is we're looking at

polymorphs using differential scanning

calorimetry if you're not familiar with

the technique of DSC many of the we're

not going to go into fundamentals

because we covered that in the previous

component of this series in part one we

are now in part two but even if you're

unfamiliar with DSC we'll be talking

about the concepts throughout because

that's primarily the data that we'll be

investigating so what we have here is

what you would generally expect in a DSC

trace of a pure pharmaceutical compound

on the y-axis of course we have what is

called the heat flow this is either

endothermic or exothermic events

associated with the heating or cooling

of a compound what we have here is a

ribavirin this is an anti antiviral

compound that we've got here we've got a

sample mass of 1.0 two milligrams we're

heating or heating our sample at ten ten

degrees Kelvin or 10 degrees centigrade

per minute in aluminum crucibles

nitrogen atmosphere and what we can see

in the range of 140 degrees C up through

180 degrees C is we have a very clearly

defined onset of our of our melting so

as we're heating the material we have no

no endothermic or exothermic events

until we reach the melting of this

crystalline compound so we have the

so-called onset temperature which is an

extrapolation of the baseline and the

tangent here with the this the slope of

this melting peak we also have the maca

point of maximum heat flow so you can

see the onset and the peak temperature

and you're in your pharmacopoeia you

should have both of these very well

defined as well as one thing we can look

at is the area under the curve here so

this is again a very well defined

melting of a crystalline compound very

straightforward but let's look into

let's look into a little bit more you

know more complex examples

this particular compound can exist as

two different polymorphic forms a and B

and as I mentioned we're looking at

polymorphs using DSC so we'll be

defining polymorphs and looking more

into DSC throughout the course of this

of this webinar so how do we get these

different forms of this same material

form a we're looking at a material

that's crystallized out of aqueous

ethanol so ethanol with a with a water

component and it has a given melting

range from 166 to 168 degrees Celsius

but then there's also a form be

crystallized from pure ethanol and that

has a melting range that is a bit higher

174 to 176 degrees Celsius so for this

particular substance it has the same

stoichiometry the same chemical formula

but the physical structure of the

substance is different so for certain

substances we have not only one melting

point but we can have multiple melting

points the example that we showed before

is of course form a based on the melting

point that onset and the peak

temperature that we saw but one of the

ways that we can distinguish between

polymorphs as we can see if I have a and

B and I can I can find out what their

melting is of my real sample this allows

me to distinguish between the two

different polymorphic forms and this is

a very basic very first form of using

DSC for polymorph studies so let's dig

in just a little bit deeper into what is

polymorphism so this is the exact same

material and we're looking at solid

materials existing in different crystal

structures so again same stoichiometry

same chemical composition you might be a

little bit more familiar if this is a

difficult concept to grasp or this if

this is new to you

chemical elements also exhibit this not

just compounds not just maliki not just

you know hetero molecules but also just

individual elements so for example

example carbon can exist as a graphite

as diamond or as these so-called

fullerenes we have the same carbon

molecules they're simply arranged


in a stacked and layered form either

small groups with graphite or large

sheets with graphene we can have a

tetrahedral arrangement of these same

molecules with diamond or of course then

these buckyballs or fullerenes

buckminsterfullerene x' as they're

sometimes known in a completely

different orientation but still the same

still the same element

so that's polymorphism so the same

material existing in different crystal

structures we also have pseudo

polymorphism and this is going to be a

special case of polymorphism in which

the different crystal types are simply

the result of water addition so

hydration or salvation so crystal

structures that have incorporated within

them some of their some of the local

solvent molecules be that water or be

that another solvent molecule so no no

overview is complete without a very

brief historical view going back all the

way into the 80 end of the 18th century

so 1798 Martin Clapp Roth looked and saw

that calcite and aragonite are just

simply two different forms of calcium

carbonate so the exact same chemical

structure this calcium carbonate exists

in an orthorhombic or in a rhombohedral

geometry and based on thermodynamics and


so basic basically looking at what

temperatures is the material existing at

and at what time how long has it been at

that given temperature one form can then

be converted into the other form within

a few days we can also see that there's

then even a third polymorph with a

different crystal structure which is

even of course less stable then and then

the actual term polymorphism come

meaning multiple shapes coming from that

that Greek base was defined already

early in in the 1820s so this is nothing


but we do have new means of studying

these tech you know these different

structures and why is it important and

this is really a the question ultimately

when we're going to be look using

analytical instrumentation or spending

our time investigating substances what's

what's the real critter

reason for us to do this so having the

same material the same substance within

with difference different organization

you know different polymorphic forms

this can adjust the melting point the

sublimation temperature as you can see

here heat capacity volume viscosity so

those are all physical properties but

from a pharmaceuticals perspective the

real impact is that the process ability

of drug substances and the shelf-life so

we're looking at stability how

hygroscopic the material is that is how

much water does this absorb and then of

course that result that relates again

back to shelf life and that can relate

to bioavailability and we're also we

also need to look at dissolution so the

same crystal strike you know say the

material with different crystal

structures can dissolve differently they

can be available for for activity within

the body differently and they can have

different shelf lives even though they

have they came same chemical chemical

structure so I mentioned we would go

into DSC and we will be looking a little

bit more into the DSC I apologize here I

actually have some German slides content

I know this is the English language

version but we'll work through it as is

the convention we can display the DSC

traces either with endotherms so heat

flow into the sample up or downwards it

is the standard European convention to

display endotherms in an upward fashion

so melting Peaks will be displayed in an

upward fashion I know that here in the

US where I'm broadcasting from many many

providers of thermal thermal analytical

instrumentation have the opposite

convention all software packages can

allow you to reverse this so really we

get the exact same information it's just

a question of is endo up or endo down so

we also have this nicely

color-coordinated red and blue red

intuitively for heating blue more

intuitively for cooling and so we have

here is indium this is one of the most

commonly known calibration standards

used for looking at DSC now what makes

it a calibration standard this is just a

pure metal and it has a very very

clearly defined onset temperature

literature one 56.7 we can see or see

here the onset temperature of melting

again as the tangent of the intersection

of the tangents of the baseline and this

rise in the melting this onset

temperature of 156 point six so very

very close to literature here the other

important point is the area

sorry that's again German here but the

area under the curve in joules per gram

so how much heat energy is required how

much energy is required to go from solid

to a completely liquid state so that's

the area under the curve here and you

can see as we heat the material up we

get a melting that is then complete we

can then take the material and go from

these high temperatures down to low

temperatures and we go through then the

crystallization we can then see very

very closely matched the area under the

curve but what you might also notice

then is that we have an offset there's

going to be a different process that

describes or that or that dictates

crystallization from melting melting is

a thermodynamic event whereas the

crystallization is a kinetic event if

these terms are a little unfamiliar to

you the thermodynamic event is going to

be is going to take place at the given

temperatures relatively independent of

of time scaling however when we go from

the liquid to a solid we're going

through a crystallization what's going

to happen is that crystallization is

going to be time dependent it actually

it takes time for those there's liquid

molecules to orient themselves in

relation to each other and form that

crystal lattice so that's why we're

seeing this offset known as hysteresis

between the onset of melting and then

the onset of cooling but really what's

important for for a calibration standard

of any type is that you have

repeatability repeatability

reproducibility precision and so what we

have is simply seven runs of the same

material heated cooled heated cooled

overlaid and very nicely consistently

showing us the exact same properties

this is this is one of the ways that

we're going to not only calibrate our

system but we're also going to use this

as a consent

chill tool to understand what's going on

in our system with a simple metal as

we're heating and cooling we're getting

complete reproducibility not necessarily

so with our polymorphic materials you

know this is a simple metal when we're

looking at more complex structures and

we get into polymorphism it's not always

so straightforward and it doesn't have

to be a complex structure for us to get

something that's that's not 100%

straightforward if we take ice very very

pure deionized filtered water we make

sure that we use a very clean crucible

this is an aluminum crucible that's been

it's been a solvent cleaned baked we

make sure that the water is extremely

pure and as we go from solid to liquid

of course everyone understands the

melting of ice very nice and easy

conceptual process we have that onset

right near zero degrees Celsius and then

it's completed but then what we see

under cooling is we get the material all

the way down here to negative 15 degrees

Celsius and there's still no

crystallization that's taking place

again that that melting is a

thermodynamic process the cooling is a

kinetic process and this is something

that you might have seen before this is

known as super cooling we reach here

some critical point where there's the

onset of crystallization after which

point we have a small seed crystal and

then that provides a lattice after which

the rest of the water molecules which

are already well below their actual

freezing point then arrange themselves

around that we get a very rapid rapid

cooling so looking at polymorph so we've

we've looked at some basic calibration

materials and very common materials

indium and water when we're talking

about polymorphs it would be very nice

if there were a standardized way of

describing these different forms of the

same substance and in the beginning what

we what was what was attempted was to

use you know Greek letters alpha beta

Roman numerals one two latin uppercase a

B to characterize these forms in a

systematic way and the idea was

the modification or the form of the

material with the highest melting point

isn't generally named form one alpha or

a unfortunately this is not always the

case sometimes a more stable form is

found later and then things are changed

so while in general you can see form one

or a or alpha as as the highest melting

point temperature or highest melting

point form this is of course this is not

always the case so take this with a

grain of salt you'll also say things

metastable modifications so things that

are not pure that are not permanently

stable sometimes marked with with this

Prime and so we need to form let's

formulate some some concepts around

polymorphism before we actually look at

the DSC data what I'm going to do is go

through a few more conceptual ideas

before we go through and look at about

four different examples of polymorphism

studies and what can what we can pull

out of it from the DSC data so let's

take this empirical observation so we

take a material and Kulim the material

from the melt what we end up finding is

that the thermodynamically least stable

modification crystallizes first and then

you get this rearrangement this

recrystallization and you get a step by

step formation of more stable forms so

these are these are discrete steps this

isn't the material partially

crystallizing and then partially

crystallizing depending on the kinetics

depending on the rate of cooling what we

can arrive at is a discrete series of

these metastable States and then we end

up in this more stable phase which again

will have that higher melting point

along with these crystalline forms again

depending on the cooling because it's a

kinetic process a time-based process we

can also end up with an amorphous phase

so phases a morphic crystalline let's

look at a little brief overview of

different states of material different

phase transitions so that we're all on

the same page this is coming from a text

it's a text from the 1980s very common

very commonly used in Germany also

translated into other languages but what

have here on the left-hand side we've

got entropy and enthalpy of our

materials increasing as we go up we have

density of our material decreasing then

as we go down so if we look at a

crystalline form of a material let's say

the Alpha form the most stable form this

is something we're very familiar with we

know this terminology we have melting

and crystallization as we go from that

crystal form to a liquid form we could

also have a less stable form and we

could have that solid transition so we

can have that crystal transition we can

have a solid solid transition from

crystal form a to crystal form B of

course we also know the terminology here

basic physical chemistry going from

liquid to a gaseous state we've got

evaporation and condensation but these

don't describe the full the full range

of possibilities we have sublimation and

and then D sublimation going from

gaseous directly just solid forms you'll

see this with with things like liquid

nitrogen and solid and dry ice but then

we have the so called liquid crystals

which are a little bit more liquid but

have retained some crystalline structure

to them we also have glassy state

materials which are more solid but which

retain some degree of that liquid that

liquid form so there is quite a large

range of different material phases and

the important thing is that based on

their thermal thermodynamic stability

and the kinetics we can use the

differential scanning calorimeter to

pull out what phase of material do I

have or what phases of material do I

have so let's get into some of the

examples this is what I know a lot of

you are waiting for and this is to me

the really exciting part is we've looked

at some of some of the background let's

look and see the the data what can we

actually pull out from some some

concrete examples you might recognize

sorbitol this is a a c6 sugar this is

something that you'll find in a lot of

different food products also a sweetener

for various different pharmaceuticals

and what all what I will show at the at

the bottom are just experimental

conditions often with these materials

we'll have somewhere between one and

five milligrams

um most of the USP the u.s.

pharmacopoeia as well as other other

standards recommend for doing basic

studies we're looking at about 10 Kelvin

per minute heating and cooling but if we

want to do particular studies we can of

course change this we can go slower we

can go faster depending on the

particular goal of our experiments and

whether or not we are using a particular

standard or we're doing our own

individual Rd we might see aluminum

crucibles either pierced or oh you know

pierced and open to the environment or

closed and sealed since we're showing

nitrogen atmosphere of course this would

then be a not sealed component not

sealed experiment

so as we're heating our material what we

see is a single crystalline phase we

have the onset onset here and then we

have one simple melting but then after

we cool let's say we cool this material

we take it from solid to liquid we then

cool the material and we cool it at 10

degrees Kelvin per minute then we heat

it up one more time now this looks

completely different what's what's going


what I mentioned is that cooling you

know the crystallization is is a kinetic

of that event or a series of kinetic

events it's not simply that we reach a

certain temperature and the material

crystallizes it's a time-based process

so if we're cooling this material at 10

degrees Kelvin and then we immediately

start heating the material we go through

this step change we go through a glass

transition and then we don't see a

crystallization what this means with a

little bit of knowledge of the materials

and and DSC is that we were actually in

an amorphous state glass transitions are

associated with materials with with an

amorphous phase however if we then go

through and store this material one day

at room temperature we allow the

kinetics of crystallization to occur we

then see a different we do see a

difference we see a different story

we heat up the material again and then

we have not this melting you know at

this onset of 95 degrees but then we

have an onset melting of you know down

in the upper 70s we have an onset of

melting down here around 50 or 45

so we have different phrases of this

material simply after being stored for a

number of days so depending on them on

the shelf life that we're interested in

we need to look at storage conditions we

need to look and see well if we want to

maintain an amorphous phase do we store

it at room temperature or do we need to

store at a lower temperature and what

are the actual physical properties of

these of these you know what is my goal

what is what form do I actually want to

have so this is going to dictate our

storage conditions but this is how we

can use DSC as a very simple tool to

evaluate what form have I received my

material in vs. how do I want to then

process that material and send it out so

this leads us into a few more

definitions we don't only have one

simple type of polymorph there are

multiple different types of polymorphs

there are those that are reversible they

go through reversible phase transitions

there are also those which go through

irreversible phase transmissions so we

have the so-called and anti tropic

transitions these are reversible so as

we go from a low temperature state to a

high temperature modification we have

one form that is under all conditions

more stable at higher temperatures and

one that is more stable at lower

temperatures so if we look at a plot of

the Gibbs free energy of the two

different phases are the two different

polymorphs below a transition

temperature this TT we can see the pink

material has a lower Gibbs free energy

ie will be more stable below this

transition temperature whereas this

alpha the Alpha form above the

transition temperature has the lower

Gibbs free energy and this will be more

stable and at this higher temperature

range so depending on whether we want to

let's say that we want to have primarily

polymorph a what I can do is allow the

kinetics of recrystallization to happen

above the transition temperature and

this higher temperature this at this

higher temperature I'll end up with more

of this polymorpha a if I'm interested

in in polymorph B then what I can do is

maintain my material down at this lower

temp in this lower temperature

to ensure that I have the more stable

material down here and we can formulate

we can formulate you know reverse

reversible and irreversible polymorphic

transitions and look at these different

in ENT tropic forms there's a few

different rules that came out of the

1970s these were published in

microquimica ACTA by a professor Berger

and these are so-called the Berger rules

and the RAM Berger rules and these are

these relate to solid solid

transformations again going between

different solid states of of these

materials going between different

polymorphic forms and so what are these

rules say how do they guide us the only

reason we want to have rules is to help

us think through and understand what's

going on so the transition enthalpy of

these two in anti tropic forms that is

reversible forms so above the transition

point we're going to have an endothermal

effect going and below the transition


we actually have an exothermic event

endothermic and exothermic that's when

we're looking back to our DSC trace and

seen do I have heat flow into my

material endotherm or heat flow out of

my material exothermic so this is going

to be a nice guide for us looking at us

at a DSC trace and knowing whether I'm

getting heat flow into or out of am i

above or below my that transition

temperature so that's the heat of

transition rule if we're looking at the

heat of fusion rule so this is instead

of looking at transition heats what

we're doing is here is looking at

differences in the melting heat so the

melting enthalpy so if I have form a

versus form B am I going to get am I

going to have a higher heat of fusion or

higher heat of melting of form a or form

B how am I going to know well the higher

melting form so this is the material

that melts at a higher temperature it's

going to have a lower melting enthalpy

so that's the area under the curve for

this for this particular and antium a nd

sorry enantiomorphic pair so it's a

really nice simple rule the higher

melting form has a lower melting

enthalpy if the higher melting form

though also has a higher melting


the relationship is mana topic what does

that mean it means instead of

enantiotopic having this reversible

transition if the higher melting form

also has the higher melting enthalpy we

all we have a situation which is mono

tropic and we'll talk about monitor

traffic forms here in just a little bit

so you can imagine we have a lot of

different ways that DSC traces could

look going and we're looking here at

some data that's very very nice this was

again published in the Journal of

thermal analytical calorimetry this was

from a Swiss researcher working with

pharmaceuticals and this shows heating

curves from a DSC and a lot of different

potential ways that the curves could

look and we'll break them down one at a

time so what do we have so in this case

we have two different forms we have a

pharmaceutical compound which has two

different forms simply a and B and so

what's going on as we're heating our

material we have in this first case an

endotherm small endotherm and then

another larger endotherm so what we have

here is stable form a transforms and

form B so this is an endothermic effect

and then that form B melts what about in

conditioned to the next one down stable

form a simply melts and then the

transformation in to form B is hindered

kinetically what does that mean that

it's it's kinetically hindered it means

that we're going through this process

too quickly where we might transition

into into a into B this heating is

happening too quickly and this process

would take time so kinetically hindered

means that the process is happening too

quickly and the thermodynamic process

takes over so what about example 3 so we

have stable form a that's melting form B

so stable form a melts here form B is

growing from that melt so we're getting

a recrystallization into form be that

recrystallization is providing this with

a Morse from a dynamically stable

compound so that's giving off heat

that's why we have an exotherm and then

a form B is melting if we look down to

to this fourth example what we could

have is a meta state

Formby it's transforming into form a so

metastable form b has a higher Gibbs you

know has a higher Gibbs free energy then

that stable form a so we're giving off

heat to trans to transition from B to a

but then a melts a trans for re a

transforms into B and then B melts so we

can see a lot of different transitions

metastable be transforming to a a

transforming to B be melting and then of

course at the very end we have the most

simple version which is metastable form

B melts we have no transition to a no

transition back back to form b so let's

look at let's go ahead and look at this

let's let's go back to some DSC data if

i if i want to look at this enantiotopic

transition i have two different phases

of cesium chloride III so I have one

phase that is a face centered cubic and

one that is a body centered cubic so

this would be a structure that is more

like sodium chloride you know whether we

have a cubic like a straight square

versus something that looks more a

little bit more orthorhombic so cesium

chloride goes through these reversible

phase transitions just like the indium

or in a similar fashion to the indium

whereas the indium melts and

recrystallizes into a single form the

cesium chloride goes between two solid

forms so on the first heating we've got

the first teethin in red and i have a

nice onset temperature that occurs here

and then cooling I then go back into the

body centered polymorphic phase

polymorphic structure and this is then

something that occurs repeatedly I then

have a second heating and once again I

have this nice stable onset we can see a

slight difference we can see a slight

difference here and you might say well

this looks very unrepeatable what is

going on why do I have this difference

between this first and second heating as

I'm going through this first insight the

first and second heating my material is

physically changing and physically

adapting to the DSC pan as I'm heating

it so

if I have a second a third fourth fifth

heating I'm going to see a nice overlap

between those those subsequent heating's

and Cooling's whereas on that first

heating maybe just the way that I've

prepared the sample means that it's not

conformed to the DSC pan in an ideal

fashion or it simply rearranged itself

you know via gravity and this transform

it transition so I'm going this is not a

solid liquid transition this is simply a

solid solid transition and then a solid

solid transition back to the previous

phase so again enantiotopic this is

completely reversible and it's happening

we're having a thermodynamic process

this in this particular one and then I'm

sorry that was a miss that was me miss

speaking so we have both of these are

actually kinetic kinetically driven

processes we talked about an anti tropic

and those were those were situations

where we had two compounds that crossed

at some transition point if we look at

mono tropic polymorphic transitions

these ones are irreversible and so what

this means is that only one of these

solid state transformation

transformations is going to be observed

so if we begin with the less stable form

once we trend once we convert it into a

more stable form that's the only

transition that we're going to see so if

you have these polymorphs alpha and beta

of a particular material so a below a

given transition temperature the pink

form is more stable but above a given

transition temperature the form B is

formed and then we never actually go

back we never actually convert back into

the to the to the modification B to this

to this beta form so once we've

processed the material into the alpha

form it stays in that alpha form as

opposed to the N anti-entropic situation

where we have multiple transitions where

we can go back and forth between a and B

in these and a numeric situations with

these monitor Opik transitions it's a

little simpler so instead of five or six

examples we now just have three and we

will start with the most same the most

simple example which is this number two

we've got stable form a

simply melting this is the example that

we saw at the very beginning when we

were looking at something like sorbitol

or the ribavirin we have one stable form

that is melting and we have that

crystalline structure melting and and we

go from solid to liquid what we could

however have is that metastable form b

that is less stable at all temperature

at all temperatures than a transitioning

and wheres if we see this exotherm so we

go from a less stable to a more stable

form we will see that exothermic event

so we're transitioning to form a and

then form a melts in a similar fashion

to a form a melting here number two the

last example that we can see here is the

metastable form b doesn't convert to a

but but for a particular compound what

might happen is the meta stable form

melts and through the melts we then have

the freedom within the melt within that

molten structure that the stable form a

can form and then form a melts and so

you might wonder so I have a neck I have

an endotherm I have an exotherm was

there this overlap of these endothermal

and exothermic events so b melting is

going to absorb energy any any solid to

liquid transition that we're going to be

looking at here is going to involve heat

put into the system but then the

rearrangement of the molecules into a

stable form a into a more

thermodynamically stable arrangement

gives off heat so I go from adding heat

to the system to the system and that

sample then giving off heat and then

keeping again put into the system

through that melt so along with DSC

there are a number of different ways

that we can look at polymorphs x-ray

diffraction is a really popular

technique you can look at powders or

single crystal forms if you're able to

crystallize your material down to a

single crystal we can look at at

infrared spectroscopy or Raman

spectroscopy and in all these situations

we have you know non-destructive

techniques in which we are using a form

of you know a portion of the

electromagnetic spectrum

and then specialized detectors to

directly look at either the crystal

lattice or vibrate or vibrational

effects that are going on we can also

use thermal microscopy so using

high-powered microscopes and just like

we can do the same thing with with

thermal XRD to take our material through

different phase transitions or different

solid solid transitions and then look at

the look at the material structure

directly so if we have these other

techniques why use DSC well DSC is

something which we already have in the

lab for a lot of other purposes for

looking at looking at purity studies

looking at reaction kinetics and it's

also something that's very very easy to

handle sample prep is very easy and the

experiments can be carried out very very

quickly so these different complementary

techniques can be used and you might

find them all within your lab but DSC

can provide us as we've seen a lot of

this a lot of different information so

those were those were us building up all

this information all this knowledge

about our about our our systems let's go

through and look at some some examples

like I said we've got three examples

here to close out the webinar portion

and then we can then go through and look

at questions at a Q&A session at the

very end so what we have here as one

example is that paracetamol we've got

three different literature values so

we've got our three different forms form

one two and three as we can see we've

got form one this is following the

nomenclature that we would ideally like

to see for for most of our compounds

where the form one has the highest

melting temperature and then going down

to form two lower melting temperature

and even lower in some of these

situations we don't have a set heat of

fusion that can be found if we look up

literature values often what we'll see

is kilojoules per mole what we do then

if we want to look at this in terms of

the DSC and the way that we've prepped

our sample we can see the amount of

joules per gram so the area under the

curve of melting for these two different

for this these two different forms so

let's take some paracetamol let's heat

it up so again using a very small amount

of sample 2 milligrams heating it a

relatively fast heating

1010 Kelvin per minute is it I say fast

because one of the things that I'm used

to doing is looking at purity studies

and in purity studies were often using

0.7 to 1 degree Kelvin per minute so


polymorph studies we can do much much

faster so we're using aluminium

crucibles again with a pierced lid in

this case so they're open to the

environment and what do we see we see an

onset temperature of 160 9.2 for the

onset well if we go back to our table

this shows us a very nice very clear

indication that we have form one I don't

see any effects happening at 156 degrees

Celsius so I know I see no effects here

in this region so what this tells me

very clearly from the DSC is that I have

this stable form one so let's heat the

material I've got that first heating I

then cool the material down and then I

heat the material a second time this is

a situation where I now have an entirely

different event so if we if we think

back to the the N and nto tropic

examples we had five different examples

we could have stable form a metastable

form b form b so what we have here is is

we have one form we have an n we have an

endo sorry an exothermic event right

here we have an exothermic event which

means a recrystallization or a

structural transformation we're going

from a less stable to a more stable


we're always going to know that's

happening when we have this exothermic

event and then we have another melting

this time at 150 6.9 degrees Celsius

this then very very nicely overlays with

that form - so based on the thermal

processing on the thermal history of the

material we can very clearly see as

received we have the form one after

processing it in the finished fashion

that we have heating it up at 10k per

minute cooling it down and reheating it

we can then arrive at form at the at

this B form at this form

- rather let's take another example we

have this second compound that we're

going to be looking at which is going to

be a little bit harder to pronounce if

you want to look at the actual structure

here this carbon metazine this one has

it's not only a little bit harder to

pronounce but also has a lot more

different forms and you can see this


we've got form 1 or alpha we've got form

a 2 B 2 3 or B 4 so a lot of different

structures we've got five different

structures that are reported in the

literature with a range of different

melting temperatures and we can find

literature values for also for the heat

of fusion or the the melting enthalpy

for these for three of these different

forms so even on the very first heating

of this material as received we can see

that there's one endothermic event with

a peak temperature of 160 degrees and

then we can see a second endothermic

effect event with this onset of about

190 degrees Celsius again small sample

10k per minute heating rate pierced lid

this is gonna be critical one thing that

we can look at so we've got the first

heating we have a nice stable baseline

all the way through until we get to

these events on a second heating after

cooling with a very fast cooling rate of

40k per minute what we can see here in

the very beginning is we have a step

change we have a step change in the the

specific heat of the material as you

might remember this is a glass

transition what types of materials have

glass transitions amorphous materials so

this what this indicates is that by

cooling the material very quickly we

have kinetically hindered formation of

crystal lattice at least in a portion of

the material and so we have an amorphous

structure that goes through this glass

transition we then have this endothermal

event or sorry EXO thermal event a

little bit off this morning we have this

exit thermal event which if you recall

and we'll say it again

any of these excess thermal events are

us finding it more thermodynamically

stable form we're going from a less

stable form to

more stable form or from an amorphous to

a crystalline of a more highly ordered

structure I then have a melting with an

onset of 186 point four degrees I can go

back and look at my table and compare

and see alright which form do I have

here in this state if I look at the same

material and instead of cooling with a

very fast cooling rate of 40 degrees

Kelvin per minute I heat my heat that

same material and I cool at 10 degrees

Kelvin per minute I have a very

different situation

so first heating this is what we saw in

the last one

now I cool down on my second heating I

no longer have that glass transition I

no longer have an exothermic

recrystallization I simply have that

same melting that I'm that I saw in the

lasts in the last curve so what's going

on its kinetics I mentioned that kinetic

hindrance can play a large part in the

way that we we have different polymorphs

in which forms exist when we cooled

really quickly we froze the material in

its amorphous state when we cooled more

slowly we allowed the material time to

find a more thermodynamically stable

form during the cooling process how is

this relevant for you as as

pharmaceuticals you know scientists or

engineers this is going to dictate the

way that we process our material is is

going to then lead to which forms are

present in our material and when we're

finished with it also looking that's at

shelf-life looking at how do we store

these materials if I need form a or form

B for bioavailability or for processing

how do I need to store these materials

so that I have the form that is desired

let's go ahead and look at another very

very common drug this is a sulfathiazole

this one has four forms reported in

literature for different melting melting

temperatures different heats of fusion

one thing that I can say is that all of

these webinars in this series will then

be posted on our website you can go back

through and download these we look

through this material at your leisure

and so I'm not going to be coming back

to these tables

I'm just gonna keep on plowing forward

through these so the sulfathiazole as

received as we're looking at this

material undergoes to two of these

endothermic events

remember if we look back to the Berger

rules if we're looking at these

endothermic events we're going from it

we're going we can look back and see

that we have a more stable going through

into a metastable form and then we're

actually getting a complete melting

right here we can also see a slight

transition we can see some effect that's

happening right here right before this

onset and this this is something that we

need to look at this could either be my

material conforming to the pan but if

this is a repeatable event this is

something that I look and say this

wasn't just my sample preparation what

we have here is is we might actually

have multiple different forms in the

material as received so I could have a

mixture of forms 1 2 3 & 4 in this case

I only have 3 forms I might have forms 1

2 3 1 3 & 4 however the case might be

but as we're heating and depending on

the heating rate we could also see

transitions from from more stable forms

or sorts from less stable forms to more

stable forms as we reach this final

melting one last example we'll go

through and look at and this is we

talked about polymorphism the last thing

I want to look at is what we talked

about in pseudo polymorphism so this is

this is something that we're going to

look at this is estradiol Hemi hydrate

and you see this second portion of the

of the of the name this Hemi hydrate

Hemi hydrate meaning that we have in the

crystal structure for every molecule of

of this pharmaceutical compound we have

half a molecule of of water of course we

don't have half molecules of water what

this means that is in bulk we have a

ratio of 2 to 1 pharmaceutical compound

to water in the crystal structure so two

simple forms form one form two let's

take this material and let's heat it up

as we've done with all of our other


and this is something that we haven't

seen before in the other examples now

instead of nicely nice very clearly

defined peaks with with nice peak shapes

now along with this melting peak we also

see this large broad endotherm ranging

from about 80 degrees Celsius all the

way up to about 150 degrees Celsius this

is where it's going to be critical for

us to look at the way that we process

our samples so we've got a couple

different samples we ran we ran the

samples in a couple different conditions

so we've got you know anywhere from 1 to

5 milligrams same heating rate we ran

these in aluminum and alumina so

aluminum oxide so metal and ceramic

crucibles under nitrogen atmospheres

wanted to see all right what exactly is

going on here

sometimes we need to look at these

samples from a slightly different

perspective when you look to the

material it's a Hemi hydrate hydrate

means there's water water in a lot of

cases is going to be available for

evaporation it's not going to stay with

the material throughout its whole

throughout the whole process so as we

heat up our material let's look at a

thermo gravimetric curve so if I look at

the mass of the sample as a function of

temperature as I have this endotherm

we've been looking at these solid solid

transitions so long you might be in the

mode to think this is a solid solid

transition when in fact what we have is

a solid is a solid to gas transition so

this might be solid going to liquid

liquid liquid converting to gas so that

hydrogen that Hemi hydrate that water

that's bound in the crystal lattice is

being released from the crystal lattice

and evaporating you can see very clearly

we have a very nice overlap between the

endotherm on my DSC trace and the mass

loss that we're seeing on the same

material run in this alumina crucible on

the TGA so for the DSC I ran a slightly

smaller amount on the TGA just to make

sure I got a little bit nicer signal

I ran five milligrams of this same

heating rate that way I could overlay

very nicely so what we're having is the

loss of water followed by the melting of

this crystalline structure

this is where the way that we run our

material is very very critical so if I

run my material and an aluminum crucible

with a pierced lid that is open to the

environment the water can evaporate I

have what we just saw we have the

evaporation followed by crystallization

if I run this in a closed door aluminum

crucible in which case the water has

nowhere to go that it's retained it's

retained there within the crystal

lattice then I have a more broad melting

peak influenced by the water that's

that's present so I no longer have this

nicely pure estradiol I have the

estradiol Hema hydrate which as we saw

in the last in the last webinar within

this series we saw purity determinations

we saw that solvation effects can

decrease that onset temperature and this

is this is exactly what we see going on

here very clearly so with with net

instruments we have two primary DSC

instruments that are designed for the

analysis of pharmaceutical instruments

we have the DSC two one for nevio and we

have the DSC 204 f 1 Nephi oh we can see

both of these are equipped with an

automatic sample changer one has a nice

magazine for 20 samples for

high-throughput labs or labs that are

operating both R&D and QC or that have

different groups we have this much

larger auto sampler for up to 192

samples that have these 96-well plates

accessories with a sample loader that is

barcode barcode controlled so that you

can go through and load your your

materials in if another researcher needs

to come in and load other samples they

can load theirs come back and reinsert

your samples and your your analysis will

continue on these differ slightly in

their temperature ranges but primarily

they differ in the number of samples

that can be loaded and the automation

that is that we're capable of so in

summary we look the polymorphs we looked

at polymorphs via differential scaling

scanning calorimetry and we used the

properties that the thermo physical

properties of these polymorphs there's


stability the kinetics of cooling to

look at different forms we saw that

polymorph site can have different forms

that are more or less thermodynamically

stable some forms which are metastable

so that they we can we can have

materials that are in general less

stable that are converting into more

stable compounds some that go through

transitions from stable to metastable

and back through two separate forms

before melting one of the reasons we

want to use DSC as a as a

characterization tool in combination

with some of these other techniques is

because the results are very fast and

these are these are quite fast results

instruments are very easy to operate

they're able to be automated and as we

saw in the very end for some some

materials that might not be pure they

might have solvents attached to them

looking at DSC as well as TGA or thermo

gravimetric analysis we can get a better

picture of what's going on within our

materials and one of the fantastic

things here at Natchez we also offer the

combined DSC and TGA so that both of

these at all times can be carried out

simultaneously as you know this is part

two of a four-part webinar series we

look forward to having you back with us

next month you know in October so the

9th of October of this year we're gonna

be looking at thermal stability and

shelf life of pharmaceuticals and we'll

be doing that with infrared spectroscopy

coupled to thermo gravimetric analysis

so we'll be doing we'll be looking at

kinetics of degradation and we'll be

doing some other studies using TGA

coupled to FTIR and then the final

portion we'll be looking at what many of

you might want for your laboratories

which is is fully automated routines so

basically starting with loading a

material into my sample crucible how do

I go from a simple sample prep all the

way to a finished report that just needs

approval via 21 CFR part 11 compliant

software so that's going to be the rest

of the remainder of our series and that

concludes the webinar portion of this

series now I'm going to go through and

answer a few questions that we have so I

first off again thank you for being part

of this series thank you for the


that you have submitted and I will go

through and pull up those on the side

here and we'll get through to your

questions all right so first question

has to do with this first question is

from Steve in Illinois if question has

to do with the conventions the the DSC

traces why am I seeing D SH you know

endotherms up versus down Steve maybe

you missed the beginning of the webinar

this is just a convention this is in

many cases in the u.s. endotherms are

displayed and in doubt down words

whereas European Convention is

endotherms are displayed upwards this is

simply convention I have another

question an anonymous question this one

was on the TGA curve how come the TGA

was TGA curve was run in an Illumina

crucible versus the aluminum crucible

that was used for this for this Hemi

hydrate compound we could have run the

material and we could have run this in

an aluminum crucible that wouldn't have

been a problem this was just simply a

choice of what we used in the laboratory

in most cases for for selection of

crucibles what we want to look at is to

ensure that we have no interaction

between the crucible or pan material

with the pharmaceuticals or the

excipient Saur whatever we're were

analyzing the first consideration is non

interaction for a lot of pharmaceutical

ingredients this means that running in

aluminum is is ideal because aluminum

pans are relatively inexpensive and

disposable they can be pierced they can

be sealed and most of these transitions

actually I can say all of these

transitions that we're looking at are

occurring below the melting point of

aluminum at about 600 a little over 600

degrees Celsius so aluminum pans are

ideal for almost all of your

pharmaceutical applications needs lastly

if we're going to be doing purity

studies or we want to ensure that we

have highly clean crucibles if your

protocols or your methods dictate this

crucibles can be solvent cleaned using

acetone ethanol something that's 100%

purity or ACS grade pharma grade

solvents and then they can be baked so

we can ensure that we have we can we can

heat these up and we can handle them

appropriately to make sure that we don't

have any oils we don't have any residual

solvents this might be a little bit more

tricky looking at ceramic crucibles

ceramic crucibles also simply presents a

different heat transfer property

aluminum crucibles being metal have

extremely high heat transfer and so

we're going to have very very high

fidelity and a high response rate in our

DSC using aluminum pans versus I'm using

say the the ceramic crucibles that we

saw in the TGA on the TGA since we're

simply looking at mass loss and not heat

transfer per se we're not looking at

heat flow the simple mass loss of the

water is perfectly fine so really this

was a user consideration and that's it

we actually only had two questions

during today's webinar again thank you

if you have other questions that you

would like to submit that you maybe were

a little too shy to submit directly

through here please feel free to add

those in now any questions that you

submit after the fact will be addressed

by myself or by my colleagues over in

Germany again very last time thank you

so much for joining us we look forward

to having you next month for next

month's webinar and have a great rest of

your week

bye now