冻干显微镜下的冻干工艺-英国凯文.沃德博士课件.ppt

Formulation Design and Characterisation for Successful Freeze-Drying Cycle Development Dr. Kevin Ward, MRSCDr. Kevin Ward, MRSC凯文凯文凯文凯文. .沃德博士沃德博士沃德博士沃德博士 Director of R&D, Biopharma Technology Ltd., Director of R&D, Biopharma Technology Ltd., Winchester UKWinchester UK英国英国BiopharmaBiopharma 技术有限责任公司研发总监技术有限责任公司研发总监 Chair of Pharmaceutical & Healthcare Sciences Chair of Pharmaceutical & Healthcare Sciences Society Freeze-Drying Special Interest GroupSociety Freeze-Drying Special Interest Group冻干显微镜下的冻干工艺 1 1Synopsis of Presentationn nSome general rules for formulation development… and a few very basic rules for cycle developmentn nMethods of formulation characterisation– –Freeze-Drying Microscopy (FDM)Freeze-Drying Microscopy (FDM)– –DTA / electrical impedance analysis (DTA / electrical impedance analysis (ZsinZsinφ φ) )n nResidual moisture analysis in the lyophilised product2 2The Ideal Formulation?n nIn some respects, the active ingredient / In some respects, the active ingredient / material alone is the best formulation:material alone is the best formulation:– –Lowest solute density (therefore, lower resistance Lowest solute density (therefore, lower resistance to vapour flow)to vapour flow)– –No excipient (No excipient (in)compatibilityin)compatibility issues issues– –Lower cost of manufactureLower cost of manufacturen nHowever, the active ingredient may need to be However, the active ingredient may need to be stabilised prior to and during freeze-dryingstabilised prior to and during freeze-drying3 3Formulation Issues (1)It should be remembered that:n nFREEZING involves CONCENTRATIONn n“Freezing is in itself a form of dehydration” (Felix Franks)n nDrying may risk the removal of water involved in maintaining the structure of the API (especially for proteins)4 4Formulation Issues (2)n nLow volume of higher concentration should freeze-dry faster than larger volume of a lower concentrationn nMany proteins lose activity when freeze-dried from low concentrationsn nHowever, some materials (especially organisms) are difficult or impossible to concentrate them without damage5 5Formulation Issues (3)n nAdditionally, some products are not stable in Additionally, some products are not stable in solution, thereby requiring pH buffering solution, thereby requiring pH buffering and/or other stabilisation and/or other stabilisation even before even before freeze-drying startsfreeze-drying startsn nHowever, remember that pH buffers are However, remember that pH buffers are designed to work in solution – there are no designed to work in solution – there are no guarantees for the frozen or dried state!guarantees for the frozen or dried state!6 6API characteristicsn nCrystalline or amorphous?n nTeu / Tg’ / Tcn nBulk characteristics when freeze-driedn nSolubilityn nConcentration required prior to FDn npH-stability plotn nIEP & aggregation issues for proteins7 7What are we formulating to prevent for the API?n nDestabilisation in liquid staten nDamage by the freezing processn nLoss of activity during dryingn nDegradation during storagen nTherefore, need excipients that are chemically compatible with the API during all the above stages8 8Formulation for freeze-drying involves the use of excipients to…n nprovide provide mechanical strengthmechanical strength (bulk) (bulk)n nafford afford thermal stabilitythermal stability (a high (a high T Tcriticalcritical) during ) during lyophilisation lyophilisation andand in the dried product in the dried productn nprotectprotect the active ingredient(s) from damage the active ingredient(s) from damage before, during and after processingbefore, during and after processingn ngive correct give correct pHpH, and , and tonicitytonicity where required where required (sometimes achieved by reconstituting medium rather (sometimes achieved by reconstituting medium rather than starting solution)than starting solution)9 9Common Excipients: pros & cons*fulfil the basic requirement of remaining amorphous but protective ability depends on API*fulfil the basic requirement of remaining amorphous but protective ability depends on API**PEG often provides **PEG often provides cryocryoprotectionprotection but not necessarily but not necessarily lyolyoprotectionprotection as it can crystallise as it can crystalliseExcipientExcipientBulkBulkThermalThermalProtectionProtectionMannitol (when crystallised)Mannitol (when crystallised)GoodGoodGoodGoodPoorPoorDisaccharides: SucroseDisaccharides: SucroseGoodGoodOKOKGood*Good* Lactose LactoseGoodGoodOKOKGood*Good* Trehalose TrehaloseGoodGoodOKOKGood*Good* Maltose MaltoseGoodGoodOKOKGood*Good*GlucoseGlucosePoorPoorPoorPoorGood*Good*DextranDextranGoodGoodGoodGood?*?*PVPPVPGoodGoodGoodGood?*?*PEGPEGGoodGoodGoodGood?**?**BSA / HSABSA / HSAGoodGoodGoodGood?*?*Amino acids / dipeptidesAmino acids / dipeptidesVariableVariableVariableVariableSome good*Some good*1010“Lyo-friendly” buffersn nCitrateCitraten nTrisTrisn nGlycine / Glycine / HistidineHistidinen nPhosphate often best avoided due to pH Phosphate often best avoided due to pH shifts on freezing, resulting from shifts on freezing, resulting from didi-sodium -sodium salt crystallising outsalt crystallising outn nOther buffers such as acetate, HEPES, borate, Other buffers such as acetate, HEPES, borate, phthalate, are less well studied for freeze-phthalate, are less well studied for freeze-drying but may be suitabledrying but may be suitable1111Other issues in formulating for freeze-dryingn nMixing amorphous and “crystallising” components together:–Phase separation (ice + glass + crystals)Phase separation (ice + glass + crystals)–Unpredictable “critical temperature”Unpredictable “critical temperature”–Possible microcollapse / micromeltingPossible microcollapse / micromelting–Inhibition of crystallisationInhibition of crystallisation–Resulting Resulting metastablemetastable components could components could change over time in dry statechange over time in dry state1212“Extrascientific” issues affecting excipient selectionn nEthical acceptability in target marketn nPrevious acceptance by regulatory bodies (FDA, MHRA etc.) for each mode of use ( (e.g. in-vitroe.g. in-vitro, PO, SC, ID, IP, IM, IV), PO, SC, ID, IP, IM, IV)n nGrade of purity availablen nCostn nSupply chain reliability1313Vials of freeze-dried productGoodOKPoorPoorThe product in the “Poor” vials has become soft and dense during freeze-drying, because it has become warmer than its “Critical Temperature”!1414“What is the Critical Temperature for our product?”n nThe “Critical Temperature” will be:The “Critical Temperature” will be:– –The The eutectic temperature (eutectic temperature (TeuTeu) ) for for crystalline materialscrystalline materials– –The The collapse temperature (collapse temperature (TcTc) ) for for amorphous materialsamorphous materials (somewhere at or above the glass transition temperature)(somewhere at or above the glass transition temperature)– –The The lowerlower of the above temperatures for of the above temperatures for mixed systemsmixed systems (depending on whether micro-collapse is acceptable)(depending on whether micro-collapse is acceptable)n nWe can analyse the critical temperature of a We can analyse the critical temperature of a formulation formulation before freeze-drying it,before freeze-drying it, for example using: for example using:– –Freeze-Drying Microscopy (FDM)Freeze-Drying Microscopy (FDM)– –Impedance (ZsinImpedance (Zsinφ φ) and Thermal Analysis) and Thermal Analysis1515Freeze-drying microscopy (FDM)FDM is the study of FDM is the study of freeze-drying at freeze-drying at the microscopic the microscopic levellevelFDM allows FDM allows determination of determination of collapse, melting collapse, melting and “qualitative and “qualitative phenomena” such phenomena” such as skin formationas skin formation1616What is a Freeze-Drying Microscope?n nEffectively a ‘micro Effectively a ‘micro freeze-dryer’ where freeze-dryer’ where the freeze-drying of a the freeze-drying of a small sample may be small sample may be observedobservedn nFirst designs in the First designs in the mid- 1960smid- 1960sn nNow manufactured Now manufactured commerciallycommercially1717Sample Preparation for FDMSample holderSide DoorBlock•Sample loading takes about 60 seconds.•Routine analysis usually takes 30 – 90 minutes1818Sample Format in Lyostat2Temperature-Controlled BlockLight Source (from below)ApertureQuartz cover slip (16 mm dia.)Glass cover slip (13 mm dia.)2µl of sampleObjective Lens (usually 10 x)Metal Spacer (70µm thick)1919n nIdeally the raw formulation is usedIdeally the raw formulation is usedn nSometimes necessary to use samples that Sometimes necessary to use samples that have previously been frozen or lyophilisedhave previously been frozen or lyophilisedn nAfter loading the sample, the After loading the sample, the Lyostat2Lyostat2 is is set to cool to the desired temperatureset to cool to the desired temperaturen nThe sample is allowed to cool and freeze The sample is allowed to cool and freeze (Note: for eutectic materials, there will be more (Note: for eutectic materials, there will be more than one freezing event!)than one freezing event!)Sample Loading and Cooling2020n nWhen sample reaches When sample reaches the holding temperature the holding temperature and has been observed and has been observed to freezeto freeze, vacuum , vacuum pump is switched on pump is switched on and drying begins.and drying begins.n nSublimation interface Sublimation interface can be seen moving can be seen moving through the frozen through the frozen sample.sample.Frozen sampleDried sampleSublimation frontOn-line plotTemp/time tableINITIAL FDM IMAGE2121n nIncreasing or decreasing Increasing or decreasing the temperature of the the temperature of the sample allows you to sample allows you to view its freeze-drying view its freeze-drying characteristics.characteristics.n nBy examining the freeze-By examining the freeze-dried structure behind dried structure behind the interface, the the interface, the collapse temperature of collapse temperature of the material can be the material can be determined.determined.n nThe temperature may be The temperature may be cycled in order to cycled in order to evaluate Tc more closelyevaluate Tc more closelyFrozen sampleSublimation frontCollapsed materialINTERPRETATION OF EVENTS2222n nSample structure lost Sample structure lost when collapse when collapse temperature was temperature was exceeded.exceeded.n nStructure regained as Structure regained as sample was re-cooled sample was re-cooled to below its collapse to below its collapse temperature.temperature.Frozen sampleCollapsed sampleRegained structure Sublimation frontINTERPRETATION OF EVENTS2323n n100% structure has 100% structure has been regained by been regained by lowering the sample lowering the sample temperature.temperature.n nSample temperature Sample temperature was again increased to was again increased to above its collapse above its collapse temperature, causing temperature, causing the sample to collapse. the sample to collapse. Dried sample with structureCollapsing again on reheatingFrozen sampleSublimation frontINTERPRETATION OF EVENTS2424‘Micro-collapse’ (see (see e.g.e.g. Wang, 2004) Wang, 2004)Below Tc of amorphous phaseAbove Tc of amorphous phaseA similar effect may also be observed due to the melting of crystalline component(s) onto a rigid amorphous structure (depending on which has the lower critical temperature)Macroscopically similar but is it:Wetter?Less stable?More difficult to reconstitute?2525FDM image of an aqueous solution of 2% Mannitol + 1% Glucose-41oC, around Tc for glucose. Possible evidence of visible micro-collapse(Drying front)Frozen materialRegions with good dried structure. Just mannitol?Regions of (micro) collapse. Just glucose?2626So, what else can FDM tell us?n nEutectic melting temperature2727NaCl Below Eutectic TemperatureFrozenDry2828NaCl Above Eutectic TemperatureNote changes in appearance of frozen structureEutectic liquid2929So, what else can FDM tell us?n nEutectic melting temperaturen nMay give some indication of skin (crust) formation potential of a formulation3030Layer of concentrated solute at edge of sampleCrust formation (1)3131Crust Formation (2)Drying only occurs through breaks in the crust3232So, what else can FDM tell us?n nEutectic melting temperatureEutectic melting temperaturen nMay give some indication of skin (crust) May give some indication of skin (crust) formation potential of a formulationformation potential of a formulationn nWhether heat-annealing may be of benefitWhether heat-annealing may be of benefit– –To increase ice crystal size – and what To increase ice crystal size – and what conditions are required for this (above Tg’?)conditions are required for this (above Tg’?)– –To encourage some components to crystalliseTo encourage some components to crystallise3333Effect of annealing on ice crystal sizeSample cooled to -40°C, then warmed to -10°CSame sample after a further 15 minutes at -10°CExperiments can be carried out to compare rates of change at different temperatures, in order to establish what annealing temperature might be most efficient to use in the freeze-dryer.3434FDM setup with polarised lightPolariserAnalyserSampleCamera3535Effect of annealing on solute behaviour: FDM with polarised light functionSample quench cooled below -40°CNo sign of crystals (no light rotation)Same Sample now drying at -18°CPolariser shows presence of crystals (white areas)3636Further applications of FDMn nIt is possible to examine differences in relative drying rates:–For different formulationsFor different formulations–For a specific formulation at different For a specific formulation at different temperaturestemperaturesRef: Ref: ZhaiZhai, S., Taylor, R., , S., Taylor, R., SanchesSanches, R. and N.K.H. Slater , R. and N.K.H. Slater (2003). Measurement of Lyophilisation primary (2003). Measurement of Lyophilisation primary drying rates by freeze-drying microscopy. drying rates by freeze-drying microscopy. Chem. Chem. Eng. Sci.Eng. Sci. 5858, 2313-2323 , 2313-2323 3737DTA and Electrical Impedance analysis (Zsinφ) of Frozen Formulations3838Differential Thermal Analysis (DTA)n nEffective yet simple and inexpensive method Effective yet simple and inexpensive method of analysing frozen solutionsof analysing frozen solutionsn nGives Gives exothermicexothermic and and endothermicendothermic events, events, which can indicate:which can indicate:– –Glass transitionsGlass transitions (amorphous) (amorphous)– –Eutectic meltsEutectic melts (crystalline) (crystalline)– –CrystallisationsCrystallisations (amorphous to crystalline) (amorphous to crystalline)n nAt Biopharma, we use this in combination At Biopharma, we use this in combination with electrical impedance (Zsinwith electrical impedance (Zsinφ φ) ) analysis to analysis to give a more complete picturegive a more complete picture3939Electrical Impedance (Zsinφ) Analysisn nThis is a more sophisticated version of This is a more sophisticated version of electrical resistance (R) analysiselectrical resistance (R) analysisn nImpedance (Z) is a combination ofImpedance (Z) is a combination ofResistance + Inductance + CapacitanceResistance + Inductance + Capacitancen nLooking at Z (or more specifically ZsinLooking at Z (or more specifically Zsinφ φ) ) can give more detailed information about can give more detailed information about frozen solute frozen solute mobilitymobility (Rey, 1999). (Rey, 1999).4040Investigating Zsinφ – Methods (1)n nWe have developed a device (We have developed a device (Lyotherm2Lyotherm2) in ) in collaboration with Prof. Louis Rey, which is capable of collaboration with Prof. Louis Rey, which is capable of analysing Impedance at a frequency of 1000Hzanalysing Impedance at a frequency of 1000Hzn nLyotherm2Lyotherm2 allows both DTA and Impedance (Zsin allows both DTA and Impedance (Zsinφ φ) ) analysis to be carried out on a sample in the frozen analysis to be carried out on a sample in the frozen state – large sample volume to give stronger signalstate – large sample volume to give stronger signal4141Investigating Zsinφ – Methods (2)n nApproximately 100 single- and dual- solute samples Approximately 100 single- and dual- solute samples were analysed in order to validate the device against were analysed in order to validate the device against published datapublished datan nFor many formulations, the onset of mobility (at a For many formulations, the onset of mobility (at a point we defined as “Tpoint we defined as “TZonsetZonset”) was obtained, ”) was obtained, even even where where ModuledModuled Temperature Differential Scanning Temperature Differential Scanning Calorimetry was unable to show a thermal eventCalorimetry was unable to show a thermal eventn nAn applied, in-depth study was then carried out in An applied, in-depth study was then carried out in collaboration with Dr. Paul Matejtschuk of NIBSC (UK) collaboration with Dr. Paul Matejtschuk of NIBSC (UK) in order to establish how Zsinin order to establish how Zsinφ φ analysis might be analysis might be relevant to a further 40 multi-component biological relevant to a further 40 multi-component biological product formulations and placebosproduct formulations and placebos4242Investigating Zsinφ - PracticalZsin impedance probeSymmetry-balanced heating built into sample holding block, to give smooth baselineDTA assemblySample format for Lyotherm2 analysisSamples were cooled to below -100ºC and analysed on rewarming4343Example of Zsinφ + DTA graph4444Examples where onset of mobility increase (at TZonset) observed well below collapse temperatureFormulationFormulation(% values in (% values in w/vw/v) )T TZonset Zonset (ºC)(ºC)( (Lyotherm2 Lyotherm2 ZsinZsinφφ) )Onset Onset TcTc (ºC) (ºC)( (Lyostat2 FDMLyostat2 FDM) )NaCl (0.9%) + HSA (0.5%)NaCl (0.9%) + HSA (0.5%)-46-46-18.4-18.4NaCl (0.9%) + HSA (1.0%)NaCl (0.9%) + HSA (1.0%)-64-64-20.4-20.4NaCl (0.9%) + HSA (5.0%)NaCl (0.9%) + HSA (5.0%)-60-60-23.0-23.0Egg Egg allantoicallantoic fluid (undiluted) fluid (undiluted)-60-60-50-50Potassium phosphate bufferedPotassium phosphate bufferedHSA (0.2%) + trehalose (0.1%)HSA (0.2%) + trehalose (0.1%)-58-58-50-50HSA (0.1%) + casein (0.3%)HSA (0.1%) + casein (0.3%)in PBSin PBS-59-59-50-504545Ongoing studies from these datan nThe real-life practical implications of these data are The real-life practical implications of these data are currently being determined by freeze-drying some of currently being determined by freeze-drying some of the 140 samples we have analysed:the 140 samples we have analysed:– –Below TBelow TZonset Zonset ( (good structure)good structure)– –Between TBetween TZonsetZonset and and T Tcollapsecollapse ( (micro-collapse?)micro-collapse?)– –Above Above T Tcollapsecollapse ( (macroscopic collapse)macroscopic collapse)…and examining their appearance, structure, water …and examining their appearance, structure, water content, crystallinity, stability, reconstitution time.content, crystallinity, stability, reconstitution time.Early indications are that such a correlation does exist, and the extent to which each aspect of the final product is affected depends on the nature of the product4646Conclusions for Impedance Analysisn nImpedance (ZsinImpedance (Zsinφ φ) analysis has provided a value of ) analysis has provided a value of T TZonsetZonset for a number of samples, indicating mobility for a number of samples, indicating mobility even when no thermal event was seeneven when no thermal event was seenn nData indicate that a correlation exists between Data indicate that a correlation exists between product temperature during lyophilisation (in relation product temperature during lyophilisation (in relation to both Tto both TZonsetZonset and and TcTc) and defined aspects of the ) and defined aspects of the lyophilised productlyophilised productn nImpedance analysis may therefore represent a useful Impedance analysis may therefore represent a useful addition to the current range of analytical methods addition to the current range of analytical methods applied prior to freeze-dryingapplied prior to freeze-drying4747Cycle Development Basics (1)n nCool to below the critical temperature of the formulation (monitor product (monitor product temperature at base)temperature at base)n nAssess cooling rate effects (but not by (but not by freeze-thawing, as thawing can be damaging!)freeze-thawing, as thawing can be damaging!)n nTemperature safety margin to allow for variability, measuring device effects, scale-up…–More safety margin required for eutectics?More safety margin required for eutectics?4848Cycle Development Basics (2)n nPartial pressure of vapour should be lower Partial pressure of vapour should be lower than vapour pressure of ice at drying frontthan vapour pressure of ice at drying frontn nNumerous PAT / Endpoint determination Numerous PAT / Endpoint determination methods availablemethods availablen nSecondary drying may need to begin below Secondary drying may need to begin below “Tg at several % water”“Tg at several % water”n nFinal water content should give sufficiently Final water content should give sufficiently high Tg for desired storage temperature & high Tg for desired storage temperature & timetime4949Case StudyProduct Cycle DevelopmentProduct Cycle DevelopmentThey were discarding a high percentage of each batch due to They were discarding a high percentage of each batch due to visible defects occurring in the product during freeze-dryingvisible defects occurring in the product during freeze-dryingA client approached BTL with a product that was being freeze-A client approached BTL with a product that was being freeze-dried using a cycle that was originally developed for another dried using a cycle that was originally developed for another productproduct5050Product Cycle DevelopmentProduct Cycle DevelopmentStep 1 – Information was obtained on the critical temperatures Step 1 – Information was obtained on the critical temperatures and and thermal behaviour of the product using the Lyostat2 and thermal behaviour of the product using the Lyostat2 and Lyotherm2 instrumentsLyotherm2 instrumentsStep 2 – This data confirmed the lack of suitability of the existing Step 2 – This data confirmed the lack of suitability of the existing freeze-drying cyclefreeze-drying cycleStep 3 – Critical temperature information was used to create a Step 3 – Critical temperature information was used to create a “first approximation” cycle tailored to the needs of the “first approximation” cycle tailored to the needs of the productproductStep 4 – Data from this cycle was used to design a more Step 4 – Data from this cycle was used to design a more optimisedoptimisedcycle until a safe and efficient cycle was cycle until a safe and efficient cycle was achieved,achieved,minimising cycle time without jeopardising minimising cycle time without jeopardising product qualityproduct qualityCase Study5151Lyostat2Lyostat2 Freeze-Drying Microscopy Analysis Freeze-Drying Microscopy AnalysisSample dries well at -50.0°C, but collapse starts as the temperature is increased to -45.7°C. This can be identified by defects appearing in the dried materialAs the temperature increases to -39.6°C the structure continues to weaken and collapse becomes more evidentCase Study5252Lyostat2Lyostat2 Freeze-Drying Microscopy Analysis Freeze-Drying Microscopy AnalysisThe sample is repeated but this time with an annealing step – frozen and cooled to -50.0°C, warmed to -15.0°C and re-cooled to -50.0°C before drying. The sample dries with good structure until the temperature reaches -31.4°C and defects appearAt -30.8°C the sample is too weak to maintain any structure as the ice is removedCase Study5353Lyotherm2Lyotherm2 DTA and Impedance Analysis DTA and Impedance Analysis1234See full labels 1 – 4 on next slideCase Study54541.1.ExothermExotherm in DTA and increase in Impedance in DTA and increase in Impedance indicating a stabilisation / rearrangement of the indicating a stabilisation / rearrangement of the frozen structurefrozen structure2.2.Increase in downward gradient of Impedance curve Increase in downward gradient of Impedance curve indicating a softening of the frozen materialindicating a softening of the frozen material3.3.Onset of a sharp endotherm consistent with the melting Onset of a sharp endotherm consistent with the melting of the iceof the ice4.4.Minimum Impedance indicating complete mobility Minimum Impedance indicating complete mobility within the solute structurewithin the solute structureLyotherm2Lyotherm2 DTA and Impedance Analysis DTA and Impedance AnalysisCase Study5555Interpretation of Analysis ResultsInterpretation of Analysis Resultsn nThe inclusion of an annealing step resulted in an increase in The inclusion of an annealing step resulted in an increase in the collapse temperature of the formulation from -45.7°C to -the collapse temperature of the formulation from -45.7°C to -31.4°C, as well as increasing ice crystal size and networking31.4°C, as well as increasing ice crystal size and networkingn nTherefore, the maximum allowable product temperature Therefore, the maximum allowable product temperature during sublimation (to avoid collapse) was raised by 14.3°C during sublimation (to avoid collapse) was raised by 14.3°C by the use of annealing, thereby allowing drying to be by the use of annealing, thereby allowing drying to be carried out at higher temperatures, for a more efficient cycle. carried out at higher temperatures, for a more efficient cycle. The higher the product temperature during drying, the faster The higher the product temperature during drying, the faster the drying rate.the drying rate.From the results of these analyses, we made the From the results of these analyses, we made the following deductions:following deductions:Case Study5656Client’s Existing Cycle – 70 hoursClient’s Existing Cycle – 70 hoursTc = -45.7°C+20°C-15°C-50°C-40°CShelf TemperatureProduct TemperatureChamber Pressure 1231 – Freezing 2 – Primary Drying 3 – Secondary Drying AA – Product at risk of collapseCase Study5757Modified Cycle Created By BTL – 42 hoursModified Cycle Created By BTL – 42 hours+20°C-15°C-50°C-35°CShelf TemperatureProduct TemperatureChamber Pressure 1234Tc = -31.4°C1 – Freezing 2 – Annealing 3 – Primary Drying 4 – Secondary DryingCase Study5858This graph shows an enlarged section of the previous graph+20°C-15°C-50°C-35°CShelf TemperatureProduct TemperatureChamber PressureTc = -31.4°C1 – Freezing 2 – Annealing 3 – Primary Drying 4 – Secondary Drying3The Sublimation Cooling Effect The lowering of product temperature caused by the sublimation of iceCase Study5959The Next StepsThe Next StepsFrom the previous run we now know:1.The extent of sublimation cooling, allowing us to increase the shelf temperature / chamber pressure as high as possible whilst sublimation cooling keeps the product temperature below Tc2.When sublimation was complete in temperature-probed samples (when product temperature = shelf temperature)3.The physical appearance of the cakes produced by the cycle4.Residual moisture was measured in the final product, in order to establish whether the extent of secondary drying was sufficientCase Study6060Problem Solved!FDM and DTA/Impedance analysis provides you with essential FDM and DTA/Impedance analysis provides you with essential information on the critical temperatures of your formulation, information on the critical temperatures of your formulation, enabling you to get the best results from your productenabling you to get the best results from your productDetermination of critical Determination of critical temperaturestemperatures‘Trial and Error’ Cycle ‘Trial and Error’ Cycle resulting in loss of resulting in loss of product or costly product or costly inefficient cycleinefficient cycleNew cycle New cycle based on actual based on actual parametersparameters6161Changes in the productn nFor many APIs, freeze-drying will give a (partially) amorphous productn nNeed to be aware of:–Thermodynamic / Kinetic stabilityThermodynamic / Kinetic stability–Changes in Changes in poly(a)morphspoly(a)morphs and/or and/or hydrates that may affect the ‘location’ of hydrates that may affect the ‘location’ of water over timewater over timen nFMS vs. KF for looking at water…6262Frequency Modulation Spectroscopy (FMS)n nFMS is a method that detects molecules in FMS is a method that detects molecules in the headspace of a sealed container (such the headspace of a sealed container (such as a vial)as a vial)n nBeing non-destructive, it allows Being non-destructive, it allows the same the same vialvial to be analysed at different time points, to be analysed at different time points, thus reducing inherent time point variabilitythus reducing inherent time point variabilityn nThe Lighthouse Instruments FMS-1400 The Lighthouse Instruments FMS-1400 measures vial pressure and headspace measures vial pressure and headspace water at 1400nmwater at 1400nm6363“Where is the water in my product?”GR Scientific ‘GR Scientific ‘CouCou-Lo -Lo AquamaxAquamax’ ’A recent study by BTL showed that using FMS in combination with KF titration may provide evidence of where the water is…Lighthouse ‘FMS-1400’…and whether this changes over time6464Results - comparing KF & FMSn nFMS gives an indication of the “equilibrium FMS gives an indication of the “equilibrium free moisture” in the headspace, while KF free moisture” in the headspace, while KF gives total moisture (assuming sample is gives total moisture (assuming sample is soluble)soluble)n nBy using the 2 methods to analyse a series By using the 2 methods to analyse a series of different dried samples with a range of of different dried samples with a range of moisture levels, we were able to moisture levels, we were able to understand more about relative proportions understand more about relative proportions of ‘free’ water and water that was more of ‘free’ water and water that was more tightly associated with the caketightly associated with the cake6565Comparison of KF & FMS for sucroseDoes Y-axis intercept (1.49%) indicate level of ‘bound’ water? (sorry Felix!)… it corresponds with there being zero pressure of ‘free’ water6666Results - comparing KF & FMSn nData taken at different timepoints and Data taken at different timepoints and temperatures, and using dried vs. non-dried temperatures, and using dried vs. non-dried stoppers, have allowed further understanding stoppers, have allowed further understanding of:of:– –Movement of water into or out of a dried cake Movement of water into or out of a dried cake from headspace (and relationship with from headspace (and relationship with temperature), even if total moisture in vial temperature), even if total moisture in vial remains constantremains constant– –Ingress of water into product / headspace from Ingress of water into product / headspace from stopperstopper– –Change in crystals / hydrates / Change in crystals / hydrates / poly(a)morphspoly(a)morphs6767In Summary… (1)n nKnow your API!Know your API!n nWhat excipients / stabilisers does it need (if What excipients / stabilisers does it need (if any) to prevent:any) to prevent:– –Destabilisation in liquid stateDestabilisation in liquid state– –Damage by the freezing processDamage by the freezing process– –Loss of activity during dryingLoss of activity during drying– –Degradation during storage?Degradation during storage?n nSelect additives that are compatible with the Select additives that are compatible with the API and each otherAPI and each other6868In Summary… (2)n nDevelop a cycle that takes into account the Develop a cycle that takes into account the critical critical temperature(stemperature(s) [e.g. gained using FDM ) [e.g. gained using FDM / Thermal Analysis / Impedance Analysis]/ Thermal Analysis / Impedance Analysis]n nBuild in a safety margin in terms of product Build in a safety margin in terms of product temperature during FDtemperature during FDn nAim for a sufficiently low residual water level Aim for a sufficiently low residual water level to give dry state stabilityto give dry state stabilityn nLooking at water dynamics in the dried Looking at water dynamics in the dried product can help flag up possible issuesproduct can help flag up possible issues6969Thank You forYour attention!biopharma.co.ukFormulation Design and Characterisation for Successful Freeze-Drying Cycle Development 7070。