`production
`
`An engineering guide
`
`Edited by Bill Bennett and Graham Cole
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`E
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`The information in this book is given in good
`faith and belief in its accuracy, but does not
`
`imply the acceptance of any legal liability or
`responsibility whatsoever, by the Institution. or
`
`by the editors, for the consequences of its use or
`misuse in any particular circumstances This
`disclaimer shall have effect only to the extent
`
`permitted by any applicable law.
`
`All rights reserved. No part of this publication
`may be reproduced, stored in a retrieval
`system. or transmitted. in any form or by any
`
`means, electronic. mechanical. photocopying,
`
`recording or otherwise. without the prior
`permission of the publisher.
`
`Published by
`
`Institution of Chemical Engineers {IChernE}
`
`Davis Building
`165—189 Railway Terrace
`
`Rugby. Warwickshire CV2] SHQ, UK
`
`IChemE is a Registered Charity
`
`Offices in Rugby (UK). London {UK} and Melbourne (Australia)
`
`{3} 2003 Institution of Chemical Engineers
`
`ISBN 0 85295 440 9
`
`Typeset by Techsct Composition Limited. Salisbury. UK
`
`Printed by Antony Rowe Limited. Chippenham. UK
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`Contents
`
`Preface ........................................................................
`
`v
`
`List of Acronyms ..........................................................
`
`v
`
`Glossary ...................................................................... xi
`
`1.
`
`Introduction .........................................................
`
`1
`
`9
`
`2. Regulatory Aspects ............................................
`2.1
`Introduction .............................................................
`2.2 Key Stages in Drug Approval Process .................. 10
`2.3 Example of Requirements ..................................... 12
`2.4 Post-Marketing Evaluation ..................................... 1
`2.5 Procedures for Authorizing Medicinal
`Products in the European Union ............................ 14
`2.6 European and US Regulatory Perspectives .......... 14
`
`3. Good Manufacturing Practice ............................ 17
`3.1
`Introduction ............................................................. 1
`3.2 GMP Design Requirements ................................... 22
`3.3 GMP Reviews of Design ........................................ 34
`
`4. Validation ............................................................. 38
`4.1
`Introduction ............................................................. 3
`4.2 Preliminary Activities .............................................. 4
`4.3 Validation Master Planning .................................... 44
`4.4 Development of Qualification Protocols and
`Reports ................................................................... 5
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`4.5 Design Qualification (DQ) ...................................... 53
`4.6
`Installation Qualification (IQ) .................................. 55
`4.7 Operational Qualification (OQ) .............................. 56
`4.8 Handover and Process Optimization ..................... 58
`4.9 Performance Qualification (PQ) ............................. 59
`4.10 Process Validation (PV) ......................................... 60
`4.11 Cleaning Validation ................................................ 61
`4.12 Computer System Validation ................................. 68
`4.13 Analytical Methods Validation ................................ 71
`4.14 Change Control and Revalidation .......................... 71
`
`5. Primary Production ............................................. 75
`5.1 Reaction ................................................................. 75
`5.2 Key Unit Operations ............................................... 85
`5.3 Production Methods and Considerations .............. 96
`5.4 Principles for Layout of Bulk Production
`Facilities .................................................................. 100
`5.5 Good Manufacturing Practice for BPC .................. 109
`
`6. Secondary Pharmaceutical Production ............ 111
`6.1 Products and Processes ........................................ 111
`6.2 Principles of Layout and Building Design .............. 154
`6.3 The Operating Environment ................................... 159
`6.4 Containment Issues ............................................... 176
`6.5 Packaging Operations ............................................ 177
`6.6 Warehousing and Materials Handling .................... 188
`6.7 Automated Production Systems ............................ 190
`6.8 Advanced Packaging Technologies ...................... 192
`
`7. Safety, Health and Environment (SHE) ............. 202
`7.1
`Introduction ............................................................. 202
`7.2 SHE Management .................................................. 202
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`7.3 Systems Approach to SHE .................................... 207
`7.4
`Inherent SHE .......................................................... 209
`7.5 Risk Assessment .................................................... 211
`7.6 Pharmaceutical Industry SHE Hazards ................. 236
`7.7 Safety, Health and Environment Legislation ......... 257
`
`8. Design of Utilities and Services ......................... 260
`8.1
`Introduction ............................................................. 260
`8.2 Objectives ............................................................... 261
`8.3 Current Good Manufacturing Practice ................... 262
`8.4 Design .................................................................... 263
`8.5 Utility and Service System Design ......................... 270
`8.6 Sizing of Systems for Batch Production ................ 287
`8.7 Solids Transfer ....................................................... 289
`8.8 Cleaning Systems .................................................. 289
`8.9 Effluent Treatment and Waste Minimization .......... 291
`8.10 General Engineering Practice Requirements ......... 297
`8.11 Installation .............................................................. 299
`8.12 In-House Versus Contractors ................................ 300
`8.13 Planned and Preventive Maintenance ................... 301
`8.14 The Future? ............................................................ 302
`
`9. Laboratory Design .............................................. 304
`9.1
`Introduction ............................................................. 304
`9.2 Planning a Laboratory ............................................ 307
`9.3 Furniture Design ..................................................... 321
`9.4 Fume Cupboards ................................................... 329
`9.5 Extraction Hoods .................................................... 336
`9.6 Utility Services ........................................................ 337
`9.7 Fume Extraction ..................................................... 337
`9.8 Air Flow Systems ................................................... 340
`9.9 Safety and Containment ........................................ 344
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`10. Process Development Facilities and Pilot
`Plants ................................................................... 346
`10.1 Introduction ............................................................. 346
`10.2 Primary and Secondary Processing ...................... 347
`10.3 Process Development ............................................ 347
`10.4 Small-Scale Pilot Facilities ..................................... 352
`10.5 Chemical Synthesis Pilot Plants ............................ 361
`10.6 Physical Manipulation Pilot Plants ......................... 368
`10.7 Final Formulation, Filling and Packing Pilot
`Plants ...................................................................... 369
`10.8 Safety, Health and Environmental Reviews .......... 371
`10.9 Dispensaries ........................................................... 371
`10.10 Optimization ........................................................... 371
`10.11 Commissioning and Validation
`Management .......................................................... 371
`
`11. Pilot Manufacturing Facilities for the
`Development and Manufacture of Bio-
`Pharmaceutical Products ................................... 372
`11.1 Introduction ............................................................. 372
`11.2 Regulatory, Design and Operating
`Considerations ....................................................... 373
`11.3 Primary Production ................................................. 388
`11.4 Secondary Production ............................................ 402
`11.5 Design of Facilities and Equipment ....................... 417
`11.6 Process Utilities and Services ................................ 442
`
`Index ........................................................................... 447
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`Primary
`pharmaceutical
`production
`
`ROGER SHILIJI'OE. Pl'lll MASON and FRED SMITH
`
`This chapter considers. the production of the bulk active ingredient or bulk
`pharmaceutical chemical (BPCJ that is subsequently converted by physical
`
`means into the. final drug‘s presentation form.
`This area of the pharmaceutical industry has much in common with fine
`
`chemical manufacture. The unit operations carried out are similar and many
`fine chemical and speciality chemical manufacturers also manufacture phat-
`maceutical intermediates.
`
`Traditional ly, the bulk production was carried out on a different site to the
`R&D and secondary processing. The style of operation. attention to cGMP and
`
`culture of a primary site. was more associated with the type of chemistry or
`operation carried out.
`
`Three main influences are changing the face of the BPC‘ industry:
`
`0 regulators, particularly the FDA. are putting greater emphasis on reviewing
`
`BPC production. and recognize the effect that failure in quality can have on
`the finished dosage form;
`
`a major pharmaceutical companies are focusing on ‘Research and Develop-
`ment’ and ‘Marketing and Selling of the finished product’. Secondary
`
`manufacture to a limited extent. and primary or BPC manufacture to a
`greater extent. is being sub-contracted out to third parties:
`0 BPCs are becoming more active and tonnage requirements are dropping as a
`
`result. Linked with this. the size of the equipment used in the manufacture is
`
`reducing. The increased activity also brings increased handling considera-
`tions and limits for exposure, which in turn drives towards closed processing
`
`operations. which is also consistent with improvements to cGMP.
`
`5.] Reaction
`
`The production of the RFC is by three main methods:
`
`0 chemical synthesis: Examples of synthetic conversions include aSpirin.
`
`diazepam. ibuprofen. This method produces the largest tonnage;
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`PHARMACEUTICAL PRODUCTION: AN ENGINEERING GUIDE
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`0 biotechnology or microbial action: Examples include antibiotics. vaccine
`
`production, blood plasma products. This method produces the high value
`products;
`a extraction: This can be by extraction of natural materials from animal or
`
`plant material such as the opium alkaloids. dioxin, heparin, insulin (pigs
`
`pancreas), thyroxine (animal thyroid gland).
`
`This chapter will concentrate on the first two methods. The extraction
`method for naturally occurring materials was the main source of drugs up to the
`19305 but was being gradually replaced with synthetic routes to products.
`
`There is resurgence now in extraction techniques linked to the biotechnology
`
`area, where specifically developed or altered organisms are allowed to grow and
`produce a desired product that is harvested and extracted. This is discussed in
`Section 5.1.2.
`
`5.l.'.l Synthetic chemistry based processes
`Various general synthetic chemical reactions are utilized in the synthesis of
`
`BPCs. These include simple liquid/ liquid reactions, complex liquid reactions
`with catalysis such as Grinards, Freidel Craft, reaction with strong reagents
`
`such as phosphorous oxychloride, thionyl chloride or elemental halogens such
`as bromine or chlorine. Gas reactions with liquids are common for example
`
`with hydrogen. hydrogen chloride or phosgene.
`Most reactions in the pharmaceutical industry are carried out on a batch
`
`basis. in non steady state operation. Continuous processing is occasionally used
`for a few generic tonnage commodity BPCs or where safety can be improved by
`
`the benefits continuous processing can bring by inventory minimization.
`
`Conventional batch reactor systems
`
`The batch reactor is the workhorse of the synthetic BPC industry. Typically
`
`made from stainless steel or glass lined mild steel, capacities ranges from 500
`litres at the small scale to 16 m3 at the large scale. Some processes employ
`reactors of even greater capacity but this is becoming unusual as the activity of
`new drug substances increases.
`
`The reactor is typically fitted with an external jacket or half pipe coils so that
`
`the temperature ofthe contents can be adjusted. Occasionally if a high heat duty
`is required, fiirther coils can be placed inside the reactor.
`Typical operating conditions are from —25“‘C to + 160°C, and full vacuum
`to 6 bar g. Generally. reactions at elevated pressures above 1 bar g are uncom-
`
`mon. with the exception of specific gas reactions such as hydrogenation.
`
`However. more processes are now being developed where working at an
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`PRIMARY PHARMfiCEU'IICAI PRGDUC'I'ION
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`elevated pressure brings benefits -— for example, it can allow the selection of
`the ideal solvent for a reaction that could not normally be used at the ideal
`
`reaction temperature because this would be above its atmospheric boiling point.
`
`The temperature is normally adjusted by indirect contact with a heating or
`cooling medium circulating through the coil or jacket. but direct heating with
`
`live steam or quench cooling with water or other materials is possible. The
`
`medium used for the heating and cooling fall into two main areas:
`
`I multiple fluids: typically steam, cooling water, refrigerated fluid such as
`ethylene glycol or brine. These are applied in sequence to the coil orjacket as
`
`required;
`
`0 single fluids: typically some form of heat transfer oil, heated or cooled by
`indirect contact with steam, cooling water or refrigerant, and blended to
`provide the. correct fluid to the eoil or jacket.
`
`Agitation is provided to the reactor to ensure good heat transfer and good
`
`mixing for reaction. Depending on the process requirements, various agitation
`regimes can be set up using different agitator profiles, speeds and locations.
`Connections are made to both the top and bottom of the reactor to allow
`
`material to be charged into the reactor, material to be distilled from the reactor,
`
`and liquids to be drained out.
`Reactors are normally fitted with a manway to allow entry for maintenance
`
`purposes. Historically, this was also the way in which solids were added to the
`
`reactor and samples were extracted, but this practice is becoming less common.
`
`Alternative reactor systems
`
`Other types of reactor Systems exist with each having their own specific
`advantages for specific processes. These include the loop reactor that specia-
`
`lizes in gas-liquid reactions at elevated pressures. such as hydrogenation. and
`the batch autoclave reactor that specializes in high-pressure reactions of
`100 bar g and higher.
`
`Materials of construction
`
`Reaction modules can be constructed from other materials dependant on the
`
`chemistry being employed and requirements for heat transfer. These include
`glass, plastics and exotic metals such as hastclloy or titanium.
`
`5.l.2 Biotechnology based processes
`The processes in biotechnology are based on cultivation of micro-organisms,
`
`such as bacteria. yeast, fungi or animal and plant cells. During the microbial
`process the micro-organisms grow the product. which is either contained within
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`PHARMAITEUTiCAL PRODUCTION AN ENGINEERING GUIDE
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`the cell or excreted into the surrounding liquor. The micro-organisms need
`carbon substrate and nutrient medium for growth and the microbial process is
`
`normally performed in water.
`There are essentially three steps to biotechnology processing, namely.“
`
`- fermentation;
`O FCCOVEI'y;
`
`o purification.
`
`The equipment in which the microbial process is carried out is called the
`fermenter and the process in which micro-organisms grow or format product is
`called fermentation.
`
`Once the product is formed it is recovered from the biomass or the liquor by
`
`downstream processing. e.g., centrifugation, homogenization or ultrafiltration.
`
`then required. Two differing
`is
`Purification of the recovered product
`techniques are required depending on whether it is for bulk large-scale or for
`small-scale genetically manipulated organisms. Large-scale recovery can be
`likened to bulk chemical organic synthesis operation.
`
`Fermentation
`
`The fer-mentor
`
`is the equipment used to produce the micro—organisms.
`
`Biotechnology applications of fennentations divide conveniently between
`microbial types and mammalian cell culture. Microbial fennentation, which
`
`can encompass very large-scale antibiotics as well as smaller scale recombinant
`products,
`is characterized by fast growth rates with accompanying heat and
`mass transfer problems. Mammalian cell culture is characterized by low growth
`
`rates and high sensitivity to operating conditions. Both techniques have
`common design principles.
`
`Several different types of vessel are used for large—scale microbiological
`
`processes. and their degree of sophistication in design, construction and
`
`operation is determined by the sensitivity of the process to the environment
`maintained in the vessel.
`
`The following is a brief description of the main types of fennenters:
`
`(a) Open tank
`
`The simplest type of fermenter is an open tank in which the organisms are
`diSpersed into nutrient liquid. These have been used successfully in the brewing
`
`industry. In the anaerobic stage of fermentation, a foam blanket of carbon
`
`dioxide and yeast develops which effectively prevents access of air to the
`process. Cooling coils can be fitted for controlling temperature during fermen-
`ration.
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`E’RIMAKY PHARMACEUTICAL PRUDUCTIUN
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`(b) Stirred tutti:
`
`Stirred-tank fermentets are agitated mechanically to maintain homogeneity. to
`
`attain rapid dispersion and mixing of injected materials. and to enhance heat-
`
`transfer in temperature control and mass—transfer in dissolving sparingly soluble
`gases such as oxygen. The extent to which these are achieved depends mainly
`
`on the povver dissipated into the medium by the agitator. so that the agitator is
`essentially a power transmission device. The efi‘ectiveness of the power input
`
`depends on the configuration of the agitator and other fennenter components.
`For aerobic fermentations. air is injected through a sparger, a single nozzle
`or a perforated tube arrangement, positioned well below the lowest impeller to
`
`avoid swamping it with gas. The sparger should have provision for drainage so
`that no culture medium remains in it after the vessel is discharged.
`
`The rate of air supply must be sufficient to satisfy the oxygen demand of the
`fermentation after allowing for the efficiency of oxygen dissolution achieved.
`
`Instead of a rotating stirrer. some systems obtain the mechanical power input
`
`by using a pump to circulate liquid medium from the fermenter vessel through a
`gas entrainer and then back into the fermenter. This separates the liquid
`movement and gas dissolution functions into separate specialized units. Two
`
`designs have evolved using this principle # the ‘loop’ fenncnter and the “deep
`jet‘ fermenter. In the loop fennentcr, the gas dissolution device is a subsidiary
`vessel into which gas is injected. and the gas-saturated liquid is recirculated to
`
`the main growth stage. In the deep-jet system, gas is entrained into a high-
`
`power jet of liquid injected into the liquid in the fermenter, re-entraining gas
`from the vessel headspaee. Exhaust gas is purged partly from the vessel
`headspace and partly from the specially designed circulation pump. from
`
`which the degassed liquid passes through a supplementary cooler before
`
`passing to the gas entraincr. This system gives high gas dissolution rate, but
`
`has correspondingly high power consumption compared to conventional
`systems. The liquid and entrained gas can also be introduced into the fermenter
`
`through a ‘bell‘. which holds the gas bubbles in contact with the recirculating
`liquid to enhance gas utilization.
`
`(cl Gas-iifi and merged—tankferal-enters
`This design has no mechanical stirrer and the power required for mixing, heat-
`
`transfer and gas dissolution. is provided by the movement of gas through the
`liquid medium. The gas is, therefore. the power transmission system from the
`
`gas compressors into the vessel. While the relatively low efficiency of gas
`
`compression seems to make this design unattractive,
`
`it has some important
`
`the absence of a
`advantages compared to the stirred-tank system. Firstly.
`rotating agitator shaft removes the major contamination risk at its entry point
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`FHARMACEHTICAL PRODUCTION: AN ENGINEERING GUIDE
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`to the vessel. Secondly. for very large vessels, the required power input for
`agitation is just too large to be transmitted by a single agitator. Thirdly, the
`evaporation of water vapour into the gas stream makes a small contribution to
`
`ecoling the fermentation. The fennenter interior does. however, need careful
`
`design to ensure that the movement pattern of the gas through the system
`produces satisfactory agitation.
`
`The various designs of non-mechanically agitated fermenters can be
`
`grouped broadly into sparged vessels and gas-lift (including air-lift} fennenters.
`
`Sparged—tank fennenters are usually of high aspect ratio, with gas introduced at
`the bottom through a single nozzle or a perforated or porous distributor plate.
`The gas bubbles rise through the liquid in the vessel and may be redispersed by
`
`a succession of horizontal perforated baffle-plates sited at intervals up the
`
`column. In the gas-lift fermenters. internal liquid circulation in the vessel is
`achieved by sparging only part of the vessel with gas. The sparged volume has a
`lower effective density than the bubble-tree volume. and the difference in
`
`hydrostatic pressure between the twu sections drives the liquid circulation
`upwards in the sparged section and, after gas disentrainment, downwards in the
`
`bubble-flee section. The two sections may be separated by a vertical draught-
`tube.
`
`Important design considerations for good fermenter operation
`The following are important design consideratioas in fennenter operation:
`
`(a) Aeration and agitation
`
`Animal cells are shear-sensitive (mild agitation is therefore required} and they
`
`are alien sensitive to air bubbles. These considerations imposa significant
`constraints on oxygen transfer design. One way in which this problem has been
`
`addressed is by the use of gas exchange impellers. Another strategy is to
`
`circulate medium through the reactor while simultaneously oxygenating it in an
`external loop. A third method is to use silicon tubing through which air diffuses
`
`into the liquid medium.
`Cell culture medium often contains serum, which has a tendency to cause
`
`foaming. Since dcfoamants may inhibit growth, agitation and aeration systems
`must be designed to minimize this potential problem. However, care must be
`
`taken in the amount of agitation applied because, although it provides good
`oxygen and heat transfer characteristics, it can result in mechanical degradation
`
`of the cells. Usually systems with gentle agitation also minimize foaming. The
`
`type of impeller, battles, and tank dimensions influences the degree of mixing.
`
`Note that mammalian cell cultures are more easily damaged by these mechan-
`ical forces than microbial cultures.
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`PRIMARY PHARMACEUTICAL PRODUCTION
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`lb) pH
`
`The internal environment of living cells is approximately neutral. yet most
`microbes are relatively insensitive to the external concentrations of hydrogen
`
`and hydroxyl ions. Many organisms grow well between pH 4 and 9, although
`
`for any particular organism the required pH range is small and accurate control
`
`is essential. Note, however, that there are exceptions where growth outside this
`range can occur.
`
`(0) Sterile design
`
`The importance of sterile design cannot be over emphasized; even the presence
`of a single contaminant will be disastrous. The fermenter must be designed to
`
`be easily cleanable (smooth surfaces and no crevices), after which it must be
`sterilized. The most effective form of sterilization is to utilize clean steam to kill
`
`both the live micro—organisms and their spores. This is usually defined as
`
`maintaining 121°C for 20 minutes. Shorter times and higher temperatures can
`
`be used but not vice versa. The quality of the steam supply is important; clean
`steam is required for mammalian cell culture, whereas, plant steam with
`
`approved additives can be used for large-scale antibiotics.
`If the fermentation design calls for sterility, the following special precautions
`
`are required:
`
`a air should be provided by an oil free compressor;
`a Clean in Place (CIP) and Sterilize in Place (SIP) systems should be
`incorporated into the design;
`
`a the femienter and all associated piping and vessels should he designed to
`
`allow sterilization initially by 1.5 bar g steam. Branch connections should be
`minimized. All lines should be free draining and have minimum dead legs
`
`with the correct type of valves specified. Selection of internal surfaces,
`piping design, and valves is critical
`in ensuring effective removal of
`
`unwanted organisms during sterilization and preventing subsequent ingress
`of contaminants from outside the sterilized system:
`
`a many fermentation media. at the large scale, can be sterilized continuously
`by heat. Economies can be achieved by incorporating heat
`recovery
`
`exchangers in the system to preheat the feed;
`. all seals and instruments must be designed to withstand steam sterilization;
`
`o the equipment should be designed to maintain sterility e. g. to include the use
`
`of steam seals on agitator inlets, double O-rings for probe insertion and
`steam blocks on transfer lines;
`
`o piping should be stainless steel:
`
`0 an integrated approach should be taken to the physical layout. the piping and
`instrumentation (P&1D) flowsheeting and the sequencing to ensure that
`
`sterility is an integral part of the design;
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`PHARMACI’UTICEL PRODUCTION. AN ENGINEERING GUIDE
`
`(:1) Temperature control
`
`The temperature for organism growth ranges from approximately —5'”C to
`80°C. However,
`the actual
`temperature is important. particularly for cell
`cultures, so temperature control is critical. The lower limit is set by the freezing
`
`point of water. which is lowered by the contents of the cell. The upper limit
`
`depends on the effect of temperature on the vital constituents of the organ-
`isms — for example. protein and nucleic acids are destroyed in the temperature
`
`range 50° to 90°C.
`
`(e) Media sterilization
`
`Medium ingredients should be controlled through a careful quality assurance
`programme. However. sterilization is also required and there are essentially
`three methods used:
`
`a continuous sterilization for large scale. The time and temperature of the
`
`continuous sterilizer shOuld be optimized based on the most heat resistant
`contaminant. The hold section of the continuous sterilizer should he
`
`designed for plug flow to prevent back mixing;
`9 in—situ batch sterilization by heat for smaller hatches;
`
`a sterilization by filtration for heat sensitive products such as cell culture.
`
`Recovery and purification
`The product separation and purification section is critical to the design of a
`
`fennentation plant; indeed, the bulk of capital and operating costs for a typical
`plant are often connected with this area. The design of product recovery
`systems encompasses both intracellular and extracellular products from both
`microbial and mammalian cell fermentation broths:
`
`{a} Large—scale extracellular products
`Technologies for recovering the simpler extracellular products consist of
`conventional unit operations such as vacuum filtration, crystallization, liquid-
`
`to-liquid extraction, main-effect evaporation. precipitation and distillation.
`
`These are similar to the basic organic synthesis processes detailed earlier in
`this section.
`
`(b) Recombinant products
`Recombinant therapeutic products can be intra— or extracellular depending
`
`upon the host micro-organism. Recovery facilities for the more complex
`intracellular protein products involve cell harvesting. debris removal, pellet
`
`washing and recovery, product concentration. desalting, purification and sterile
`
`product finishing operations.
`
`The recovery and purification of protein products from fermentation broths
`involves rapidly-evolving, state-of-the—art unit operations. The complexity of
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`IPR2020-00770
`United Therapeutics EX2020
`United Therapeutics EX2020
`Page 14 of 71
`Page 14 of 71
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`PRIMARY PHARMACELlTlEAL PRODUCTION
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`these operations is increased due to the heat and shear sensitivity of the proteins
`being recovered.
`
`The use of recombinant~DNA organisms can also affect the design of the
`cell recovery area. if the organisms are not killed in the fermentation area, the
`recovery area handling the live organisms must be designed in accordance with
`applicable guidelines for containment.
`
`Typical methods for recombinant product isolation and purification inc ludc:
`
`(a) Cell disruption
`
`is inside the cells. The
`For intracellular products the product of interest
`objective of cell disruption is to release this product for further separation.
`
`Cell disruption is usually carried out by mechanical means. This can be by use
`
`of homogenizers, grinding by beads or by high pressure liquid jet impacting.
`Other methods are use of sound. pressure changes or temperature changes and
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`chemical methods. The separation of product from the cell debris alter cell
`disruption is usually done by centrifiigation.
`
`(bl Cenh'r'fiigm‘ion
`Centrifuges are commonly used for cell harvesting, debris removal. and pellet
`washing operations. Cclls can be separated using disc-stack or scroll decanter
`
`centrifuges. The latter allows cell washing prior to subsequent processing. The
`arrival of steam sterilizablc, contained designs have made the use of such
`machines more suitable.
`
`(r) Ultrafilrron'on
`
`Ultrafiltration is Widely utilized in the recrwery and purification of protein
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`products. The main uses of ultrafiltration are as follows: concentrating protein
`products; desalting product solutions by diafiltration; exchanging product
`
`buffer solutions by diafiltration; and depyrogenating of buffer solutions used
`in the process. Ultrafiltration is also finding increasingly wider use in the cell
`harvesting operation. It has an advantage over centrifugation in this situation
`since it subjects the protein to less heat and shear el’fects. Ultrafiltration is
`excellent for processes using cell recycle and in particular for mammalian cell
`applications.
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`(of) Electrodiabsrs
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`Electrodialysis is sometimes used to remove salts, acids and bases from
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`fermentation broths. A unit will consist of compartments separated by alternate
`
`anion and cation exchange membranes. A direct electric current is then passed
`through the stack to effect the separation.
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`83
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`|PR2020-OO77O
`
`IPR2020-00770
`United Therapeutics EX2020
`United Therapeutics EX2020
`Page 15 of 71
`Page 15 of 71
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`PHARMACEUTICAL I‘RDDUC'IION. AN ENGINEERING GUIDE
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`(2) Chromatography
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`Chromatography is the main technique for final