`
`Review Paper
`
`Excipients used in lyophilization of small molecules
`
`Ankit Baheti , Lokesh Kumar , Arvind K. Bansal
`a
`b
`
`b*
`
`a
`
`b
`
`Department of Pharmaceutical Technology (Formulations) National Institute of Pharmaceutical Education and Research (NIPER)
`Sector 67, S.A.S. Nagar, Punjab-160062, India
`
`Department of Pharmaceutics, National Institute of Pharmaceutical Education and Research (NIPER) Sector 67, S.A.S. Nagar,
`Punjab-160062, India
`
`Received 12 March 2010; Accepted: 7 May 2010
`
`ABSTRACT
`
`This review deals with the excipients used in various lyophilized formulations of small molecules.
`The role of excipients such as bulking agents, buffering agents, tonicity modifiers, antimicrobial
`agents, surfactants and co-solvents has been discussed. Additionally, a decision making process for
`their incorporation into the formulation matrix has been proposed. A list of ingredients used in
`lyophilized formulations marketed in USA has been created based on a survey of the Physician Desk
`Reference (PDR) and the Handbook on Injectable Drugs. Information on the recommended
`quantities of various excipients has also been provided, based on the details given in the Inactive
`Ingredient Guide (IIG).
`
`KEY WORDS: Lyophilization, excipients, bulking agent, small molecule, primary drying
`
`INTRODUCTION
`
`Lyophilization, or freeze drying, is a process in
`which water is frozen, followed by its removal
`from
`the sample,
`initially by sublimation
`(primary drying) and
`then by desorption
`(secondary drying).
`In
`this process,
`the
`moisture content of the product is reduced to
`such a
`low
`level that does not support
`biological growth or chemical reactions. The
`
`* Corresponding author: Department of Pharmaceutics,
`National Institute of Pharmaceutical Education and Research
`(NIPER) Sector 67, S.A.S. Nagar, Punjab-160062, India, Tel:
`+91-172-2214682, Fax: +91-172-2214692, e-mail:
`akbansal@niper.ac.in
`
`in
`therefore, finds special use
`technique,
`formulation development of drugs which are
`thermolabile and/or unstable
`in aqueous
`medium (1-3).
`
`Lyophilization is based on the principle of
`sublimation of ice, without entering the liquid
`phase. The phase diagram of water (Figure 1)
`show that two phases coexist along a line under
`the given conditions of
`temperature and
`pressure, while at the triple point (0.0075 °C at
`0.61kPa or 610 Nm ; 0.01 °C at 0.00603 atm),
`-2
`all three phases coexist. Lyophilization
`is
`performed at
`temperature and pressure
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 41
`
`Mylan Ex 1010, Page 1
`
`

`

`conditions below the triple point, to enable
`sublimation of
`ice. The entire process
`is
`performed at low temperature and pressure,
`hence is suited for drying of thermolabile
`compounds.
`
`Steps involved in lyophilization start from
`sample preparation
`followed by
`freezing,
`primary drying and secondary drying, to obtain
`the final dried product with desired moisture
`content (Figure 2). The concentration gradient
`of water vapor between the drying front and
`condenser is the driving force for removal of
`water during lyophilization. The vapor pressure
`of water
`increases with an
`increase
`in
`temperature during
`the primary drying.
`Therefore, primary drying temperature should
`be kept as high as possible, but below the
`critical process temperature, to avoid a loss of
`cake structure
`(4-6). This critical process
`temperature is the collapse temperature for
`amorphous substance, or eutectic melt for the
`crystalline substance (1, 7, 8).
`
`During freezing, ice crystals start separating out
`until
`the
`solution becomes maximally
`concentrated. On
`further cooling, phase
`separation of the solute and ice takes place.
`
`Review Paper
`
`Figure 2. Steps involved in lyophilization from sample
`preparation to final product formation. Annealing is an
`optional step, occasionally used
`to crystallize
`the
`formulation component(s).
`
`If the solute separates out in crystalline form, it
`is known as the eutectic temperature. In
`contrast, if an amorphous form is formed, the
`temperature is referred to as the glass transition
`temperature (T ’).
`g
`
`Determination of this critical temperature is
`important for development of an optimized
`lyophilization cycle. During primary drying,
`drying temperature should not exceed the
`critical temperature, which otherwise leads to
`‘meltback’ or ‘collapse’ phenomenon in case of
`crystalline or amorphous substance respectively
`(Figure 3).
`
`In the majority of lyophilized formulations,
`excipients are
`included
`to
`improve
`the
`functional properties and stability of
`the
`
`Figure 1 Phase diagram showing the triple point of water
`at 0.01°C, 0.00603 atm. Lyophilization is carried out
`below the triple point to enable conversion of ice into
`vapor, without entering the liquid phase (known as
`sublimation).
`
`Figure 3 Flowchart showing the concept of eutectic
`temperature and T , and their importance during
`g
`primary drying
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 42
`
`Mylan Ex 1010, Page 2
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`

`

`Review Paper
`
`CRI TERIA FOR SELECTION O F
`EXCIPIENTS
`
`lyophilized
`in a
`Selection of excipients
`formulation employs a need based approach, to
`develop a
`simple,
`stable and elegant
`formulation, with an economical process.
`Figure 6 depicts a flow chart for selection of
`different excipients for lyophilization of small
`molecules.
`
`EXCIPIENTS FOR LYOPHILIZATION OF
`SMALL MOLECULES
`
`Bulking agent
`
`Bulking agents, as the name implies, form the
`bulk of the lyophilized product and provide an
`adequate structure to the cake. These are
`generally used for low dose (high potency)
`drugs that per se do not have the necessary bulk
`to support their own structure. These are
`particularly more important when the total solid
`content is less than 2% (17). In such cases, a
`bulking agent is added to the formulation
`matrix (18, 19). The structure of the lyophilized
`
`lyophilized product. The International Pharma-
`ceutical Excipients Council has defined
`excipients as: “…substances
`other
`than
`the
`pharmacologically active drug or prodrug which are
`included in the manufacturing process or are contained
`in a finished pharmaceutical product dosage form” (9).
`Excipients also provide an aesthetic appeal to
`the product in terms of good cake structure.
`Inclusion of excipients also helps in developing
`a robust and economical lyophilization process.
`
`Neema et al. listed the excipients and frequency
`of
`their usage
`in marketed
`injectable
`formulations (10). Polwell et al. tabulated all the
`excipients used in parenteral formulation with
`reference to individual products (11). Strickley
`compiled the parenteral formulation of small
`molecules marketed in the United States (12-
`14). However,
`to our knowledge, a
`comprehensive analysis of the excipients used
`in lyophilization of small molecules does not
`exist in the literature, until now. This review
`therefore focuses specifically on the issues
`related to excipient selection in lyophilized
`formulations of small molecules. Proteins and
`peptides have not been included in the scope of
`this article. A comprehensive list of excipients
`used
`in
`lyophilized formulations of small
`molecules marketed in USA has been compiled
`from
`the Physician Desk Reference and
`Handbook on Injectable Drugs. Finally, the
`regulatory status of these excipients with
`respect to limits mentioned in IIG has been
`compiled.
`
`Table 1 lists the excipients used in marketed
`lyophilized preparations, highlighting
`the
`frequency of
`their use
`in
`lyophilized
`formulations (Figure 4). About 67% of the
`lyophilized marketed preparations of small
`molecules contain excipient(s)
`in
`their
`formulation.
`
`CLASSIFICATION OF EXCIPIENTS
`
`The excipients commonly used
`in
`lyophilization of small molecules have been
`classified in Figure 5.
`
`Figure 4 Distribution of commonly used excipients in
`marketed lyophilized formulations of small molecules.
`About 67% of marketed lyophilized formulations of small
`molecules contain excipients.
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 43
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`Mylan Ex 1010, Page 3
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`

`

`Table 1 List of excipients used in lyophilized formulation of small molecules, as marketed in USA (15, 16)
`
`Review Paper
`
`Drug
`
`Amifostine
`
`Category
`
`Excipients
`
`Route of administration
`
`Marketed name
`
`Cytoprotective agent
`
`-
`
`IV infusion over 15-30 min
`
`Amphotericin B cholesteryl
`sulfate
`
`Antifungal
`
`Amphotericin B
`
`Antifunal
`
`Sodium cholesteryl sulfate
`Lactose
`Tris
`EDTA
`
`Hydrogenated
`soyaphosphatidylcholine
`Disteroylphosphatidly glycerol
`Cholesterol
`Alpha tocopherol
`Sucrose
`Disodium succinate
`
`-
`
`-
`
`á-cyclodextrin
`Lactose
`
`Lactose
`Sodium citrate
`Benzyl alcohol
`
`IV infusion at 3-4 mg/kg/hr
`
`Ethyol (MedImmune
`®
`Oncology)
`
`Amphotec (Sequus
`®
`Pharmaceuticals)
`
`IV infusion at 3-5 mg/kg/hr
`
`Ambisome (Astellas)
`®
`
`IV infusion over 1 hr
`
`IV infusion
`
`Intracavernosal
`
`Intracavernosal
`
`IV bolus, IV infusion
`
`IV infusion
`
`IM, IV bolus, IV infusion
`
`IV infusion
`
`IM, IV bolus, IV infusion
`
`IM, IV bolus, IV infusion
`
`Zovirax ®
`(Glaxo Wellcome)
`
`Aloprim (Nabi
`®
`Biopharmaceuticals)
`
`Edex ®
`(Schwarz Pharma)
`
`Caverject (Phamacia and
`®
`Upjohn)
`
`Imuran ®
`(Glaxo Wellcome)
`
`Zithromax (Pfizer)
`
`®
`
`Azactam ®
`(Bristol Myers Squibb)
`
`BiCNU ®
`(Bristol Myers Squibb)
`
`Kefzol®
` (Lilly)
`
`Ancef ®
`(GlaxoSmith-Kline)
`
`Acyclovir sodium
`
`Allopurinol sodium
`
`Alprostadil
`
`Alprostadil
`
`Azathioprine sodium
`
`Azithromycin
`
`Aztreonam
`
`Carmustine
`
`Cefazolin sodium
`
`Cefazolin sodium
`
`Antiviral
`
`Anti-gout
`
`Erectile dysfunction
`
`Erectile dysfunction
`
`Immunosuppressive
`antimetabolite; management
`of severe rheumatoid arthritis
`
`-
`
`Antibiotic
`
`Antibiotic
`
`Antineoplastic
`
`Antibiotic
`
`Antibiotic
`
`Citric acid
`
`L- arginine
`
`-
`
`-
`
`-
`
`Chlorothiazide sodium
`
`Diuretic and hypertensive
`
`Mannitol
`Thiomersol
`
`Cisplatin
`
`Antineoplastic
`
`IV bolus, IV infusion
`
`IV infusion
`
`Diuril ®
`(Merck)
`
`®
`
`Diltiazem
`
`Antianginal
`
`Mannitol
`
`IV bolus, IV infusion
`
`Intratracheal
`
`Platinol (Bristol Myers
`Oncology)
`
`Exosurf neonatal (Glaxo
`Wellcome)
`
`®
`
`IM, IV bolus, IV infusion, IP,
`Intrapleural
`
`Cytoxan ®
`(Bristol Myers Squibb)
`
`IV bolus, IV infusion
`
`Cosmegen (Merck)
`
`®
`
`IV bolus, IV infusion over 1 hr
`
`Dantrium (Procter & Gamble)
`
`®
`
`IV infusion
`
`IV
`
`IV
`
`Cerubidine (Bedford)
`
`®
`
`®
`
`Zinecard (Pharmacia &
`Upjohn)
`
`®
`
`Cardizem (Hoechst Marion
`Roussel)
`
`Rubex (Bristol Myers Squibb)
`
`®
`
`IV infusion over 30-60 min
`
`IV infusion
`
`Etopophos (Bristol Myers
`Squibb)
`
`®
`
`Flolan (Glaxo Wellcome)
`
`®
`
`Slow IV bolus, IV infusion
`
`Sodium edecrin (Merck)
`
`®
`
`IV infusion over 30 min
`
`Fludara (Berlex)
`
`®
`
`IV infusion at 5mg/kg over 1
`hr
`
`Cytovene (Roche)
`
`®
`
`Colfosceril palmitrate
`
`Prevention and treatment of
`respiratorydisease syndrome
`in low birth weight infants
`
`Cyclophosphamide
`
`Antineoplastic
`
`Dactinomycin
`
`Dantrolene sodium
`
`Daunorubicin HCl
`
`Dexrazoxane
`
`Antibiotic
`
`Muscle relaxant
`
`Antibiotic
`
`Cardioprotective agent
`
`-
`
`Mannitol
`Sodium chloride
`
`Cetyl alcohol
`Tyloxapol
`Sodium chloride
`
`Mannitol
`
`Mannitol
`
`Mannitol
`
`Mannitol
`
`Doxorubicin HCl
`
`Antineoplastic
`
`Etoposide phosphate
`
`Antineoplastic
`
`Epoprostenol sodium
`
`Antihypertensive
`
`Ethacrynate sodium
`
`Diuretic
`
`Fludarabine phosphate
`
`Antineoplastic
`
`Lactose
`Methyl paraben
`
`Sodium citrate
`Dextran 40
`
`Mannitol
`Sodium chloride
`Glycine
`
`Mannitol
`
`Mannitol
`
`Ganciclovir sodium
`
`Treatment of CMV retinitis in
`immunocompromized patient
`
`-
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 44
`
`Mylan Ex 1010, Page 4
`
`

`

`Review Paper
`
`Table 1 List of excipients used in lyophilized formulation of small molecules, as marketed in USA (15, 16)
`
`Drug
`
`Gemcitabine HCl
`
`Hemin
`
`Category
`
`Antineoplastic
`
`Treatment of acute
`intermittent porphyria related
`to mensuration
`
`Excipients
`
`Mannitol
`Sodium acetate
`
`Sorbitol
`Sodium carbonate
`
`Route of administration
`
`Marketed name
`
`IV infusion over 30 min
`
`Genzer (Lilly)
`®
`
`IV infusion
`
`Panhematin (Abbott)
`®
`
`Hydromorphone HCl
`
`Opioid analgesic
`
`Indomethacin sodium
`
`NSAID
`
`-
`
`-
`
`Lansoprazole
`
`Proton pump inhibitor
`
`Mannitol
`Meglumine
`Sodium hydroxide
`
`IV, IM, SC
`
`IV bolus
`
`IV
`
`Levothyroxine sodium
`
`Hormone relacement
`
`Mannitol
`Sodium phosphate tribasic
`
`IM, IV
`
`Dilaudid-HP (Abbott)
`®
`
`Indocin I.V. (Merck)
`®
`
`Prevacid (TAP)
`®
`
`Synthrod (Knoll)
`®
`
`Melphalan HCl
`
`Antineoplastic
`
`Povidone
`Diluent: Water, propylene
`glycol, ethyl alcohol, sodium
`citrate
`
`IV infusion over 15-20 min
`
`Alkeran (Celgene)
`®
`
`Methohexital sodium
`
`Anesthetic
`
`Anhydrous sodium carbonate
`
`IV, IM
`
`Brevital sodium (KING)
`®
`
`Methyl prednisolone succinate
`sodium
`
`Hormone replacement
`
`Sodium phosphate
`Lactose
`Benzyl alcohol
`
`IM, IV bolus, IV infusion
`
`Solu-Medrol (Pfizer)
`®
`
`Metronidazole
`
`Mitomycin
`
`Antibacterial
`
`Antineoplastic
`
`Pamidronate disodium
`
`Inhibition of bone resorption
`
`Pentostatin
`
`Antineoplastic
`
`Phentolamine mesylate
`
`Antihypertensive
`
`Pipecuronium bromide
`
`Long acting neuromuscular
`blocking agent
`
`Mannitol
`
`Lactose
`
`Mannitol
`
`Mannitol
`
`Mannnitol
`
`-
`
`-
`
`IV bolus, IV infusion
`
`Flagyl (Pfizer)
`
`®
`
`IV infusion
`
`IV
`
`Mutramycin (Bristol Myers
`Squibb)
`
`®
`
`Aredia (Novartis)
`
`®
`
`Slow IV bolus, IV infusion
`
`Nipent (Supergen)
`
`®
`
`IM, IV bolus, IV infusion
`
`Regitine (Novartis)
`
`®
`
`IV bolus
`
`Arduran (Oryannon)
`
`®
`
`IV bolus, IV infusion
`
`®
`
`Pralidoxime chloride
`
`Remifentanil HCl
`
`Streptozocin
`
`Antidote for overdose due to
`anticholinesterase
`
`Analgesic
`
`Antineoplastic
`
`Tazobactam sodium and
`Piperacillin sodium
`
`Antibacterial combination
`
`Glycine
`
`Citric acid
`
`EDTA
`Sodium citrate
`
`Thipental sodium
`
`Short acting anesthetic
`
`Sodium carbonate
`
`Thiotepa
`
`Antineoplastic
`
`-
`
`Thiothixene HCl
`
`Ticarcillin disodium
`
`Tigecycline
`
`Topotecan
`
`Antipsychotic
`
`Antibacterial
`
`Antibacterial
`
`Antineoplastic
`
`Trimetrexate glucuronate
`
`Treatment of pneumonia
`
`Vancomycin HCl
`
`Antibiotic
`
`Vecuronium bromide
`
`Muscle relaxant
`
`Vinblastine sulfate
`
`Warfarin sodium
`
`Antineoplastic
`
`Anticoagulant
`
`Mannitol
`
`-
`
`-
`
`Mannitol
`Tartaric acid
`
`-
`
`-
`
`Mannitol
`Citric acid
`Sodium phosphate dibasic
`
`-
`
`Mannitol
`Sodium chloride
`Sodium
`phosphate,monobasic,
`monohydrate
`Sodium phosphate, dibasic,
`heptahydrate
`
`IV infusion
`
`IV bolus, IV infusion
`
`IV infusion
`
`IV infusion
`
`IV bolus, Intracavitary,
`Intravesical
`
`Protopam (Baxter
`Healthcare)
`
`Ultiva (GlaxoWellcome)
`
`®
`
`®
`
`Zanosar (Pharmacia &
`Upjohn)
`
`Zosyn (Lederle)
`
`®
`
`Pentothal sodium (Baxter)
`
`®
`
`Thioplex (Immunex)
`
`®
`
`IM
`
`Navane (Pfizer)
`
`®
`
`IM, IV bolus, IV infusion
`
`Ticar (Smith Kline Beecham)
`
`®
`
`IV infusion
`
`IV infusion
`
`IV infusion
`
`IV infusion
`
`IV bolus, IV infusion
`
`IV bolus
`
`Slow IV over 2 min
`
`Tygacil (Wyeth)
`
`®
`
`®
`
`Hycamtin (Smith Kline
`Beecham)
`
`Neutrexin (U.S. Biosciences)
`
`®
`
`Vancocin HCl (Lilly)
`
`®
`
`Norcuron (Organon)
`
`®
`
`Velban (Lilly)
`
`®
`
`Coumandin (Bristol Myers
`Squibb)
`
`®
`
`HCl – hydrochloric acid; i.v. – intravenous; i.m. – intramuscular; s.c. – subcutaneous; PDR- Physicians Desk Reference; EDTA – ethylenediaminetetraacetic acid
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 45
`
`Mylan Ex 1010, Page 5
`
`

`

`Review Paper
`
`Figure 5 Classification of commonly used excipients used in lyophilization of small
`molecules. A need based approach is utilized to select the appropriate excipient(s) for
`lyophilization.
`
`cake is important, since proper cake formation
`leads to proper pore formation that provides
`the means for vapor to escape from the
`product during the drying cycle. Loss of
`product structure blocks the path for vapor
`removal leading to an increased resistance in
`moisture removal and thus higher moisture
`content in the final product (Figure 7). Such
`localized high moisture content may lead to
`degradation of
`the active pharmaceutical
`ingredient during the shelf life. Kovalcik and
`Guillory demonstrated the need of a bulking
`agent for cyclophosphamide. They also showed
`that the moisture containing cake showed
`better stability with mannitol, than with lactose
`as bulking agent (20). Mannitol and glycine, are
`the most commonly used bulking agents,
`followed by glucose, sucrose, lactose, trehalose
`and dextran (21).
`
`The bulking agent may appear as crystalline
`and/or amorphous solid at
`the end of
`lyophilization process. Crystallization of the
`bulking agent, however, might adversely
`affect the physical stability of the product in
`certain instances, for which, an amorphous
`bulking agent would be preferred. Herman et al.
`studied
`the
`rate of hydrolysis of
`methylprednisolone sodium succinate in the
`presence of mannitol and lactose as the bulking
`agents, and observed that formulation with
`mannitol showed a faster degradation
`in
`comparison to
`that with
`lactose. It was
`hypothesized to be due to the crystallization of
`mannitol during lyophilization, unlike lactose,
`which remained in the amorphous state (22).
`The non-hygroscopic nature of crystalline
`mannitol, led to an increase in the amount of
`water available with the drug. In contrast, the
`hygroscopic amorphous lactose held water
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 46
`
`Mylan Ex 1010, Page 6
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`

`

`
`
`Preformulation
`Stability - thermal, pH, photostability
`Solubility - pH solubility profile
`Compatibility - excipients and CCS
`Dose - single/multiple units
`
`Review Paper
`
`Is pH critical
`for stability?
`
`Yes
`
`Add buffering agent
`
`No
`
`Is the drug
`solubility
`low?
`
`Yes
`
`Add solubilizing agent
`
`No
`
`Is the product
`for multiple
`use?
`
`Yes
`
`Add antimicrobial agents
`
`No
`
`Lyophilize using prototype
`lyophilization cycle
`
`#only in vial lyophilization
`
`No
`
`Bulking agent
`required
`
`Change buffering
`agent
`
`No
`
`Is cake
`structure
`adequate?
`
`Yes
`
`Is the desired
`pH maintained
`during
`lyophilization
`and after
`reconstitution?
`
`
`Yes
`
`#
`
`Figure 6 Decision tree for selection of excipients for lyophilization of small molecules (continued next page)
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 47
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`Mylan Ex 1010, Page 7
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`

`

`#
`
`Is drug
`crystalline?
`
`Yes
`
`Is
`process
`lengthy?
`
`Review Paper
`
`Add organic
`solvent to reduce
`processing time
`
`Yes
`
`No
`
`No
`
`Is annealing
`sufficient to
`crystallize the
`drug?
`
`
`No
`
`Does addition
`of cosolvent
`crystallizes
`the drug?
`
`
`No
`
`Does combination
`of cosolvent and
`annealing
`crystallize drug?
`
`
`No
`
`Drug remains amorphous
`after lyophilization
`
`Yes
`
`Perform annealing to
`crystallize drug
`
`Yes
`
`Add cosolvents to
`crystallize drug
`
`Yes
`
`Use a combination of
`cosolvents and annealing
`to crystallize
`
`Is the
`process
`lengthy?
`
`Yes
`
`Add collapse
`temperature modifiers
`
`No
`
`Lyophilized product
`
`CCS – Container closure system
`
`
`
`Figure 6 Continued from previous page
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 48
`
`Mylan Ex 1010, Page 8
`
`

`

`molecules, leading to a relative decrease in the
`amount of water available
`in the micro-
`environment of drug.
`
`The bulking agent may also crystallize in
`different polymorphic
`forms during
`lyophilization, which could also have serious
`implications on physical stability of drug
`product. Liao et al. reported that mannitol
`crystallizes as a mixture of ä-mannitol and
`mannitol hemihydrate during annealing. Release
`of water from hemihydrate form during
`shipping and storage conditions might cause
`degradation of moisture sensitive drug product
`(23).
`
`The nature of lyophilized cake also depends on
`the ratio of drug and bulking agent, showing an
`increased crystallization with an increase in
`amount of bulking agent. Korey and Schwartz
`studied the lyophilization of active constituents
`such as atropine sulfate, sodium cefoxitin,
`cefazolin
`sodium
`and procainamide
`hydrochloride. They showed that crystallization
`was induced by excipients such as glycine,
`alanine,
`serine, methionine, urea, and
`niacinamide, whereas lyophilization with some
`excipients such as mannitol and lactose gave an
`amorphous product. The degree of
`crystallization increased with an increase in the
`mole fraction of the excipient. For example,
`when the mole fraction of glycine in the
`solution was 0.41, the lyophilized cake of
`cefazolin sodium was amorphous. As the mole
`
`Figure 7 Good cake structure ensures proper pore
`formation which provides the channels for vapor removal
`(A) whereas in case of cake collapse, a poor structure is
`obtained, leading to increased resistance in removal of
`vapor (B).
`
`Review Paper
`
`to 0.80, complete
`increased
`fraction was
`crystallization of cefazolin
`sodium was
`observed. It was hypothesized that (i) the active
`constituent and excipient forms a molecular
`complex that crystallizes during the freezing
`process and/or (ii) the excipient crystallizes
`during the freezing process and serves as a seed
`crystal for the active constituent and/or (iii) the
`active constituent and excipient formed a
`eutectic during the freezing process (24).
`
`Buffering agent
`
`Control of pH is critical to avoid degradation of
`drug during processing,
`storage and
`reconstitution, thereby necessitating addition of
`buffering agent in the lyophilized formulation.
`The choice of buffer depends on the pH
`stability profile of active ingredient as drug
`needs to be reconstituted and stored for some
`time before it could be administered to the
`patient. For this purpose, the pH of maximum
`stability of drug should be known and
`maintained. Selection of a suitable buffer and
`its concentration is important for sensitive
`molecules. For example
`in aspartame
`lyophilizates, the presence of 0.1M phosphate
`buffer caused the half life of the material to
`decrease from 921 days in unbuffered material
`to 98 days; increasing the buffer concentration
`further causes a reduction to 77 days (25).
`
`The buffering agent should have a high collapse
`temperature, be non-volatile and have a high
`glass transition temperature (T ) (26). A high
`g
`collapse temperature would facilitate a faster
`primary drying, and its non-volatile nature
`would prevent pH drift,
`that might be
`detrimental
`to
`the product
`stability.
`Additionally, a high glass transition temperature
`(T ) would ensure stability during storage. In
`g
`this context, acetate buffer is not used due to its
`volatile nature, as it can be partially lost during
`lyophilization (27). Crystallization of buffer
`components can also lead to a drastic shift in
`pH, resulting in degradation of the active
`component. Sodium and potassium phosphate
`salts are not often used in the lyophilization,
`since these crystallize during cooling and in
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 49
`
`Mylan Ex 1010, Page 9
`
`

`

`frozen solution, which leads to a decrease in
`pH of about 4 units (28). Shalave et al. studied
`citrate, succinate and tartrate buffer for their
`crystallization behavior and its effect on pH of
`the formulation. Citrate buffer was found to be
`the most preferred as it remained amorphous,
`with the shift
`in pH being minimal,
`in
`comparison to succinate and tartrate, which
`crystallized during lyophilization (29).
`
` “pH memory” is a term used to denote the
`relationship between pH-activity and pH-
`stability profiles, in the solution and dried state
`respectively, as the pH of the solution before
`drying has an impact on the rate of chemical
`reactivity in the resulting amorphous material
`(30-33). Guo et al. found that depending on the
`initial pH of an aqueous solution of quinapril
`hydrochloride,
`lyophilization produced a
`mixture of quinapril hydrochloride and the
`neutralized form. Increase in pH led to a
`proportionate
`increase
`in
`the amount of
`neutralized form. Since neutralized form is less
`stable than hydrochloride salt, the stability of
`the
`lyophilized sample decreased with an
`increase
`in the pH of solution used for
`lyophilization (34). In certain cases, however,
`pH stability of a drug in lyophilized form may
`show an unusual behaviour in comparison to
`the pH-stability in solution state. For example,
`moexipril is most stable at a pH of 4.5 in
`solution. However, when moexipril was
`lyophilized using a solution of pH range of 2-
`11, it was found to be most unstable at a pH of
`5.1. This was attributed
`to
`the altered
`mechanism of reactivity in absence of water in
`lyophilized product, compared to the aqueous
`solution (35). A similar effect was shown for
`the ionization of sulfonephthalein in trehalose-
`citrate systems, where protonation of indicators
`were higher in the lyophilized sample than in
`solution at a particular pH (36).
`
`Collapse temperature modifiers
`
`Lyophilization of amorphous material requires
`the primary drying temperature to be kept
`below
`the collapse
`temperature of
`the
`formulation. However, some excipients (37) in
`
`Review Paper
`
`amorphous state have a very low collapse
`temperature, thus increasing the duration of
`primary drying significantly. In such case,
`collapse temperature modifiers are utilized,
`which shift the overall collapse temperature
`higher (owing to their high individual collapse
`temperatures), with a consequent reduction in
`the primary drying
`cycle, without
`compromising
`the product quality
`(25).
`Commonly used collapse temperature modifiers
`®
`are dextran, Ficoll
`, gelatin
`and
`hydroxyethylstarch
`(38). Their collapse
`temperatures (along with other critical process
`temperatures for different excipient classes) are
`summarized in Table 2. However, it must be
`noted that their use in lyophilized formulations
`is not very frequent.
`
`Table 2 Critcal process temperatures of various
`excipients used in lyophilization
`Excipient
`T ’
`T
`g
`
`References
`
`c
`
`BULKING AGENT
`Sucrose
`Lactose
`Trehalose
`Mannitol
`Sorbitol
`
`Glucose
`
`Raffinose
`Glycine
`Histidine
`PVP (K40)
`
`-32, -35
`-28
`-27, -29
`-35, -28
`-46
`
`-43
`
`-27
`-62
`-33
`-20
`
`BUFFERING AGENT
`Sodium citrate
`Sodium phosphate
`Sodium hydroxide
`Meglumine
`Tris base
`Tris HCl
`Tris acetate
`
`-41
`-45
`
`-51
`-65
`-54
`
`TONICITY M ODIFIER
`Sodium chloride
`Dextrose
`
`-44
`
`COLLAPSE
`TEM PERATURE
`M ODIFIER
`Dextran
`Ficoll
`Gelatin
`Hydroxyethyl starch
`
`-10
`-19
`-9
`
`-34,-32
`-31, -32
`-29.5, -34
`
`-45
`-40, -41.5
`-43
`-26
`
`-23
`
`(39-41)
`(39, 40, 42)
`(39, 41, 42)
`(39, 42)
`(40, 42, 43)
`
`(40, 41, 44)
`
`(43)
`(45)
`(39)
`(40, 42, 43)
`
`(39)
`(39)
`
`(39)
`(39)
`(39)
`
`(42)
`
`(38)
`
`-9, -10
`-19.5, -20
`-8
`-5
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 50
`
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`
`

`

`Review Paper
`
`Miscellaneous Excipients
`
`Tonicity modifiers
`
`Solubilizing agent
`
`Surfactants
`
`Parenteral formulations should be isotonic with
`human plasma so as to avoid damage to the
`tissues. However, not all drugs at
`their
`recommended dosage are isotonic with blood,
`thus requiring the addition of a tonicity
`adjusting agent to the formulation. The most
`commonly used tonacity agent is dextrose,
`while others, such as glycerol and sodium
`chloride are less commonly used. The addition
`of tonicity modifiers to the
`lyophilization
`mixture, however, can complicate
`the
`formulation development, since they may lower
`the collapse
`temperature of
`the entire
`formulation owing to their very low col- lapse
`temperatures,
`thus
`increasing
`the primary
`drying
`time
`significantly. An alternative
`approach is to add the tonicity modifier to the
`reconstitution diluent rather than the freeze
`dried product. Deviations from isotonicity may
`be acceptable when the injection volumes are
`small and the infusion rate is slow (46).
`
`Antimicrobial agents
`
`Antimicrobial agents are added to multi-dose
`formulations
`to prevent microbial growth
`during its shelf life. Benzyl alcohol and a
`mixture of ethyl- and methyl- parabens are
`commonly used. Additionally, phenol and m-
`cresol are utilized in lyophilization. At very low
`(i.e., #0.05% w/w
`levels
`in
`solution),
`antimicrobial agents generally do not alter the
`collapse
`temperature of
`the
`formulation.
`Compatibility of antimicrobial agent with other
`ingredients in the formulation needs to be
`checked, when to the lyophilized cake (8).
`However, since the antimicrobial agent is not
`needed during
`the
`lyophilization process
`antimicrobial agents are often typically included
`in the diluent for reconstitution (27).
`
`Surfactants may be added at low levels (e.g.,
`~0.05% w/w aqueous
`solution)
`to aid
`reconstitution if the drug does not show good
`wetting behaviour. Surfactants are added to low
`dose products to minimize losses due to surface
`adsorption. Their addition in large quantity is
`not recommended due
`to
`the
`low glass
`transition temperature of commonly used non-
`ionic surfactants (e.g. Polysorbate 80) (47).
`Halikala et al. showed the effect of surfactant
`on the crystallization behavior of mannitol. It
`was observed
`that with
`increase
`in
`concentration of polysorbate 80,
`the
`crystallinity of mannitol
`increased and at
`concentration of 0.01% and above, increased
`amount of ä–mannitol was formed (48). They
`can be included in the diluting medium so as to
`keep the lyophilized formulation simple.
`
`Co-solvents
`
`Water is the most commonly used solvent for
`lyophilization. However, organic solvents are
`sometimes used to increase the primary drying
`rate by
`increasing
`the sublimation rates,
`improve product
`stability, decrease
`reconstitution
`time by
`improving drug
`wettability or solubility, and also enhance the
`sterility assurance of the sample solution (49).
`Since lyophilization works on the principle of
`vapor pressure differential, it is necessary for
`excipients to have low vapor pressure, to
`minimize their loss during the lyophilization
`process. However, co-solvents are added due to
`their high vapor pressure to facilitate faster
`removal from
`the product during drying
`process and thus speeding up the lyophilization
`process. It has been reported that tert-butyl
`alcohol has been used to speed up the
`lyophilization process
`(50). The
`rate of
`degradation of active constituent
`in
`the
`presence of water can be reduced by addition
`of non-aqueous solvents (51, 52). In some
`cases, solvent may play no role in stabilization
`
`This Journal is © IPEC-Americas Inc
`
`J. Excipients and Food Chem. 1 (1) 2010 - 51
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`Mylan Ex 1010, Page 11
`
`

`

`of the active constituent but improve the
`aesthetic property of the final product (50).
`
`The use of co-solvents, however, requires
`proper handling and storage, consideration of
`the level of residual solvent and its potential
`toxicity. The most commonly used solvent is a
`tertbutanol/water combination. Tertbutanol
`modifies the crystal habit of ice and promotes
`its sublimation, thus reducing the duration of
`primary drying (53, 54). Telang et al. studied the
`effect of solvents on the crystallization of
`cefalothin sodium. The lyophilized cake formed
`with ethanol and isopropanol were found to
`collapse, while tert-butyl alcohol gave a good
`cake
`structure.
`Increased degree of
`supersaturation and nucleation during freezing,
`followed by annealing due to co-solvent, led to
`a faster crystallization rate
`(55). Ethanol,
`although being the most generally used solvent
`in the laboratory, is less preferred as a co-
`solvent, as a shelf temperature of about -120°C
`is required to attain frozen state, thus requiring
`a tremendous rise in energy consumption. In
`contrast, tert-butyl alcohol does not require
`additional cooling (50).
`
`Complexing agent
`
`Complexation is sometimes used to improve
`the solubility of drug in the solvent especially
`water. Kagkadis et al. used hydroxypropyl-â-
`cyclodextrin (HP-â-CD) complex of ibuprofen
`to increase the solubility of lyophilized product
`(56). The addition of complexing agent may,
`however, lead to a reduction in the critical.
`temperature of
`the
`formulation,
`thus
`complicating the formulation development.
`
`LEGAL STATUS OF EXCIPIENTS
`
`Excipients added to the lyophilized cake should
`have regulatory acceptance, as they are intended
`for parenteral administration. The
`list of
`approved excipients with their maximum limit
`for
`lyophilization of small molecules, as
`reported
`in the IIG (Inactive Ingredients
`Guide) is listed in Table 3 (37).
`
`Review Paper
`
`Table 3 List of excipients used in ‘Powder, for
`injection solution, lyophilized’ with the maximum
`allowable limits as per IIG (37)
`
`Inactive ingredient
`
`Mannitol
`
`Mannitol
`a
`Mannitol
`Mannitol
`a
`Mannitol
`Anhydrous lactose
`Anhydrous lactose
`Lactose
`Lactose
`Lactose
`Lactose m onohydrate
`Lactose m onohydrate
`Lactose m onohydrate
`Lactose m onohydrate
`Lactose, hydrous
`Sucrose
`Sucroseb
`Sucrose
`Sucrose
`Sucrose
`Sucrosea
`Glycine
`Glycine
`Glycine
`Dextran 40
`Anhydrous citric acid
`Citric acid
`Citric acid m onohydrate
`Sodium citrate
`Sodium citrate
`Sodium citrate
`Sodium citrate
`Trisodium citrate dihydrate
`Sodium phosphate
`dihydrate
`Sodium phosphate,
`dibasic
`Sodium phosphate,
`dibasic anhydrous
`Sodium phosphate,
`dibasic anhydrous
`Sodium phosphate,
`dibasic anhydrous
`Sodium phosphate,
`dibasic, dihydrate
`Sodium phosphate,
`dibasic, heptahydrate
`Sodium phosphate,
`dibasic, h