Volume 23 Number 4
`
`INTERNATIONAL
`
`Aprll 2010
`
`The Science & Business of Biopharmaceuticals
`
`ww1w.biapha.rminternational.com
`
`26
`
`FORMULATION
`A Better Way to Study
`Leachables from
`Container Closures
`A systematic approach facilitates
`formulation component selection
`PROCESS DEVELOPMENT
`Harness the Power
`of CFD for Bloprocess
`Troubleshooting
`Computational fluid dynamics is a
`powerful tool·to optimize processes 38
`DOWNSTREAM PROCESSING
`Optimize Pollshlng
`In a Two-Step Process
`I High throughput process development
`allows rapid screening of
`46
`chromatographic parameters
`
`REGUlATORY BEAT
`Complex REMS programs
`raise manufacturers' concerns
`I OUTSOURCING INSIGHTS
`I Is the balance of power shifting
`toward clinical supply chain CROs?
`
`16
`
`DOWNSREAM TRENDS
`What's inhibiting downstream
`technology innovation?
`
`21
`
`BURRlll ON BIOTECH
`Initial public offerings are back,
`
`Here's what to expect in 2010.
`
`l but not everyone will make the cut.
`
`AN ~
`
`-PUBLICATIO ... N-~~
`
`FINAL WORD: Biotech Needs Policies that Encourage Innovation
`
`58
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 1
`
`

`

`Extractables & Leachables
`
`Xioochun Yu, PhD, is a senior research
`scientist, Don De(ou, PhD, Is a senior research
`scientist, Derek Wood is a laboratory manager,
`Sleven Zdrovkovic Is a senior scientist, Heolher
`Schmid! is an associate research scientist,
`Laurie Slockmeier is a scientist, Robert Piccoli,
`PhD, is an associate research scientist, Doniel
`Rude is a research scientist, and Xiaoyo Ding,
`PhD, is the director of scientific and technical
`affairs, all at the cGMP laboratory, PPD, Inc.,
`Wilmington, NC, 910.558.7585,
`xiaoya.ding@ppdi.com.
`
`A Study of Leachables
`for Biopharmaceutical
`Formulations Stored
`in Rubber-Stoppered
`Glass Vials
`
`Xiaochun Yu, Don DeCou, Derek Wood, Steven Zdravkovlc,
`Heather Schmidt, Laurie Stockmeler, Robert Piccoli, Daniel Rude, Xiaoya Ding
`
`ABSTRACT
`This article describes a systematic approach to evaluating the leachables profile
`from various biopharmaceutical formulations stored in rubber-stoppered glass
`vials. We evaluate leachables from rubber stoppers in 11 different formulations
`containing typical biopharmaceutical exclpients to assess how differences in
`various formulation excipients affect leachables. The information obtained from this
`type of study is critical to support the Quality by Design paradigm of incorporating
`product quality into the product design process and using risk-based approaches to
`managing quality.
`
`B iopharmaceutical packaging
`
`materials (container/closure
`systems) must be evaluated
`for compatibility with the
`drug formulation in the early stages
`of development to ensure the product
`is safe for use throughout its shelf life.
`This requirement also applies to the
`development of biopharmaceutical
`manufacturing processes including
`single-use components used in manu(cid:173)
`facturing equipment and in-process
`production conditions.1 Although
`compendia! tests provide some pre(cid:173)
`liminary extractables information
`about the container/closure systems,
`they do not identify or quantitate
`individual extractables. Therefore,
`they cannot correlate extractables
`
`from the container/closure systems to
`the leachables in the drug products.
`This article describes a systematic
`approach to determining extractables
`and leachables based on the concepts
`proposed by the Product Quality
`Research Institute's Leachables and
`Extractables Working Group for the
`development of orally inhaled and
`nasal drug products in concert with
`relevant regulatory guidelines.2-4 The
`study design focuses on identify(cid:173)
`ing and elucidating the correlation
`between leachables caused by bio(cid:173)
`pharmaceutical formulations stored
`in rubber-stoppered glass vials and its
`potential application in supporting
`Quality by Design (QbD) in the bio(cid:173)
`pharmaceutical development process.
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 2
`
`

`

`Extractables & Leachables
`
`Formulation
`
`Buffer with
`co-solvent
`
`pH variations
`
`Chelating
`agent
`
`Surfactant
`
`Bulking agent
`
`2
`
`3
`
`4
`
`5
`
`6
`
`7
`
`8
`
`9
`
`10
`
`11
`
`EDTA: ethylenedlemlnelelraacetlc acid
`
`Buffer
`(20 mM)
`pH 6.8
`phosphate
`pH 6.8
`phosphate
`pH 5.0
`citrate
`pH 6.8
`phosphate
`pH 8.2
`phosphate
`
`pH 6.8
`phosphate
`
`7%
`sucrose
`7%
`sucrose
`7%
`sucrose
`
`7%
`sucrose
`
`pH 6.8
`phosphate
`
`7%
`sucrose
`
`pH 6.8
`phosphate
`
`pH 6.8
`phosphate
`
`pH 6.8
`phosphate
`pH 6.8
`phosphate
`
`7%
`sucrose
`
`7%
`sucrose
`
`7%
`mannitol
`7%
`trehalose
`
`Table 1. Aqueous formulations used for leachables evaluations
`Formulation
`Bulking
`number
`agent
`
`Stabilizer
`
`Tonicity
`modifier
`
`Chelating
`agent
`
`Surfactant Co-solvent
`
`2% glycerol
`
`Sucrose
`
`Sucrose
`
`Sucrose
`
`Sucrose
`
`Sucrose
`
`Sucrose
`
`Sucrose
`
`Trehalose
`
`150mM
`NaCl
`150mM
`NaCl
`150mM
`NaCl
`
`150 mM
`NaCl
`
`150mM
`NaCl
`
`150 mM
`NaCl
`
`150 mM
`NaCl
`
`150 mM
`NaCl
`150 mM
`NaCl
`
`0.1 mM
`EDTA
`disodium
`0.5mM
`EDTA
`disodium
`0.1 mM
`EDTA
`disodium
`0.1 mM
`EDTA
`dlsodium
`
`0.1%
`Tween 80
`
`0.5%
`Tween BO
`
`Glass vials and rubber stoppers are
`widely used as container/closure systems
`for biopharmaceutical products and other
`drug formulations. Organic compounds
`in the stoppers, such as oligomers, anti(cid:173)
`oxidants, and curing agents, can leach out
`into drug formulations and affect drug
`safety and efficacy. In addition, typical
`biopharmaceutical drug product formula(cid:173)
`tion ingredients, such as co-solvents, sur(cid:173)
`factants, chelating agents, bulking agents,
`and pH modifiers, can alter the physico(cid:173)
`chemical properties of the drug formula(cid:173)
`tion itself and also may have an impact on
`the leaching of organic compounds from
`rubber stoppers. This paper evaluates leach(cid:173)
`ables from rubber stoppers in 11 different
`formulations containing typical biophar(cid:173)
`maceutical _excipients, and addresses how
`differences in various formulation excipi(cid:173)
`ents affect leachables.
`In this study, typical biopharmaceutical for(cid:173)
`mulation components were chosen to create a
`variety of test formulations. The extractables
`
`profiles of chlorobutyl rubber stoppers were
`determined. The major extractables were then
`chosen as targets for a leachable assessment
`in which the test formulations were stored
`at 40 °C and 75% relative humidity (RH) for
`one month. The following formulation varia(cid:173)
`tions were evaluated: pH (5.0, 6.8, 8.2), chelat(cid:173)
`ing agent (0.1 to 0.5 mM EDTA), surfactant
`concentration (0.1-0.5% Tween 80), bulking
`agent (sucrose, mannitol, or trehalose), and
`the presence of a co-solvent (2% glycerol).
`STUDY DESIGN
`Eleven aqueous placebo formulations were
`evaluated in this study (Table 1). The various
`test formulations included a simple phosphate
`buffer at pH 6,8, a buffer with a glycerol co(cid:173)
`solvent, three formulations in which the pH
`was varied from slightly acidic to slightly alka(cid:173)
`line, two formulations with different amounts
`of ethylenediaminetetraacetic acid (EDT A)
`included as a chelating agent, two formulations
`with different amounts of Tween 80 included
`as a surfactant, and two formulations with
`
`28 BioPharm International www.biopharmlnternational.com April 2010
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 3
`
`

`

`Extractables & Leachables
`
`either mannitol or trehalose added as bulking
`agents. Commercially available glass vials and
`rubber stoppers were used in the study.
`Separate glass vials were filled with one
`of the 11 formulations listed in Table 1 and
`crimp-sealed with rubber stoppers. The
`stoppered vials containing various formu(cid:173)
`lations were inverted and then stored at
`40 °c and 75% RH. After one month, the
`formulations were tested for volatile, semi(cid:173)
`volatile, and nonvolatile leachables using
`headspace gas chromatography-mass spec(cid:173)
`trometry (GC-MS), direct injection GC-MS,
`and gas chromatography-ultra violet mass
`spectrometry LC-uv.:.Ms, respectively. The
`portions of each sample to be analyzed
`were transferred to headspace GC vials and
`high performance liquid chromatography
`(HPLC) vials without further treatment
`for headspace GC-MS and LC-UV-MS
`analysis, respectively. For direct injection
`GC- MS analysis, the samples were back
`extracted into methylene chloride, which
`is a more suitable solvent for this analysis.
`Control samples (portions of each formula(cid:173)
`tion that were stored in glass volumetric
`flasks at S °C for the same time duration as
`the samples) also were analyzed.
`To correlate the leachables with the
`extractables from the stoppers, the stoppers
`were extracted with water and isopropanol
`and analyzed with the same headspace
`GC-MS, direct injection GC-MS, and LC(cid:173)
`UV-MS conditions. For headspace GC-MS
`analysis, the stoppers were extracted in
`headspace GC vials at 90 °C for 24 h, and
`the vials were directly used for volatile
`extractables analysis, without opening the
`vial cap. All volatile extractables, including
`water insoluble extractables, were detect(cid:173)
`able using this approach. For direct injec(cid:173)
`tion GC-MS and LC-MS analyses, the
`stoppers were extracted by refluxing with
`water and isopropanol for 16 h. Tables 2-4
`show the chromatographic conditions used
`for each analysis.
`RESULTS AND DISCUSSION
`Headspace GC-MS Analysis
`Extractables Evaluation
`The water extracts of stoppers yielded seven
`extractables peaks (Figure 1). Six of the seven
`peaks were identified as 2-methylpentane
`(4.24 min), 3-methylpentane (4.62 min), hex-
`
`Table 2. Headspace GC-MS chromatographic conditions
`Column
`
`DB-VRX 30 m x 0.25 mm, 1.4 µm film thickness or
`equivalent
`Oven program
`35 •c (15 min), 10 •c/min, 220 •c (10 min)
`Injector port temp
`220 ·c
`Flow (He)
`1 ml/minute (constant flow)
`Split ratio
`5:1 (split flow column flow)
`Run time
`43.5 min
`Mass range
`35-350 amu
`Ionization
`El
`75 ·c
`Oven temperature
`GC-MS: gas chromatography-mass spectrometry
`
`Table 3. GC-MS chromatographic conditions
`Instrument
`Agilent 5973 GC-MSD
`GC column
`HP-5MS, 30 m, 0.25 mm id, 0.25 µm film thickness or
`equivalent
`Flow
`1.0 ml/min helium
`Temperature program
`40 •c (2 min), 15 °C/min, 320 •c (20 min)
`Mode
`Splitless
`Purge time
`0.5 min
`280 ·c
`Inlet temperature
`1.0 µl
`Injection volume
`Mass range
`35-650 amu
`GC-MS: gas chromatography-mass spectrometry
`
`Table 4. LC-MS chromatographic conditions
`UPlC system
`Waters Acquity
`MS detector
`Waters Acqulty TOD
`Mobile phase A
`Water
`Mobile phase B
`Acetonitrile
`Column
`Waters Acquity UPlC BEH C18, 2.1 x 50 mm, 1.7 µm
`Flow rate
`0.8 ml/min
`60 ·c
`Column temperature
`UV wavelength
`280 nm
`Injection volume
`5 µl
`Gradient
`Water and acetonitrile gradient
`Ionization mode
`APCI (-) and APCI (+)
`Mass range
`150-1,200 amu
`LC-MS: liquid chromatography-mass spectrometry
`
`ane (5.14 min), methylcyclopentane (6.24
`min), cyclohexane (7.93 min), and butylated
`hydroxytoluene (BHT, 33.26 min). The peak
`at 28.60 min was not identified; the GC-MS
`library search and manual spectral interpre(cid:173)
`tation did not produce a good match or a ten(cid:173)
`tative identification. The amounts for each
`peak are summarized in Table 5. The peak
`at 28.60 min was quantitated using cyclo-
`
`April 2010 www.blopharmlnternational.com BioPharm International 29
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 4
`
`

`

`Figure 1. Headspace GC-MS chromatogram of water extracts
`of rubber stoppers
`
`400.000
`
`340.000
`
`320,000
`
`5.2,4;1 --
`-260,000
`
`300,000
`
`C:
`
`e!
`"' "C
`C:
`:::,
`..0
`<(
`
`240,000
`
`22?,0IIO
`
`200,000
`
`180,000
`
`160,000
`
`140.000
`
`U0.000
`
`100.000
`
`60.000
`
`80,000 •
`......
`
`20,000
`
`Methylcyclopentane
`
`~
`
`3-methylpentane
`
`Hexane
`
`2-methylpentane
`
`BHT
`
`Unknown
`
`29.100
`
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`21.00
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`
`16.00
`
`28.00
`
`)Q.00
`
`~
`32.00
`
`Time(min)
`
`GC-MS: gas chromatography-mass spectrometry; BHT: butylated hydroxytoluene
`
`hexane as the surrogate standard.
`Because the relative response fac(cid:173)
`tor of the unknown peak against
`cyclohexane is not determined,
`the amount reported for this peak
`is only considered a semi-quanti(cid:173)
`tative estimate.
`
`Leachables Evaluation
`Six leachables peaks were observed
`in the headspace GC- MS analysis
`from the formulations : 2-meth(cid:173)
`ylpentane, 3-methylpentane,
`hexane, methylcyclopentane,
`cyclohexane, and BHT. These
`leachables peaks correlate to stop(cid:173)
`per extractables . The unknown
`extractables at 28.60 min were not
`
`observed as leachables. This may
`be because of insolubility of the
`compound in the aqueous media.
`The amount of leachables in dif(cid:173)
`ferent formulations is summarized
`in Table 5. The effect of various
`formulation ingredients on the
`leachables profile is dis.cussed in
`detail below.
`
`Phosphate Buffer with or
`without Glycerol Co-Solvent
`Five leachables peaks were
`observed in the pH 6.8 phosphate
`buffer: 3 methylpentane, hexane,
`methylcyclopentane, cyclohexane,
`and BHT. All of the peaks were
`very small. Methylcyclopentane
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 5
`
`

`

`Extractables & Leachables
`
`Table &. ·ouantitation of extractables/leachables with headspace GC-MS
`I I
`
`- - , -
`
`- -
`
`- -
`
`- - - -
`
`-
`
`t~
`Formulation ,
`' 2-methyl-
`i
`pentane
`
`I
`
`-
`
`! 3-methyl-
`,
`pentane
`
`Compound and amount (ng/vial or ng/stopper)
`I
`'
`
`Hexane
`
`Methylcyclo-
`pentane
`
`•
`
`I
`
`-
`
`Cyclohexane
`
`-
`
`- - •
`
`-
`
`-
`
`- -
`
`J
`
`BHT
`
`: Unknown
`
`182.6
`
`805.7
`
`434.1
`
`Stopper
`extract
`ND
`4.8
`3.6
`1
`ND
`4.9
`4.9
`2
`1.8
`4.0
`4.4
`3
`ND
`ND
`1.5
`4
`ND
`3.8
`4.2
`5
`ND
`3.5
`1.9
`6
`ND
`2.4
`2.7
`7
`8.9
`8.5
`2.6
`8
`6.8
`37.7
`28.6
`9
`ND
`4.8
`2.5
`10
`ND
`ND
`ND
`11
`GC-MS: gas chromatography-mass spectrometry; BHT: butylated hydroxytoluene
`
`1420.9
`
`320.9
`
`12.5
`13.1
`11.2
`5.4
`11.9
`10.3
`7.4
`21.6
`53.8
`11.0
`2.5
`
`2.3
`2.1
`1.4
`ND
`1.2
`1.2
`ND
`2.6
`7.1
`ND
`ND
`
`26.1
`
`3.0
`3.6
`2.9
`ND
`1.0
`2.3
`2.7
`ND
`ND
`ND
`ND
`
`40.9
`
`ND
`ND
`ND
`ND
`ND
`ND
`ND
`ND
`ND
`ND
`ND
`
`was the largest leachables peak, probably
`because it is more soluble in aqueous media
`than the other compounds. The addition
`of 2% glycerol in the phosphate buffer as a
`co-solvent did not significantly affect the
`amount of leachables observed.
`
`Formulation pH
`Formulation pH had an impact on leachables.
`The neutral pH (pH 6.8) provided slightly
`lower amounts of leachables compared to the
`slightly acidic (pH 5.0) and the slightly alka(cid:173)
`line (pH 8.2) formulations (Figure 2). The
`acidic and alkaline formulations had similar
`leachable profiles.
`
`EDTA Chelating Agent
`Based on the leachables profiles of formula(cid:173)
`tions containing 0, 0.1, and 0.5 mM EDTA,
`EDTA did not significantly affect the amount
`of organic leachables.
`
`The impact of different
`formulation excipients does
`not always affect all
`leachables components to
`the same extent.
`
`in formulations without Tween 80. This is
`because the surfactant increases the solubil(cid:173)
`ity of these compounds in the aqueous for(cid:173)
`mulation (Figure 3). However, the addition
`of Tween 80 did not affect the amount of
`leached BHT because no BHT was detected in
`the formulations with 0.5% or 0.1% Tween 80.
`
`Tween SO
`The excipient Tween 80 had the most sig(cid:173)
`nificant effect on leachables compared to
`other formulation ingredients. The addi(cid:173)
`tion of Tween· 80 significantly increased the
`leached amount of 2-methylpentane, 3-meth(cid:173)
`ylpentane, hexane, methylcyclopentane,
`and cyclohexane. The leached amounts of
`these compounds in the formulation with
`0.5% Tween 80 is five to 10 times higher than
`
`Bulking Agents: Sucrose, Mannitol, and Trehalose
`The use of bulking agents affected the
`amount of leachables observed. Higher
`amounts of leachables were observed in the
`formulation with 7% mannitol compared to
`the formulation with 7% trehalose, in which
`few leachables were observed (Figure 4). The
`formulation with 7% sucrose had leachables
`levels between those seen in formulations
`with mannitol and trehalose.
`
`32 BioPharm International www.biopharminternatlonal.com April 2010
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 6
`
`

`

`Extractables & Leachables
`
`Figure 2. Headspace GC-MS chromatograms of formulations with
`different pH conditions (the large peak at around 33.1 min is from the
`formulations, not a leachable peak)
`
`.,,,.
`.....
`
`Tlmefmlnl
`
`GC-MS: gas chromatography-mass spectrometry; BHT: butylated hydroxytoluene
`
`Figure 3. Headspace GC-MS chromatograms of formulation with and
`without Tween 80
`
`2-methylpentane
`
`35,000 1
`
`30,oooj
`
`1
`
`•
`
`25,000
`
`j ~,000
`
`Direct Injection GC-MS Analysis
`Extractables
`There was only one semivolatile extractable
`peak observed from the water extracts. The
`peak was identified as BHT and is very small
`because of its limited solubility in water.
`There were many extractables peaks
`observed in the isopropanol extracts (Figure
`5). The two dominant peaks are BHT (12.18
`min) and an unknown hydrocarbon at 13.66
`min. Other compounds observed were pal(cid:173)
`mitic acid (15.35 min) and stearic acid (16.62
`min). All other peaks appear to be various
`hydrocarbons.
`
`Leachables
`Leachables were determined by comparing
`the stressed formulations to the formula(cid:173)
`tion blanks and the stopper extractables
`profile. There is no appreciable interference
`from the formulation blanks for the main
`extractables peaks . The only leachables
`observed was BHT, which was reported
`in formulation 3 (pH 5.0), formulation 6
`(0.1 mM EDTA), and formulation 7 (0.5
`mM EDTA). This result was consistent with
`headspace GC-MS data. The BHT peak is
`very small. No other peaks observed in the
`isopropanol extract were observed in any
`of the formulations . This can be explained
`by the insolubility of these compounds in
`the aqueous formulations .
`There were two additional peaks observed
`in formulation 1 (pH 6.8 phosphate buf(cid:173)
`fer) and formulation 2 (phosphate buffer
`with 2% glycerol). The first elutes at 11.27
`min and the second elutes at 12.11 min.
`The peak at 12.11 min is tentatively identi(cid:173)
`fied as 2,4-t-butylphenol, while the peak at
`11.27 min was not identified. For all other
`formulations, the peak at 11.27 min was not
`observed. For the peak at 12.11 min, there
`is a co-eluting interference peak from all
`the formulation blanks except formulations
`1 and 2, so it is not conclusive if the peak
`observed in formulations 1 and 2 was also
`present in those formulations. The source of
`the two peaks requires further evaluation.
`
`LC-UV-MS Analysis
`No extractables were observed with LC-MS
`from the water extracts of rubber stoppers.
`The isopropanol extract of the stoppers con(cid:173)
`tained BHT (5.02 min) and three fatty acids
`
`15,000
`
`GC-MS: gas chromatography-mass spectrometry
`
`Tlme(mln)
`
`(myristic acid, 5.21 min; palmitic acid, 5.80
`min; and stearic acid, 6.30 min).
`The leachables were evaluated by com(cid:173)
`paring the stability samples to the formu(cid:173)
`lation blanks and the stopper extractables
`profile. There is no appreciable interfer(cid:173)
`ence from the formulation blanks for the
`main extractables peaks. There are no
`leachables observed in the LC-MS analysis
`of the 11 formulations. The observation
`of BHT in the GC-MS analysis but not in
`the LC-MS analysis is likely because of
`
`April 2010 www.blopharmlnternatlonal.com BioPharm International
`
`33
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 7
`
`

`

`Extractables & leachables
`
`Figure 4. Headspace GC-MS chromatograms of formulations with
`different bulking agents
`
`3-methylpentane
`
`Hexane
`
`---
`
`UD
`Time(mln)
`
`I.SO
`
`1M
`
`1~
`
`GC-MS: gas chromatography-mass spectrometry
`
`Figure &. GC-MS chromatogram of isopropanol extracts of rubber
`stoppers
`
`5,500,000
`
`5,000,000
`
`4,500,000
`
`4,000,000
`
`3,500,000
`
`3,000,000
`
`2,500,000
`
`2,000,000
`
`1,500,000
`
`1,000,000
`
`~ .. "tJ
`.c <
`
`C:
`::,
`
`.09
`
`500,000
`
`1535 16.f t 17.98 19.03 20.0020.91 2192
`11.31
`I
`I
`,· 1
`I
`. •
`11-~~
`• .,.,.., 1 .J. . • ,, , 1 "•i · "•i•IJ, .,
`I " l •
`,
`I I,
`1 .. ,, .. I
`,
`9.00 10.00 11.00 12.00 13.00 14.00 15.00 16.00 17.00 18.00 19.00 20.00 21.00 22.00
`Time(mln)
`
`BHT
`
`12.18
`
`/J:::,:,
`I I l
`
`of note is Tween 80, which signifkantly
`increased the amount of leachables as com(cid:173)
`pared to other formulations.
`• The impact of different formula(cid:173)
`tion excipients does not always affect
`all leachable components to the same
`extent. For example, Tween 80 signifi(cid:173)
`cantly increased the leachables 2-meth(cid:173)
`ylpentane, 3-methylpentane, hexane,
`inethykyclopentane, and cyclohexane.
`However, it does not appear to have an
`impact on BHT. The bulking agents used,
`such as sucrose, mannitol, and treha(cid:173)
`lose, lead to different leachables levels,
`with trehalose being a superior bulking
`agent from the standpoint of minimizing
`leachables.
`• The volatility of the extractables in the
`stopper material and the solubility of
`those compounds in the formulations are
`the determining factors for leachables.
`All leachables observed in this study
`were volatile compounds and observed in
`headspace GC-MS analysis. Volatile com(cid:173)
`pounds migrate more easily than nonvol(cid:173)
`atile compounds and are more likely to
`leach out into formulations. The relative
`solubility of leachables in formulation is
`directly proportional to the amount that
`will be present in the formulation as a
`leachable from the stopper material.
`• This study demonstrates that biophar(cid:173)
`maceutical excipients, formulation stor(cid:173)
`age conditions, and to some extent the
`container/closure material, can have a
`significant impact on the success of a
`potential commercial formulation.
`A well-designed study can help a bio(cid:173)
`pharmaceutical company build quality and
`efficiency into its biopharmaceutical prod(cid:173)
`uct design and in-process development and
`commercialization. This approach provides
`a knowledge base for a better selection
`of product formulation components, with
`respect to leachables.
`Scientific understanding based on data
`obtained from this particular study can be
`used to support other pharmaceutical prod(cid:173)
`uct development processes. For instance, for
`different rubber stopper materials or other
`types of container/closure materials, the
`potential leachables profile can be predicted
`based on its extractables profiles and the
`intended formulation and excipients regime
`
`GC-MS: gas chromatography-mass spectrometry; BHT: butylatad hydroxytoluene
`
`the increased sensitivity of GC-MS over
`LC-MS.
`CONCLUSION
`Eleven different typical formulations of
`biopharrnaceutical products were evaluated
`to determine their effect on the leachables
`profile of a common rubber stopper after
`storage at 40 °C, 75% RH for one month.
`The key findings are as follows:
`• Different formulation excipients have a.
`significant impact on leachables. Especially
`
`34 BioPharm International www.biopharminternational.com Aprll 2010
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 8
`
`

`

`Extractables & Leachables
`
`- - - - - - - - - - -
`
`Figure 8. LC-MS [APCI (-)] chromatogram of isopropanol extracts of rubber stoppers
`
`LABS\plccolrl
`MSMS-39
`Sel0350
`
`RAPRASel03a
`
`uo
`
`IPA Stopper Extract
`
`1: Scan AP(cid:173)
`TIC
`3.02e7
`
`Palmltlrn~~
`
`BHT~
`
`5.02
`I
`
`5.10 i
`
`Myristic acid
`
`0
`
`0.50
`
`1.00
`
`1.50
`
`2.00
`
`2.50
`
`3.00
`
`3.50
`
`4.00
`
`4.50
`
`5.00
`
`5.50
`
`6.00
`
`6.50
`
`7.00
`
`7.50
`
`8.00
`
`8.50
`
`9.00
`
`-
`
`• ¥' ,;'
`
`. _ . . . _ ;.- . }• •• • •
`
`5-?1
`
`e.pa
`
`LC-MS: liquid chromatography-mass spectrometry; BHT: butylated hydroxytoluene
`
`_Time(min)
`
`View publication stats
`View publication stats
`
`with greater accuracy for the model systems
`studied. The knowledge and understanding
`accumulated will in turn aid the product
`development process and make it possible
`to develop safer and more efficacious medi(cid:173)
`cines faster.
`Based on our study, changes in the com(cid:173)
`ponents in a biopharmaceutical formula(cid:173)
`tion can lead to appreciable changes in
`the leachable profiles. A thorough under(cid:173)
`standing of how various formulation com(cid:173)
`ponents may affect a leachables profile
`can aid in selecting appropriate formula(cid:173)
`tion constituents based on the container/
`closure materials and storage conditions.
`Such an understanding overcomes the lim(cid:173)
`itations of a traditional selection process
`and allows the early adoption of a risk(cid:173)
`based approach and QbD at the beginning
`of product development. Consequently,
`drug development duration can be short(cid:173)
`ened significantly while product quality,
`safety, and efficacy are maintained. •
`
`REFERENCES
`1. Yu X, Wood D, Ding X. Extractables and
`leachables study approach for disposable
`materials used In bioprocessing. BioPharm
`Int. 2008;21(2):42-51.
`2. Product Quality Research Institute.
`Leachables and Extractables Working Group.
`Safety Thresholds and Best Practices for
`Extractables and Leachables in Orally Inhaled
`and Nasal Drug Products. Arlington, VA; 2006.
`3. US Food and Drug Administration. Guidance
`for Industry: Container closure systems for
`packaging drugs and biologics, chemistry,
`manufacturing, and control documentation.
`Rockville, MD; 1999 May.
`4. Europeans Commission (Enterprise
`Directorate General). EMEA guideline on
`plastic immediate packaging materials.
`Brussels, Belgium; 2005 May.
`5. United States Pharmacopeia (USP). General
`chapter <661> containers-polytethylene
`containers. Rockville, MD.
`6. USP. General chapter <381> elastomerlc
`closure for Injections. Rockville, MD
`7. European Pharmacopeia. Chapter 3. Materials
`for containers and containers. Strasbourg,
`France.
`
`36 BloPharm International www.blopharmlnternatlonal.com April 2010
`
`Opiant Exhibit 2319
`Nalox-1 Pharmaceuticals, LLC v. Opiant Pharmaceuticals, Inc.
`IPR2019-00685, IPR2019-00688, IPR2019-00694
`Page 9
`
`