157
11
Formulation Development Concepts
11.1 Preformulation
Preformulation is often used to describe the physical and
chemical characteristics of substances and mixtures used
in pharmaceutical products. It is part of the research and
development (R&D) unit of the pharmaceutical industry.
Preformulation is used synonymously with physical phar-
macy. It is considered to be part of a larger science known
as pharmaceutics. Both preformulation and formulation are
parts of pharmaceutics, and their application in formula-
tion development in the industry is known as the industrial
pharmacy. Characterization of chemicals can be done on
the molecular, particulate, and bulk levels (Brittain
1999). Nowadays, during the R&D stage of the pharmaceu-
tical product’s progress, scientists depend heavily on using
instrumentation to characterize or identify a chemical or a
final product. Instruments used in research have become
extremely complicated and similar to a piano and a musi-
cian’s ability to play it, the same instrument can produce
bad, good, or excellent data, depending on how it is being
run (Scarlett 2003). The biopharmaceutical industry alone
has reported an expenditure of $173 billion on R&D in 2017
(Hilleke and Kars 2019). Because of the importance of the
characterization and the production of drug products, the
pharmaceutical and biotechnology industries have become
the number one target for cybercriminals worldwide
(Douthwaite 2020).
According to recent estimates, only one out of 10,000 com-
pounds evaluated becomes an approved pharmaceutical
product for a new active pharmaceutical ingredient (API).
During this process that lasts on average between 10 and
14 years, an excess of $1 billion is being spent on the pro-
duct’s development (Brown et al. 2019). For New Drug
Application (NDA) and Biologics License Application
(BLA), the Food and Drug Administration (FDA) has
approved an average of 100 NDAs and BLAs in the period
between 2010 and 2018, 38% of which are new molecular
entity (NME) types. The NMEs were either a single active
(BLA and NDA), a biosimilar (BLA), or a combination type
Integrated Pharmaceutics: Applied Preformulation, Product Design, and Regulatory Science, Second Edition.
Antoine Al-Achi, Mali Ram Gupta, and William Craig Stagner.
© 2023 John Wiley & Sons, Inc. Published 2023 by John Wiley & Sons, Inc.
Companion website: www.wiley.com/go/Al-Achi/IntegratedPharmaceutics
(NDA) (Bassart 2020). Many techniques and instrumenta-
tions are needed during the discovery and preformulation
phase to establish a baseline characteristics profile (physi-
cal/mechanical, chemical, and biological) for the new API.
Preformulation studies are conducted to examine the com-
patibility of the API with various common excipients.
Among the physicochemical properties of the API that are
established during preformulation investigations include
melting point/freezing point, porosity, bulk volume, bulk
density, bulkiness, equivalent diameters for powder parti-
cles, specific surface, angle of repose, powder flowability, dif-
ferential scanning calorimetry (DSC), thermogravimetric
analysis (TGA), viscosity, viscoelasticity parameters, zeta
potential and physical stability parameters (creaming/sedi-
mentation and flocculation) for dispersed systems, specific
gravity/density, solubility characteristics in water and
various solvents, dissolution profile, disintegration charac-
teristics of the dosage form, partition coefficient, pH, pKa,
surface tension/interfacial tension, Karl Fischer (KF) titra-
tion for water content, water sorption analysis (dynamic
vapor sorption; DVS), X-ray diffraction analysis (XRD,
PXRD, or XRPD), polymorphism characteristics/metastable
crystals, amorphous forms, and solvate forms, Raman spec-
troscopy, infrared spectroscopy and imaging, Fourier-
transform infrared spectroscopy (FTIR), among others. For
example, Stagner et al. (2020) investigated the factors
involved in the compressibility, compactibility, and tabletibil-
ity of flufenamic acid: nicotinamide cocrystal (flufenamic
acid is an analgesic, anti-inflammatory, and antipyretic
agent; nicotinamide is a water-soluble vitamin B-3). The
cocrystal formation improves the aqueous solubility of flufe-
namic acid (Fabian et al. 2011). They aimed to study the
latent effect of storage conditions and material compression
parameters on tablet mechanical stress. Their study used var-
ious techniques that included morphological studies on the
cocrystal using scanning electron microscopy, the tablet’s
mechanical stress measurement, Shapiro analysis, and
Heckel analysis to describe initial particle arrangement and
fragmentation during compression. Parameters obtained
158 11 Formulation Development Concepts
from the analysis were fitted to multivariate analysis and
latent variable statistical methodologies such as explora-
tory factor analysis, principal component analysis, and
principal component regression. The statistical methods
were used to describe complex hidden main, interaction,
and quadratic effects of the storage conditions and the
compression parameters on the cocrystal tablet mechani-
cal strength (Stagner et al. 2020). Interestingly, as recently
as 2019, six out of ten product applications in the United
States were given accelerated approval via priority review,
fast track, or breakthrough (Thomas, 2020b). For this
acceleration to be accomplished, pharmaceutical scien-
tists have to produce quality measurements in a shorter
allotted time, often 18–24 months earlier than typical
schedules (Browne and Savla 2019). Moreover, the devel-
opment of hybrid products under the European Union
guidelines or their corresponding regulations in the US
505(b)(2) (see Chapter 22 for section 505) require higher
technological innovations to achieve the desired better
therapeutic effectiveness. It was estimated that one out
of each two New Drug Applications were subjects to 505
(b)(2) in 2017 (Anonymous 2018). This is in agreement
with the pharmaceutical industry is becoming more and
more patient-centric in its approach to designing newer
dosage forms (Thomas, 2020b).
The solubility of chemicals in water is one of the main
challenges that hinder the chemical process of becoming
pharmaceutical products. Inclusion complexes formed
between the chemical agent and a carrier system can
render the chemical more water-soluble. Cyclodextrins
are often used to enhance the water solubility of various
substances by creating an inclusion complex. An example
of this formulation strategy is the use of a cyclodextrin ini-
tially discovered by a group of scientists (Professor Valen-
tino Stella and Dr. Roger Rajewski) at the University of
Kansas (Sarah Motter 2020). This cyclodextrin is known
as CaptisolÒ (sulfobutylether-cyclodextrin; Ligand Phar-
maceuticals) that combines with various drugs to form
water-soluble complexes (to date, 15 products have been
approved by various world regulatory agencies using Cap-
tisolÒ) (Pipkin et al. 2020). An example of a CaptisolÒ-
containing pharmaceutical product is Veklury® (remdesi-
vir) by Gilead. The drug is an antiviral agent and suffers
from poor solubility in water. The use of CaptisolÒ in this
formulation in the ratio of 30 : 1 (CaptisolÒ:API) as a lyo-
philized powder or 60 : 1 as a solution for injection has
facilitated its complete dissolution in water (Pipkin
et al. 2020).
Al-Achi et al. (2019) examined the enhancement of aque-
ous solubility of the antihypertensive drug nifedipine by
complexing it with hydroxypropyl-β-cyclodextrin. Physical
characterization of the complex was accomplished by mate-
rial science methodologies (TGA-DSC, attenuated total
reflection analysis (ATR), and PXRD analysis). The ATR
is a FTIR technique. The following is a synopsis of their
work (Al-Achi et al. 2019):
Inclusion complex can enhance the drug solubility,
stability, and masking smell. Cyclodextrin (CD) is a
commonly used host molecule for inclusion, with
three main types of CD, exist, α, β, and γ. They are
classified based on their chemical structure. The
derivative of β-CD, (Hydroxypropyl-β-Cyclodextrin;
P-β-CD), is widely used in pharmaceutical formula-
tions. HP-β-CD works by a dynamic equilibrium
process for inclusion. Nifedipine is a Class II drug
(Biopharmaceutical Classification System) that
has poor solubility in water, and high permeability
through the cellular membrane. The Phase-
solubility profile of nifedipine/HP-β-CD complex
showed an AL type (according to Higuchi and Con-
nors’ method) indicating a complex ratio of 1 : 1. The
objective of this study was to investigate the pH
effect on the inclusion process of nifedipine/HP-
β-CD. The results of this study showed that ioniza-
tion pH conditions improved nifedipine solubility
in water to a limited extent however it was not as effi-
cient as the unionization pH conditions since union-
ization conditions produced higher Complexation
Efficiency (CE) and the stability constant (K1 : 1)
value. Therefore, maintaining the drug in a union-
ized form is perhaps a more efficient way of produ-
cing inclusion complex with HP-β-CD. In order to
characterize the inclusion complex thus formed
between the drug and HP-β-CD, a solvent evapora-
tion method was used to prepare the complex which
was compared to a physical mixture, pure drug, and
pure HP-β-CD. Significant differences were found to
exist between the inclusion complex and all the
other groups based on the TGA, DSC, ATR, and
PXRD analysis.
The results from the TGA, DSC, ATR, and PXRD are
shown below, respectively (Al-Achi et al. 2019).
TGA-DSC test: After TGA run, the four samples
were subjected to decomposition temperatures.
Based on the temperature data the DSC maximum
temperature should not be set higher than the
decomposition temperature. The inclusion complex
had completely turned to amorphous form which
was similar to that of pure HP-β-CD, whereas the
DSC curve for pure nifedipine is a typical crystalline
form curve which had a sharp endothermic peak at
170 C. The curve of the physical mixture had a
smaller endothermic peak which indicated the drug
was partially included into the HP-β-CD.
ATR test: The inclusion complex has a strong peak
in 1678 cm−1, this peak specifically reflects -C=O
 11.1 Preformulation 159

group, where nifedipine has two ester groups which
are lipophilic that can be captured inside of the cav-
ity of cyclodextrin, the outside of the cavity is hydro-
philic that have more -OH groups, it has more peak
intensity than pure drug in 3327 cm−1. Additionally,
for HP-β-CD, hydroxypropyl group connects the
cyclodextrin by an oxygen atom which forms an
ether group. Once the drug is included inside, the
inclusion complex has a stronger peak compared
to the pure drug in 1022 cm−1. This was specifically
demonstrated for the ether (C-O-C) group as seen in
the pure HP-β-CD spectra.
PXRD test: A sharp contrast was observed between
the inclusion complex and pure drug. The pure drug
showed a typical crystalline form that had many
sharp peaks; for inclusion, the crystal form almost dis-
appeared and the position of the two main broaden
peaks at the same angle compared with pure cyclo-
dextrin suggests that nifedipine was included into
the HP-β-CD. For the physical mixture, it has few
sharp peaks as compared to pure drug and the general
shape of the graph is more similar to pure HP-β-CD.
From TGA-DSC, ATR, PXRD studies, all the results
indicate nifedipine molecule can form an inclusion
complex with cyclodextrin using solvent evaporation
method in the presence of 40% acetonitrile as the sol-
vent. (Al-Achi et al. 2019).

102
Pure HPBCD
Physical mixture
Pure Nifedipine
Inclusion complex
100

98

96
Weight (%)

94

92

90

88
0 50 100 150 200 250 300 350
Temperature (°C) Universal V4,5A TA

TGA: Source: Al-Achi et al. (2019) / Ocimum Scientific Publishers Pty Ltd / CC BY 4.0.

2 Inclusion complex
Pure HPBCD
Physical mixture
Pure Nifedipine

0
Heat flow (W/g)

–2

–4

–6
0 50 100 150 200 250 300
Exo Up Universal V4.5A TA In
Temperature (°C)

DSC: Source: Al-Achi et al. (2019) / Ocimum Scientific Publishers Pty Ltd / CC BY 4.0.
 Drug with inclusion complex
101
100
95

90

85 3325.94 cm–1

1679.21 cm–1
80 3327.38 cm–1 1349.22 cm–1
1431.79 cm–1 829.07 cm–1
%T

1495.61 cm–1
75 1226.91 cm–1 793.63 cm–1
1348.86 cm–1 711.95 cm–1
1678.32 cm–1 744.47 cm–1
70
1119.98 cm–1
65 1225.52 cm–1
1021.88 cm–1

60
1022.80 cm–1
55
54
4000 3500 3000 2500 2000 1500 1000 650
cm–1
Name Description
Complex label 011 by PERC Date Monday, May 18 2015
Pure drug label 014 by PERC Date Monday, May 18 2015

ATR: Source: Al-Achi et al. (2019) / Ocimum Scientific Publishers Pty Ltd / CC BY 4.0.

100 Fingerprint zone of NI COM CD

95

90

85

80 1679.21 cm–1 1349.22 cm–1

1431.79 cm–1 1349.24 cm–1
1495.61 cm–1 829.07 cm–1 744.47 cm–1
75 1527.38 cm–1 1348.88 cm–1 1226.95 cm–1 793.63 cm–1
%T

1678.32cm–1 711.95 cm–1

70

65 1119.98 cm–1
1225.52 cm–1 1021.48 cm–1
60

55
1022.54 cm–1
50
49
1755 1600 1400 1200 1000 800 598
cm–1
Name Description
Pure drug label 014 By PERC Date Monday, May 18 2015
Complex label 011 By PERC Date Monday, May 18 2015
Pure CD label 012 By PERC Date Monday, May 18 2015

ATR: Source: Al-Achi et al. (2019) / Ocimum Scientific Publishers Pty Ltd / CC BY 4.0.

1500

1000

500

0
10 20 30 40 50 10 20 30 40

Inclusion complex Physical mixture

1000

500

0
10 20 30 40 50 10 20 30 40

Pure drug Pure HP-β-CD

PXRD: Source: Al-Achi et al. (2019) / Ocimum Scientific Publishers Pty Ltd / CC BY 4.0.

On the biological front, preformulation testing is also
important to characterize the biological profile of the
API when needed. For example, in evaluating novel anti-
microbial peptides (bacteriocin “Garvicin KS; GarKS”
was identified from Lactococcus garvieae KS154), Thapa
et al. (2020) utilized various techniques and instrumenta-
tions in preformulation testing of this new peptide-
antibiotic. These techniques included the use of
high-performance liquid chromatography instruments,
determination of the physical stability of the peptides
by adsorption and aggregation studies, determination of
the chemical stability of the novel drug, determination
of the antimicrobial activities of the peptides, and deter-
mination of fibroblast cell viability and proliferation (cell
viability and proliferation studies). In addition, statistical
analysis methods were used to detect statistical signifi-
cance in the data collected. Among the instruments that
were utilized in the preformulation studies were (Thapa
et al. 2020):
•
UV–Vis spectrophotometer (UV-2401 PC, Shimadzu,
Kyoto, Japan).
• HPLC system from Shimadzu Corporation (Kyoto, Japan)
that consisted of an auto-injector SIL-9A, a Prominence
LC-20AD pump, a Chromatopac C-R5A integrator, and
a Prominence SDP-10A UV detector. A reversed-phase
column (Inertsil ODS-SP; 5 μm, 4.6 × 150 mm; GL
Sciences Inc, Tokyo, Japan) equipped with a Guard Col-
umn (Inertsil ODS-SP; 5 μm, 4.0 × 10 mm; GL Sciences
Inc, Tokyo, Japan).
• For the aggregation studies: A dynamic light scattering
instrument (ZetaSizer, Malvern Instruments, Worcester-
shire, U.K.)
•• For the adsorption studies: HPLC method.
For the chemical stability studies: HPLC method.
• The antimicrobial activities of individual and combina-
tion peptides were evaluated in methicillin sensitive S.
Aureus MSSA; ATCC 6538) and methicillin resistant S.
aureus (MRSA, Xen 31; parental strain ATCC33591).
The turbidity (OD600) of bacterial growth was analyzed
to determine the (minimum inhibitory concentration)
MIC value for each peptide.
• Cell culture studies: Adult human dermal fibroblasts
(ATCC®PCS-201–012™) were obtained from ATCC
(Manassas, VA, USA) and cultured in DMEM supple-
mented with 10% FBS, 100 U/mL penicillin, 100 μg/mL
streptomycin, and 250 μg/mL fungizone (referred to as
“complete DMEM media”) in tissue culture flasks. The
cells were incubated at 37 C in a humidified atmosphere
of 5% CO2 and sub-cultured twice a week (cells in
passages 3–10 were used for the experiments). (Thapa
et al. 2020)
11.1 Preformulation 161
Cell culture research is fast developing with new
instrumentation for studying cells in real time. Systems
such as the ones developed by Sartorius/Essen BioScience
(Essenbio.com/oncology; Ann Arbor, Michigan) (e.g.
IncuCyte® S3 Live-Cell Analysis System) allow research-
ers in the field of oncology to study cancer cells in their
living environment noninvasively. Phenomena such as
apoptosis, angiogenesis, cell invasion/migration, and anti-
body internalization may be detected by these devices. In
addition to the cell culture studies, in vivo studies are also
conducted in experimental animals, including toxicologi-
cal studies where applicable. Thus, it is of importance that
the pharmaceutical scientist has a broad comprehension
in various process and production techniques including
a thorough knowledge in statistical analysis (Chapter 10).
Also, pharmaceutical scientists should be familiar with the
various current pharmacopeia requirements in particular
those of the United States, European Union, and Japan,
in order to meet pharmacopoeia standards (Wiggins and
Albanese 2020). Another consideration is that related to
the mode of pharmaceutical production. Although other
industries such as aerospace, car making, food, and metal
products have been employing continuous manufacturing
processes for decades (Smith 2020), the pharmaceutical
industry is slowly moving away from one-batch produc-
tion (the traditional batch processing) to continuous pro-
duction, which is recognized to provide a better product
quality, according to the FDA. Applications such as con-
tinuous production offer new challenges to formulation
scientists, especially those in biopharmaceutical industry
in regard to process and equipment robustness operating
over several days to several weeks at a time (e.g. tubing
and mechanical pump degradation) (Brooks 2020).
Although continuous processing is financially sound,
the industry still perceives that this way of production
comes with a “higher regulatory risk” tag (Smith 2020).
The following brief discussion describes results from
preformulation studies that were carried out on human insu-
lin (HINS) for the purpose of administering HINS orally or
transdermally. These routes of administration for HINS
remain unattainable for various natural obstacles that govern
the handling of HINS by the body: in the GI tract, the presence
of proteolytic enzymes; transdermally, HINS has high molec-
ular weight and hydrophilic which render the hormone inca-
pable to undergo spontaneous diffusion through the skin.
Faced with these challenges, Al-Achi and collaborates
reported the following findings on HINS (available from
recombinant DNA technology) (Humulin®R; Eli Lilly; Indi-
anapolis, IN; 100 U/mL) from in vitro, cell culture, in situ, and
whole experimental animal studies. It should be noted that
the physicochemical characteristics of HINS are well known
and widely reported in medical literature.
162 11 Formulation Development Concepts
• In vitro study on the diffusion of HINS through
polycarbonate membrane (PM) (Al-Achi and Green-
wood 1995): The PM has been used as a physical support
surface for cellular growth in cell culture studies. The
membranes were selected with pore sizes of 0.2, 1.0,
and 8.0 μm (membrane diameter of 13 mm). The diffu-
sion of HINS through the membrane depended on the
• Preparation of HINS with carrier systems: Several
pore size, pH, and the presence or absence of collagen
on the membrane’s surface. All experiments were carried
at 37 C. HINS diffusion was optimum at alkaline pH and
reached a minimum diffusion point near its isoelectric
point (pH ≈ 5.3). The presence of collagen on PM’s sur-
face slowed the rate of HINS diffusion and resulted in
a lesser total amount of HINS to diffuse over 24 hours.
Larger pores of PM (8.0 μm vs. 0.2 or 1.0 μm) resulted
in a more significant amount of the hormone diffused
(Al-Achi and Greenwood 1995). Moreover, HINS precipi-
tated out from its solutions when the pH was between 4
and 5.8. The diameter of precipitated HINS particles
increased as the pH increased from 4 to 5.8 and reached
its maximum diameter at pH 5.8 (Al-Achi et al. 1998).
• Cell culture studies on HINS diffusion through a
layer of Caco-2 cells (Greenwood and Al-Achi 1997):
Caco-2 cells are known to undergo a spontaneous differ-
entiation (15–21 post-incubation) in cell culture to enter-
• Preparation of the dosage forms (HINS suspen-
ocytic cells. This human colon adenocarcinoma cell line
is commonly used as an experimental model to study
drug diffusion through the intestinal epithelium. The
cells were grown on the surface of polycarbonate-colla-
gen-coated membranes (Corning® Transwell® polycar-
bonate membrane cell culture inserts) at a density of
60,000 cells/cm2. The permeability coefficient of HINS
through a differentiated layer of Caco-2 cells was (6.5 ±
1.2) × 10−7 cm/s. This permeability coefficient was similar
• Diffusion of HINS through excised skin: These experi-
in value to that reported elsewhere from an everted rat
small intestine model (Greenwood and Al-Achi 1997).
• In vitro studies on the stability of HINS: Soybean
powder (mean volume-surface diameter dvs = 45.5 μm;
average bulk volume 44.5 mL; average bulk density
0.625 g/mL; average angle of repose 38.33 ; and average
Carr index 26.72%) was prepared by grinding the beans
(Glycine max) in a coffee grinder (Mr. Coffee, Sunbean
Products, Hatiesburg, MS), and an aqueous extract
(pH = 6.5–7.0; density = 1.0178 g/mL; viscosity = 1.39
centipoises) was prepared from the resulting soybean
powder (Al-Achi et al. 2003). The aqueous extract was
prepared by mixing 5 g of the powder with 25 mL water
and the mixture was incubated for one hour at 37 C. The
mixture was then centrifuged and filtered to produce a
clear extract (Al-Achi et al. 2006). Soybean contains sub-
• In situ experiments (Al-Achi and Greenwood 1993b):
stances in its protein fraction with anti-proteolytic activ-
ities, inhibiting trypsin and chymotrypsin from reacting
with HINS. In the presence of the soybean extract, insulin
degradation by these enzymes was significantly reduced,
and the protection afforded by soybean extract was more
pronounced against trypsin and chymotrypsin than that
against pepsin (Al-Achi et al. 2006).
carrier systems were chosen to study their suitability to
interact with HINS. These carriers were erythrocyte
ghosts (EG) and sonicated erythrocyte ghosts to produce
erythrocyte vesicles (EV); some of these membranes were
further coated with liposome-type film (LEG and LEV).
All carriers were found to associate with HINS, with
EG exhibited the highest percent of this association
(approximately 50% of the initial amount of HINS was
found associated with EG following a 24-hour incubation
period at 37 C) (Al-Achi and Greenwood 1993a). In addi-
tion, the interaction of HINS with soybean powder
(Glycine max) was investigated. The maximum amount
of HINS that can be adsorbed on was experimentally esti-
mated to be 106 U/g of powder. The interaction between
HINS and soybean powder was relatively weak, which
allowed easy dissociation of the hormone upon contact
with aqueous media (Al-Achi et al. 2003).
sion): HINS solution was added to soybean extract (see
above) in a ratio of soybean extract:HINS solution
(100 U/mL) 1 : 9, and the pH of the mixture was adjusted
to 4.0. At this pH, HINS precipitated from solution forms
a suspension with a dispersed phase average diameter
(dvs) of 29.93 μm. The shelf life of the HINS suspension
was estimated to be approximately 18 days at 4 C
(Al-Achi and Parekh 2004).
ments were done using excised hairless mouse skin. The
skin was mounted on Franz cell apparatus, and the diffu-
sion of HINS through the skin with or without absorption
enhancers [limonene solution in alcohol (5%), iodine tinc-
ture, and ethyl acetate in alcohol (1 : 1)] was documented.
Limonene solution in alcohol facilitated the diffusion of
HINS, the most compared with the other two enhancers
(the permeability coefficient of HINS through the skin
was 9.47, 4.42, and 2.78 × 10−6 cm/s for limonene, iodine,
and ethyl acetate, respectively). The pretreatment with
limonene alcoholic solution produced a HINS flux
through the skin of 1.23 U/cm2/h, which imitated the pan-
creatic normal secretion rate. There was no HINS diffu-
sion through the skin when the hormone was applied
without an absorption enhancer (Al-Achi et al. 2011).
HINS was administered intraduodenally to anesthetized,
streptozocin-induced diabetic rats. EG, EV, LEG, and
LEV HINS-carrier systems and the free hormone were

administered locally, and the blood glucose of rats was
monitored over five hours. Free HINS did not produce
any hypoglycemic activities, while HINS-carrier systems
showed varying degrees of hypoglycemia; LEV was the
most effective carrier for delivering HINS to the circula-
tion via the intraduodenal route as indicated by a pro-
nounced hypoglycemic state (Al-Achi and Greenwood
1993b). The LEV-HINS carriers were further investigated
and found to be equally effective (i.e. causing hypoglyce-
mic effect) at doses of 200, 400, or 600 U/kg body weight.
Interestingly, the administration of free HINS via this
route in high enough doses (600 U/kg body weight) also
produced a hypoglycemic response (Al-Achi and Green-
wood 1994).
• Whole experimental animal studies (Al-Achi and
Greenwood 1993c): HINS free or associated with the car-
riers mentioned above were administered either orally
(by gavage) or buccally in streptozocin-induced diabetic
rats. The latter form of administration resulted in a low-
ering in blood glucose level following free HINS or EG-
HINS. These findings led to the conclusion that HINS
could pass through the buccal mucosa, reaching the cir-
culation to affect a state of hypoglycemia (Al-Achi and
Greenwood 1993c). Expect for LEG-HINS, which was
ineffective in lowering blood glucose level, the oral
administration of the other carriers-HINS mentioned
above or free HINS (333 U/kg of rat body weight) resulted
in a hypoglycemic response (Al-Achi and Greenwood
1998). This response has demonstrated once again that
a large enough oral HINS dose could bring down blood
glucose level, even when HINS was in its free form in
solution. The oral administration (by gavage) of soybean
extract-HINS suspension (1 mL soybean extract was com-
bined with 1 mL of Humulin®R; 167 U/kg of rat body
weight) in diabetic rats resulted in a significant reduction
in blood glucose level that was sustained for four hours
post-administration. The lowering in blood glucose was
observed as early as 30 minutes following the oral dose.
In contrast, control groups (saline or soybean extract) did
not show any hypoglycemic effects (Al-Achi et al. 2012).
The administration of a 200 U/kg of rat body weight
HINS oral dose to diabetic rats did produce any reduction
in blood glucose either (Al-Achi and Greenwood 1994).
It has been stated that only 1 out of 11 drugs entering
phase 1 clinical investigations reach the market, and only
one-third of the drugs that make it to the market cover their
development cost (Dubin 2006). It was also predicted that
the market for drug delivery systems would grow at an
annual rate of 10%, reaching $132 billion in 2012 (Dubin
2009). A compound annual growth rate (CAGR) of 4.9%
for advanced drug delivery technologies was expected
between 2015 and 2020 (Saharan 2019). And, according
11.1 Preformulation 163
to the thepharmaletter (https://www.thepharmaletter.
com/article/growth-forecast-for-drug-delivery-systems-
market (accessed 14 September 2020)), due to a forecasted
rise in future infections that require specialized instrumen-
tations (e.g. ventilators), CAGR for drug delivery systems
market is expected to be at 7% between 2020 and 2025.
Another driving force behind the rise in predicted growth
in drug delivery technologies is the extension of a product’s
life while minimizing cost (Hamilton and Lutz 2005).
Development of a new delivery system for a given drug
begins in research and development (R&D) departments
in the pharmaceutical industry, utilizing the principles of
preformulation. However, there is no guarantee whatso-
ever that a specific formulation that was generated in
R&D will be applicable in the production area. In addition,
the pharmaceutical industry utilizes what is known as proc-
ess analytical technologies (PATs). The main purpose of
PAT is for control and assessment during manufacturing.
For instance, the compression force signal during tableting
can be used to accept or reject a tablet during processing
(Manley et al. 2019). Moreover, near-infrared spectroscopy
is being employed as PAT for many steps in tablet manufac-
turer such as blending uniformity, as well as for moisture
content detection in the finished tablets (Rajan et al.
2010). It is important for the R&D department and the pro-
duction operation facility to coordinate their use of PAT
and to allow communication concerning PAT to flow
smoothly back and forth (Arrivo 2003). The obvious advan-
tage of PAT is that it provides online, in-place analysis for
an active pharmaceutical ingredient (API) during the pro-
cessing steps, eliminating the need for repeated sampling
during the manufacturing operation. Thus, it reduces the
exposure of personnel to APIs, a matter of importance
when handling highly potent drugs. PAT also reduces both
the overall cost and the time of operation (Arrivo 2003). In
September 2004, the US FDA issued a Guidance for Indus-
try document concerning PAT. The use of PAT allows the
industry to produce a pharmaceutical product of high qual-
ity in a consistent and predictable manner (Dziki 2008).
The purpose served by R&D is often questioned, since a
considerable amount of information learned there may not
be applicable to production. In this chapter, we shed some
light on issues related to transferring a formulation from
R&D to production (i.e. the scale-up process), rate-
controlled drug delivery systems, novel drug delivery
technologies to enhance oral bioavailability, possible
drug–excipient or excipient–excipient interactions, con-
tamination issues, and other issues. To make the point in
another way, according to Arthur Calder-Marshall, “out
of sight, out of mind” translated into Russian by a com-
puter, then back into English, became “invisible maniac”
(Tingstad 1988). Thus, clear and open communications
between a R&D department and production personnel
164 11 Formulation Development Concepts
cannot be overemphasized for the ultimate success of the
final product.
Eight principles determine whether or not a process is suit-
able for manufacturing a drug product. The process must be
robust, simple, safe, standardized, scalable, minimize the
cost and time of production, and transferable/scalable
(Brower et al. 2020).
11.2 Scale-up Considerations
Issues related to scaling-up dosage forms from the R&D
stage to the pilot stage and thereafter to production are
numerous and often lead to failure in producing the
planned dosage form unless addressed and resolved. Most
problems encountered during scale-up are related to solid
dosage forms such as tablets and suppositories (Russo
1984; Chowhan 1988), as well as lyophilization techniques
(Tchessalov et al. 2007). Variability in the chemicals with
respect to their physical characteristics and equipment var-
iability are perhaps the main areas of concern during scale-
up (Russo 1984; Pondell 1985). For example, during tablet
formulation, variations exist during milling and mixing,
and in the types of presses used (Russo 1984). Moreover,
a fluid-bed granulation process is often used in industry
in tablet manufacturing. The basic fluid-bed granulation
unit consists of an air-handling unit, an exhaust air turbine,
an exhaust air filter, a processing zone (i.e. dryers, granula-
tors, spray nozzles), and product discharge components
(Olsen 1989). This technique allows the production of gran-
ules with ideal features of spherical shape, high porosity,
and narrow size range. These features provide excellent
flow characteristics and compressibility for the resulting
granulation. Production batches are naturally higher in
weight and require more time for processing. Fluid beds
employed in the production stage produce higher shearing
forces to form the desired agglomeration. They utilize
warmer fluidizing air than that used with R&D units,
and the rate for delivering the binder dispersion is many-
fold higher than that of a R&D application. These two fac-
tors present in the production unit produce granulation
containing a lower moisture content just prior to the drying
stage. Production tablets often have higher disintegration
times, albeit not significant enough to affect the dissolution
profile of tablets. To achieve a smooth transition from R&D
to production utilizing fluid-bed technology, factors such as
binder delivery rate and moisture content must be consid-
ered (Gore et al. 1985). Overall, and for the fluid-bed gran-
ulation scaling-up procedure, close attention should be
directed to factors such as batch size, type of equipment
used, delivery rate of binders, pressure of the atomization
air, volume of the fluidization air, and temperature of the
inlet air (Gore et al. 1985; Mehta 1988). Granulations made
by high-shear mixtures present another type of challenge
for scale-up. There appear to be four areas of concern
regarding this scaling-up operation: the first three being
variations in the API and the excipients, in-process con-
trols, and failure to recognize the fact that scaling-up for-
mulations from R&D to production is not a linear
process. However, the most significant factor affecting
the process of transition to large-scale production was
found to be the variation encountered with the API
(Chowhan 1988).
Film coating is another area of concern during scale-up.
For film coating, two major types of equipment are availa-
ble: fluid-bed type and pan coaters. Each of these coaters
exists in many types, and those used in the R&D stage
are obviously smaller than those employed in production.
Smaller instruments handle materials in a much gentler
way. Large production large equipment could damage a
product at any step before and after coating (Pondell
1985). Therefore, care must be exercised to protect a prod-
uct during handling in the production areas, despite evi-
dence of “stability” throughout the R&D process. Other
factors that may contribute to the quality of the finished
coating film include the solid content of the coating disper-
sions, the number of spray guns used, the pan speed (in the
case of pan coating devices), and the coloring agent if used
in the coating dispersion. All these factors play a significant
role during processing of the final product formulation
(Porter and Saraceni 1988).
Several drying technologies are utilized in industry for
the purpose of improving product’s stability. These technol-
ogies include lyophilization (freeze-drying via sublimation
of water), spray drying, and reduced-pressure drying
(Kawasaki et al. 2019). While freeze-drying techniques
are useful for a variety of products, including biologicals,
spray-drying techniques are limited to low-molecular-
weight compounds due to the heat and sheer-stresses used
during spay drying cycles (Lowe et al. 2018). Lyophilization
is a commonly employed process in industry for the prep-
aration of sterile powdered dosage units. This technique
requires several steps that may create challenges during
the scale-up process. Lyophilization process involves three
main stages; these are freezing stage and the formation of
ice crystals, primary drying stage (sublimation of ice crys-
tals; it is the longest of the three stages), and secondary dry-
ing stage (desorption of unfrozen water) (Kawasaki et al.
2019). Several instrumentations may be used to monitor
the temperature and pressure during the freeze-drying
cycles. These monitoring devices include capacitance man-
ometers, Pirani gauges, resistance temperature detectors,
and thermocouples (Sacha 2020). In addition to these chal-
lenges faced during cycle development, formulators have
also to consider other areas of the freeze-drying process
 11.2 Scale-up Considerations 165

such as cake characterization and process optimization
(Zhang et al. 2020). From the point of view of the scal-
ing-up technique, temperatures during the freezing, pri-
mary, and secondary drying steps must be similar for the
laboratory and manufacturing processes. This guarantees
not only the same product characteristics but also the same
stability profile (Tchessalov et al. 2007). Thus, two para-
meters for a successful scale-up of the lyophilized product
have emerged for their importance in this respect; these are
the water vapor transfer resistance of the dried layer that is
formed during the freeze-drying cycle and the vial heat
transfer coefficient. The latter is the algebraic sum of three
other coefficients, namely the contact heat transfer, the gas
heat transfer, and the radiant heat transfer coefficients
(Kawasaki et al. 2019). The vial heat transfer coefficient
value is directional proportional to the mass loss over time
and the latent heat of ice and inversely proportional to the
cross-sectional area of vial (calculated from its outer diam-
eter) and the difference between the shelf surface temper-
ature and the product temperature. The water vapor
transfer resistance is inversely proportional to the mass loss
over time and directly proportional to the cross-sectional
dried layers and the difference between the equilibrium
vapor pressure of ice and the vacuum pressure (Kawasaki
et al. 2019).
At the beginning of the twenty-first century, the transfer
of a lyophilized product from R&D to production necessi-
tated that the production team relied on trial-and-error sort
of approach to produce the lyophilized product in industrial
quantities. This was dramatically changed, toward the end
of the first decade of this century, with the establishment of
the principles of quality-by-design (QbD) with its critical
elements, namely the Design Space and Process Analytical
Technology (PAT). These principles indicated that “a
systematic approach to development that begins with pre-
defined objectives and emphasizes product and process
understanding and process control, based on sound science
and quality risk management” be implemented (Kawasaki
et al. 2019). The guiding principles of QbD along with their
attributes related to lyophilization process are summarized
in Table 11.1 (Kawasaki et al. 2019).
However, what QbD principles apply to small-molecule
production may not necessarily be fully applicable to bio-
pharmaceuticals. These latter products have significantly
more quality attributes than those encountered with small
molecules (Menezes et al. 2019). An example of these appli-
cations to biopharmaceuticals is applying QbD principles to
develop a high-dose, self-injectable delivery system of bio-
logics where the need to inject a viscous liquid with a high
volume (2 mL or more) via an auto-injector exists. For such
a case, a patient-centric approach is needed that provides
robustness and acceptability by the user (Le Gal 2019).
Lyophilization can also be applied to liposomal type for-
mulations. This is done to improve the shelf life of the lipo-
somal preparation. The challenges that face the formulator
in this process are related to the freezing and the dehydra-
tion of the liposomes that lead to destabilization of the
vesicular structure with the concomitant drug leakage
(Franzé et al. 2018). Experimental data have shown that
substances such as fructans (inulins; linear fructose poly-
mers; isolated from plants such as chicory roots and dahlia
tubers), when combined with glucose are capable of stabi-
lizing the phospholipid structure of liposomes during the
lyophilization (Hincha et al. 2000). Others (Kannan et al.
2015; Guimarães et al. 2019) have found that the inclusion
of sucrose as a lyoprotectant during the lyophilization proc-
ess of liposomes has greatly improved the physical stability
of the formulation. It is important to note here that glucose

Table 11.1 QbD: guiding principles.

Lyophilization parameters
Guiding principles Description or attributes

Critical Quality Physical, chemical, biological, or microbiological properties or

•• Glass transition temperature

••
Attributes (CQAs) characteristic that should be within an appropriate limit, range, or Eutectic temperature
distribution to ensure the desired product quality Cake collapse temperature

•
Product temperature
Water vapor transfer resistance of
the dried layer

Critical Process Process parameters whose variability has an impact on a CQA and •• Freezing temperature

••
Freezing rate
Parameters (CPPs) therefore should be monitored or controlled to insure the process produces
Annealing temperature/time
the desired quality
Primary and secondary drying
temperature, pressure, and time

Critical Material Attributes of input materials whose variability has an impact on a CQA and •• Related substances

••
Appearance
Attributes (CMAs) therefore should be monitored or controlled to insure the process produces
Water content
the desired quality
Reconstitution time
Source: Adapter from (Kawasaki et al. 2019).
166 11 Formulation Development Concepts
and sucrose exhibit a collapse temperature (Tc) and glass
transition temperature of maximum freeze concentrate
(T g) of (−40 C; −43 C) and (−32 C; −32 C), respectively
(Franzé et al. 2018). If the prevailing temperature is below
T g, the material resembles solid glass; and when the ambi-
ent temperature is below Tc, the material can support and
maintain its “rigid” structure. Other commonly used lyo-
protectants during the lyophilization process include
(Franzé et al. 2018) arginine, dextrans (various molecular
weights), gelatin, histidine, HP-β-cyclodextrin, kaemp-
ferol-3-O-glucoside, lactose, lysine, maltoexaose, malto-
heptaose, maltose, maltotetraose, maltotriose, mannitol,
quercetin-3-O-glucoside, quercetin-3-O-rhamnoside, and
trehalose. It should be noted that the lyoprotectants vary
in their effectiveness with the type of liposomes being used
(Franzé et al. 2018; Guimarães et al. 2019).
As stated above, the main advantage for lyophilization is
to render the product more stable than if it were in a liquid
dosage form. Thus, lyophilization techniques require low
temperatures during processing, which is highly suited to
heat-sensitive APIs. In addition, from the cost point of view,
it is cheaper to transport lyophilized units than liquid-filled
containers due to weight differential between the two dos-
age products (Thomas 2020a). It should be noted, however,
that not all formulations can be subject to lyophilization.
For instance, those containing high salt concentration have
a reduced freezing point (based on the colligative properties
of solutions) and are difficult to freeze-dry. Moreover, for-
mulations with volatile components that have high vapor
pressure (e.g. acetate buffers) and can be lost during the
vacuum cycle. Techniques, such as lyophilization, also
require highly trained pharmaceutical scientists specializ-
ing in this area of formulation applications (Thomas
2020a). Such expertise is available at companies that spe-
cialize in this type of technology. Exelead (Indianapolis,
IN) is such a company that offers supports in developing
liposomal and PEGylation formulations for parenteral ster-
ile drug products (in areas such as enzyme replacement
treatment, infectious diseases, and oncology). Incidentally,
PEGylation is the process of attaching a polyethylene oxide
polymer to a pharmacologically active drug molecule. The
purposes of which are to extend the biological half-life of
the drug and, more importantly, to diminish the host’s
immune responses. Overall, PEGylated drug molecules
have better therapeutic actions (Okutgen et al. 2020).
11.2.1 Vaccines
The primary purpose of administering vaccines is to stim-
ulate the immune system to identify and protect the indi-
vidual from impending infections. Some vaccines are
made to fight cancer development or to help in its treat-
ment. Examples of the formers are the vaccine used against
human papillomavirus and another against hepatitis
B virus. Examples of the vaccines that are approved for
treating cancer are sipuleucel-T (Provenge®; Dendreon
Pharmaceuticals LLC, for the advance stage of prostate
cancer – metastatic castration-resistant prostate cancer)
and talimogene laherparepvec (Imlygic®; Amgen; for mel-
anoma of the skin and lymph nodes) (Baluja 2018). It has
been reported that by mid-2017, there was an estimated
total of 240 new vaccines and immunotherapies against
cancer being investigated in clinical trials (O’Connor
2019). Vaccines that are used to fight infectious diseases
are defined as antigenic substances that are made from
the causative agent of a disease or a synthetic substitute
or carry a genetic material (e.g. mRNA) of the infectious
agent to promote the making of antigens by the host cells
that trigger the immune system to build antibodies against
them (e.g. vaccines made by Pfizer-BioNTech and Moderna
against COVID-19 viral infection, caused by SARS-CoV-2).
Vaccines are used to provide immunity against one or mul-
tiple diseases (Calle 2020). This immunity is often granted
early in life during childhood. Pediatric vaccines are many
and include conjugate, live-attenuated, toxoid, and inacti-
vated vaccines (Wei Ling 2020). Some of the vaccine formu-
lations may also contain substances that are known
collectively as “adjuvants.” According to the US FDA, an
adjuvant is “a substance added to some vaccines to enhance
the immune response of vaccinated individuals.” The FDA
lists several of those adjuvants that are found in some of the
vaccine preparations. The most important of which are alu-
minum salts, monophosphoryl lipid A, an oil-in-water
emulsion composed of squalene, a combination of mono-
phosphoryl lipid A and QS-21 (a natural compound found
in the central Chilean evergreen soapbark tree Quillaja
saponaria Molina), AS04, which is a combination of alumi-
num hydroxide and monophosphoryl lipid A, and CpG
1018 (cytosine phosphoguanine – a synthetic form of
DNA, which resembles the genetic composition of bacteria
and viruses). Vaccines that do not need adjuvants for their
activation include the messenger ribonucleic acid (mRNA)
types. The mRNA vaccines rely on introducing a synthetic
viral mRNA to the recipient’s cells, allowing the cell to man-
ufacture viral proteins. These, in turn, are recognized by the
organism as antigens, and the immune system raises antibo-
dies against them. Multiple dosing is commonly needed with
the mRNA vaccinations to achieve their desired protective
effect against viral infections. Recent examples of mRNA
vaccines are the ones developed by Pfizer-BioNTech (New
York, NY) and Moderna (Cambridge, MA) to combat novel
coronavirus infection (COVID-19). The active ingredient in
the Pfizer COVID-19 and Moderna vaccines is nucleoside-
modified mRNA encoding the viral spike (S) glycoprotein
of SARS-CoV-2. They differ in their inactive ingredients;
the Pfizer-BioNTech vaccine contains 2[(polyethylene glycol
(PEG))-2000]-N,N-ditetradecylacetamide (protects/shields
mRNA from cell’s microenvironment and lipid lubricant

to facilitate cell entry), 1,2-distearoyl-sn-glycero-3-phospho-
choline, (4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis
(2-hexyldecanoate) (protects/shields mRNA from cell’s
microenvironment and lipid lubricant to facilitate cell entry),
sodium chloride (stabilizer), potassium chloride (stabilizer),
and dibasic sodium phosphate dehydrate (pH adjustment).
Those in the Moderna vaccine are PEG2000-DMG: 1,2-
dimyristoyl-rac-glycerol (protects/shields mRNA from cell’s
microenvironment and lipid lubricant to facilitate cell entry),
methoxypolyethylene glycol (protects/shields mRNA from
cell’s microenvironment and lipid lubricant to facilitate cell
entry), 1,2-distearoyl-sn-glycero-3-phosphocholine (pro-
tects/shields mRNA from cell’s microenvironment and lipid
lubricant to facilitate cell entry), SM-102: heptadecan-9-yl
8-((2-hydroxyethyl) (6-oxo-6-(undecyloxy) hexyl) amino)
octanoate (protects/shields mRNA from cell’s microenviron-
ment and lipid lubricant to facilitate cell entry), trometha-
mine (pH adjustment), tromethamine hydrochloride (pH
adjustment), acetic acid (pH adjustment), and sodium
acetate (pH adjustment). They both contain cholesterol
(protects/shields mRNA from cell’s microenvironment and
lipid lubricant to facilitate cell entry) and sucrose (cryopro-
tector). The Johnson & Johnson/Janssen vaccine against
COVID-19 is based on viral vector technology and has as
its active ingredient the recombinant, replication-
incompetent Ad26 vector, encoding a stabilized variant of
the SARS-CoV-2 Spike (S) protein. The vaccine formulation
also contains inactive ingredients: Polysorbate-80 (Tween-
80; surfactant/stabilizer), 2-hydroxypropyl-β-cyclodextrin
(solubilizer/stabilizer), citric acid monohydrate (pH adjust-
ment/stabilizer), trisodium citrate dehydrate (pH adjust-
ment/stabilizer), sodium chloride (stabilizer), and ethanol
(solvent). All three vaccines are free from eggs, preservatives,
latex, and metals (iron, nickel, cobalt, lithium, rare earth
alloys, or any manufactured products) (https://www.cdc.
gov/vaccines/covid-19/clinical-considerations/covid-19-
vaccines-us.html#Appendix-C (accessed 16 August 2021)).
Another relatively new development in this field is that
the FDA granted Seqirus Inc. (Maidenhead, UK) authoriza-
tion in February 2020 to market Audenz (influenza A/
H5N1 monovalent vaccine for persons six months of age
and older). This approval would be the first time that the
FDA has permitted a cell-based, adjuvanted (adjuvant
was MF59®) vaccine in the United States. The company
is producing the vaccine in its facility in the US (Holly
Springs, NC).
Some of the available vaccines are given via parenteral
administration, and some are administered orally. The oral
route of administration offers the advantage of being safer
and easier to deliver than the injectable forms, leading to
improving compliance. From the manufacturing point of
view, it is relatively easier, in general, to produce an oral
dosage form than a parenteral one. When vaccines are
11.2 Scale-up Considerations 167
given orally, they also tend to interact with the lymphatic
system elements populating the digestive tract pathway,
which leads to activation of the immune system quicker
than that observed from a parenteral dose administration
(Vela Ramirez et al. 2017). Vaccines may be dispensed as
a single-unit container or a multi-dose container. For the
multi-dose containers, a preservative is needed to prevent
bacterial growth within the product over its shelf life.
A list of licensed vaccines in the United States is found
on the FDA website (https://www.fda.gov/vaccines-
blood-biologics/vaccines/vaccines-licensed-use-united-
states (accessed 9 September 2020)). Vaccines for viral
infections can be prepared with live attenuated virus, inac-
tivated virus, or by recombinant methods. Those made to
protect against a bacterial infection are prepared from
whole-cell inactivated bacteria, bacterial sub-units, toxoid,
or conjugate forms (Plotkin et al. 2017).
Scale-up of vaccines (Markarian 2020) to produce mil-
lions or even billions of units can be a magnificent chal-
lenge. The cost for establishing a new vaccine facility is
estimated to be in the hundreds of millions of US dollars
for mammalian-cell-based facilities (Robinson 2009).
Depending on the medical urgency, such as during pan-
demics, the units to be produced occasionally have to be
manufactured at different locations and within a short
period of time. Thus, in addition to controlling for the
raw materials quality (e.g. containers) and manufacturing
procedures (e.g. maintaining sterility during processing),
issues related to training personnel at the different facilities
to get them to the same level of competency is an added
consideration for the production team to plan for
(Markarian 2020). Also, for manufacturers with other pro-
ducts being manufactured within the same facility, this
raises the question of cross-contamination. In situations
like these, the manufacturer has to have “cleaning-in-
place” procedures for all surfaces, including those of the
equipment that is used for the end units, as well as for
the intermediate products (Challener 2020). Of course,
the issue of cross-contamination can be greatly minimized
or totally eliminated by the use of single-use platforms such
as the automated one available from Sartorius (High Perfor-
mance Single Use Platform; BIOSTAT STR Generation 3)
for bioreactors (Sartorius, Goettingen, Germany). This sys-
tem provides real-time data collection and it is data-driven;
the system is completely controlled by BIOBRAIN, an auto-
mation platform. Pall Corporation (a global company) has
introduced the iCELLis bioreactor system that offers an
automation platform with a single-use capability, which
greatly facilitates the scale-up of vaccine manufacturing
(Castillo 2013). And the demand for these types of reactors
is on the increase (13.5% per year) (Mirasol 2020). Some of
these new bioreactors have double the upper range capacity
of 2000 L, reaching upward to the 6000 L volume capacity
168 11 Formulation Development Concepts
(experts argue over their usefulness beyond the current
2000 L bound) (Mirasol 2020). It should be noted that, in
general, single-use platforms are more suited for persona-
lized cell/gene therapy, while continuous processing appli-
cations are more utilized in large production volume
methodologies, such as vaccine or virus manufacture
(Anonymous 2020a). Although the development of a new
vaccine is highly time consuming, Novavax, Inc.
(Rockville, MD) has developed a rapid system for preparing
influenza vaccine that is an insect-cell based (a new vaccine
for influenza can be made within 10–12 weeks). This insect-
based system allows the use of disposable reactors due to its
high yield production (Robinson 2009). In 2020, Novavax,
Inc. platform was utilized to produce a vaccine against
the novel coronavirus (COVID-19) pandemic. At the writ-
ing of this section, a dozen of other companies are engaged
in developing vaccines against this virus that was discov-
ered in China during the fall season of year 2019. The virus
spread worldwide and has resulted in hundreds of thou-
sands of people dying from the viral infection globally.
COVID-19 is particularly dangerous in patients with
comorbid conditions (e.g. diabetes, obesity, high blood
pressure) and seniors. For the latter group, their immune
system is weakened with aging, and a supportive-type vac-
cine in addition to the leading vaccine may be of impor-
tance to provide an added prolonged immune support
through T-cell-mediated immunity (Wolf 2020). One of
the major challenges that face vaccine development against
viruses is the ability of the virus to mutate. For instance,
Influenza A has 18 different subtypes and thousands of
strains for each subtype. By mutating, the virus can go
undetected by the immune system (Panjwani 2019).
Vaccine production is licensed for the actual biological
entity and for the manufacturing process for producing
the final product. Changes in the manufacturing process
can trigger licensing-related issues with regulatory agencies
(Plotkin et al. 2017). In manufacturing vaccines, several
considerations have to be identified for producing the final
product. These considerations include (1) biological aspects
such as the nature of the microorganism, (2) growth
medium aspects (the specifications of the microbial cul-
ture), (3) the phases needed for the purification processes,
(4) personnel-related knowledge and experience of the
manufacturing process, and (5) the methods used to ana-
lyze the antigens and other biological processes resulting
from vaccine production (Plotkin et al. 2017). Stabilization
of the vaccine preparation can be done by preparing the
solution under optimal pH for stability using a buffer sys-
tem or preparing a lyophilized dosage form that is less
prone to degradation (Plotkin et al. 2017). Examples of vac-
cines prepared by lyophilization are VAXCHORA™
(Cholera Vaccine, Live, Oral), Suspension for Oral Admin-
istration (PaxVax, Inc., Redwood City, CA), and BCG
VACCINE (TICE® strain) (for the prevention of tuberculo-
sis; Organon Teknika Corporation LLC, Durham, NC). A
list of licensed vaccines in the United States is found on
the FDA website (https://www.fda.gov/vaccines-blood-
biologics/vaccines/vaccines-licensed-use-united-states
(accessed 14 September 2020)).
Manufacturing platforms for vaccine production vary greatly
among pharmaceutical companies. This variability in the
production is due to the different forms of vaccines that can
be produced (active vs. inactive form; parenteral vs. oral admin-
istration). Table 11.2 summarizes some of the substances found
in vaccine products licensed in the United States (https://www.
fda.gov/vaccines-blood-biologics/safety-availability-biologics/
common-ingredients-us-licensed-vaccines (accessed 10
September 2020)).
11.3 Combination Products
Compliance with taking medications is a huge issue in clin-
ical practice. It is estimated that the cost of noncompliance
can be as high as $100 billion annually, accounting for addi-
tional health care expenditures and lack of productivity
(Dubin 2005a). Combination products offer one solution
to improving patient compliance. The notion of combina-
tion products is not a new concept in the pharmaceutical
industry. A quick perusal of clinical reference texts reveals
their presence since the industry began. However, the old
types of combination products were limited to certain drugs
that did not show compatibility issues, and their quantities
were small enough to accommodate them within a single
dosage unit. The new combination products are much more
versatile in their ability to include products in larger quan-
tities and regardless of compatibility issues. Multicompart-
ment capsules (e.g. InnerCap Technologies, Inc; Tampa,
FL) are made to house several drugs to treat the same or
different conditions, regardless of their compatibilities.
The new capsules are made of a suitable material, such
as hydroxypropyl methylcellulose, gelatin, starch, or a com-
bination. Coating materials can be applied to the outer
walls for specially targeted delivery. [Unlike hard-shell cap-
sules, soft gelatin capsule shells are to date only available as
gelatin-based units (Thomas 2020b).] Another example of
multicompartment capsules is BilDil (NitroMed Inc., Lex-
ington, MA), containing the combination vasodilator drugs
isosorbide dinitrate and hydralazine hydrochloride
approved by the FDA for the treatment of heart failure in
African-American patients (Dubin 2005b).
In general, combination products can deliver simultane-
ously, within the same unit, a drug, a biological, and a med-
ical device. Most of these products (90%) are currently being
developed for cardiovascular applications and focus on
 11.3 Combination Products 169

Table 11.2 Substances found in vaccines.

Substance name Function

AS04 (a combination of aluminum hydroxide and monophosphoryl lipid A) Adjuvant

AS03 (D,L-alpha-tocopherol (vitamin E), squalene, and polysorbate 80) Adjuvant
MF59 (an oil-in-water emulsion of squalene oil) Adjuvant
CpG 1018 (synthetic DNA sequencing) Adjuvant
AS01B (a combination of monophosphoryl lipid A and QS-21, a substance Adjuvant
extracted from the Chilean tree Quillaja Saponaria)
Aluminum salts (e.g. aluminum hydroxide) Adjuvants
Neomycin Antibiotic
Polymyxin B Antibiotic
Streptomycin Antibiotic
Gentamicin Antibiotic
Fetal calf/bovine serum Cell nutrients during manufacture
Formaldehyde (≤0.005 mg–0.1 mg/dose) Denaturing agent
Thimerosal (0.001–0.01%) Preservative
Phenol (0.25%) Preservative
2-Phenoxyethanol (100 μg/mL) Preservative
Benzethonium chloride (25 μg/mL) Preservative
Lactose Stabilizer
Sucrose Stabilizer
Glycine Stabilizer
Monosodium salt of glutamic acid Stabilizer
Human serum albumin Stabilizer
Gelatin Stabilizer

developing stent drug delivery (Prajapati et al. 2008). Other
applications involve areas related to surgical dressing and
orthopedics. Also, an emerging area of combination prod-
uct technology includes nanotechnology (Ford 2008). This
area has evolved into a new specialty known as nanomedi-
cine. Applications in this area include improved
antibacterial agent activity, acting as carriers, improvement
in implantable materials, diagnostic applications, and sen-
sory units in various devices (Prajapati et al. 2008). For
example, new technologies are being developed to over-
come issues related to post-stent placement (e.g. re-
blocking of arteries, thrombosis, blood clots). Among these
technologies, Micelle technologies’ polymer nanoparticle
coating technique allows multiple drug delivery from a
single stent, as well as controlled delivery of drugs in a
sequential manner (Andrews and McClain 2008).
A French-government-funded technological research
organization, CEA-Leti, is currently investigating the use
of a nanoparticulate system (LipidotsTM; bio-assimilable
lipids) for developing a vaccine for human immunodefi-
ciency (HIV) virus infection (Navarro 2019). Ascendia
(North Brunswick, NJ) has developed nanotechnology plat-
forms suitable for oral and parenteral administration.
Ascendia’s EmulSol allows the formation of oil-in-water
nanoemulsions (droplet sizes: 50–500 nm), with drug solu-
bilized within the droplets of the oily phase. The nanoemul-
sion is formed through homogenization. This method was
found helpful in enhancing the bioavailability of drugs and
allowing medicines that are not candidates for parenteral
administration to be given via this route (e.g. clopidogrel;
anti-thrombotic) (Huang 2020a). Unlike microemulsions
that are considered to be thermodynamically stable systems
due to the presence of stabilizers (surfactants and co-sol-
vents), nanoemulsions are subject to Ostwald ripening
effect, which tends to destabilize their formulation (see
Chapter 15) (Huang 2020a). The other platforms of
Ascendia are AmorSol, NanoSol, and controlled-release
applications. According to Ascendia, AmorSol is for the
production of amorphous solid dispersions; NanoSol is
for producing nano-sized drug particles. Applications for
controlled-release techniques are for poorly soluble drugs
and include the coated particles (membrane-diffusion
170 11 Formulation Development Concepts
mechanism) and molecularly dispersed systems (matrix-
erosion mechanism) (see Section 11.4). Ascendia nanotech-
nology platforms may be found favorable in producing drug
products under section 505(b)(2) pathway (for information
on section 505, refer to Chapter 22), which allows compa-
nies to obtain FDA approval relying in part on clinical data
or other companies’ literature (Huang 2019, 2020a).
11.4 Rate-Controlled Drug Delivery
Systems for controlling drug release from a dosage form
have been developed to maximize therapeutic benefits
while minimizing adverse outcomes of drug therapy. Diffu-
sion systems and activation approaches are the two main
classes of rate-controlled drug delivery. Diffusion systems
are of several types and include membrane permeation,
matrix diffusion, and microreservoir dissolution. The acti-
vation technologies utilize hydrodynamic, osmotic, or
vapor pressure methods. Ion or pH activation, ultrasound,
and magnetism are all activation delivery techniques
(Chien 1985).
Membrane permeation allows a polymeric barrier to con-
trol delivery of a drug housed inside a rigid compartment.
Usually, the drug is in the form of dispersion, and its deliv-
ery through the membrane follows a zero-order rate. Matrix
diffusion systems depend on dispersing the drug in a biode-
gradable matrix form. The rate of drug diffusion from the
polymeric matrix is a function of the square root of time.
The microreservoir dissolution method employs a suspen-
sion of the drug particles in an aqueous solution dispersed
throughout a hydrophilic polymeric matrix. The suspen-
sion may be housed within an additional polymeric mem-
brane to further control the rate of drug release. The rate of
drug release from the device is a function of its solubility in
the aqueous solution and in the polymeric matrix
(Chien 1985).
Some gastroretentive systems, for example, work via
membrane permeation mechanisms. Floating drug delivery
systems that contain a drug dispersed in a hydrophilic
matrix (e.g. guar gum; hydroxypropyl methylcellulose)
exhibit a density below than 1 when placed in an aqueous
environment (such as gastric fluid) and thus float on the
surface of the aqueous liquid (Jain et al. 2005). The hydro-
philic matrix can form a superporous hydrogel. Upon
absorption of water, the hydrogel material swells with
the formation of space within the matrix, known as the
effective pore size. When the effective pore size is larger than
10 μm, the hydrogel is known as a superporous hydrogel
(Amin et al. 2008a). Some examples of this technology
available on the market are Liquid Gaviscon (an antacid;
GlaxoSmithKline), which is a floating alginate raft;
Baclofen GRS (a bronchodilator; Sun Pharma), which con-
sists of a coated multilayer floating and swelling device; and
Glumetza (for blood glucose control; Depomed), which is a
polymer-based system (Amin et al. 2008a). These units
release the drug one layer at a time, starting with the most
superficial layer first. The outer layer in contact with aque-
ous fluid swells and prevents further entry of the fluid
inside the matrix. When the outer layer swells, it entraps
some air and helps to keep the unit floating on the surface
of the fluid (Dubin 2007).
Other systems contain matrices prepared from swellable
polymers and effervescent materials that upon contact with
the gastric fluid generate CO2 gas; the gas gets entrapped in
the polymer structure, helping the unit to float (Dubin
2007). Once the outer layer is completely degraded and
the drug is released from it, a new outer layer is formed,
and the process continues until the entire unit is gone.
The hydroxypropyl methylcellulose (HPMC), with appro-
priate hydration rate and viscosity, used in these floatable
units is extremely popular in the preparation of con-
trolled-release systems. Its popularity stems from several
features, including a high capacity for drug loading, easy
compression into tablets, having temperature- and
humidity-independent viscoelastic characteristics, being a
crystallization inhibitor which helps enhance poorly solu-
ble low-ionized APIs’ solubility and bioavailability (in par-
ticular those containing a neutral amine group), and its
inertness, among others (Gothoskar 2005; Stegemann
2020). Floating foam-based systems are also used to provide
gastroretentive drug delivery. The matrix of these systems
consists of porous (low density) material resembling foam
(e.g. Accurel MP; polypropylene foam powder). Drugs with
or without added polymers are adsorbed on the foam par-
ticles. The entrapped air within the matrix allows the unit
to float upon exposure to an aqueous environment. The
drug release profile from the system is independent of
the foam materials; however, it depends on the type of pol-
ymer used with the drug (Amin et al. 2008a).
Other floating matrices utilize lipid materials as a base.
For example, lipid matrices known as Gélucires
(Gattefosse, St-Priest, France), composed of mixtures of
mono-, di-, and triglycerides with poly(ethylene glycol)
esters of fatty acids, have been shown to provide a sustain-
ing effect. Gélucires have hydrophile–lipophile balance
(HLB) values in the range 1–18, with a wide melting-point
range between 33 and 65 C. Low-HLB-value Gélucires pro-
duce units with floating characteristics gastrointestinal
fluids. The absence or low content of PEG esters mixed with
Gélucires provides a sustaining effect. [Larger amounts of
PEG esters mixed with the lipid matrix accelerate the rate
of drug release (Amin 2008a, 2008b; Patel and Patel 2008).]
Glyceryl monostearate retards the release of drugs from for-
mulations. When used in combination with Gélucires, the

matrix failed to float. Mixtures of ethylcellulose, a release
retardant agent, with Gélucires provided both retardation
of drug release and floating characteristics. Gélucires com-
bined with HPMC showed an accelerated drug release pro-
file compared to Gélucires alone (Amin et al. 2008a).
Another oral delivery membrane permeation technology
is the Eudramode platform (Evonik Röhm GmbH, 2012,
Darmstadt, Germany), which is made of layered pellets.
The core of the pellets is composed of modulator ions (a salt
of weak or strong acids, such as sodium citrate) layered
with a modulating film (Eudragit NE 30 D), followed by
a layer of the drug, and a covering controlled-release layer
of quaternary ammonium ions (Eudragit RL/RS 30 D;
ammoniomethacrylate copolymers). The pellets may be
placed in a capsule form or compressed into a tablet. The
Eudramode platform was shown to deliver the drug in a
zero-order release fashion. Three factors play an important
role in controlling the drug release from the pellets; these
are the salts used in the core and the thickness of both
the modulating film and the controlled-release covering
outer layer (Ravishankar et al. 2005). Eudragit polymers
(Evonik Röhm GmbH, 2012, Darmstadt, Germany) are
copolymers derived from esters of acrylic and methacrylic
acid. Their properties are based on functional groups
located on the polymeric chain. Eudragit polymers are
available in a wide range of applications, including aqueous
dispersions (L50 D-55; NE 30 D; NE 40 D; NM 30 D; RS
30 D; RL 30 D; FS 30 D), powders and granules (L100;
L100-55; S 100; RS 100; RL 100; RS PO; RL PO; E-100;
E PO), and organic solutions (E12,5; L12,5; S12,5; RS12,5;
RL12,5) (Evonik Röhm GmbH). Eudragit polymers can
be classified into two main categories: pH-dependent and
pH-independent. As pH-dependent films, the polymers
become soluble in solution if the pH exceeds a certain
value, whereas pH-independent polymer films act as a bar-
rier, allowing diffusion of a drug through the film. When
used as a matrix, the polymer acts as a sponge, releasing
the drug in a controlled fashion. The drug diffuses out of
the pH-independent matrix, whereas it is released by ero-
sion and diffusion when a pH-dependent matrix is used
(Álvarez et al. 2007).
Activation technologies that control drug delivery by a
pressure mechanism depend on interaction between the
device and its environment following administration. Usu-
ally, the drug is housed within a physical compartment, and
its rate of release through an orifice is controlled by either
driving water inside the device through osmosis, absorbing
water by a polymeric matrix that upon swelling exerts pres-
sure on the drug reservoir or by the use of a chemical that
vaporizes at body temperature, with the gas thus formed
exerting pressure to release drug from its compartment.
These systems allow release of the drug from the device
by a zero-order process (Chien 1985; Gupta et al. 2005).
11.4 Rate-Controlled Drug Delivery 171
An example of these activation technologies is the Oros sys-
tem (Alza Pharmaceuticals, Mountain View, CA; a subsid-
iary of Johnson & Johnson). Oros drug delivery is based on
osmosis and has wide clinical applications, including drugs
used in the treatment of attention-deficit hyperactivity dis-
order, benign prostatic hyperplasia, hypertension, diabetes
(type 2), overactive bladder, pain, and schizophrenia
(Conley et al. 2006). Some drug products using Oros include
albuterol, chlorpheniramine maleate, doxazosin mesylate,
glipizide, ivermectin, nifedipine, and verapamil (Gupta
et al. 2005).
Systems that depend on pH activation are covered by a
mixture of polymers that deliver the device intact to the
intestines. Under the pH conditions available in the intes-
tine, some polymers are soluble and others are not. This
causes the polymeric membrane surrounding the device
to become porous, allowing diffusion of the drug through
the pores. Ion activation systems rely on interaction of
the drug with a resin, which upon delivery to the gastroin-
testinal tract undergoes reactions with available ions to
release the drug slowly from its resin–drug complex and
to provide controlled delivery (Chien 1985). Insulin encap-
sulated in nanoaggregate particles (100–230 nm) and later
coated with weak polyelectrolytes [(Poly(α, ß-1-malic acid)
and chitosan] exhibited a pH-dependent release of insulin
from the nanoparticles due to the sensitivity of the polyelec-
trolytes to pH. At physiological blood pH (7.4), insulin
release from the matrix was observed, whereas at pH 4–5
no insulin release was detected. Furthermore, the addition
of a higher number of polyelectrolyte layers to the surface of
the nanoaggregates resulted in a reduction of the rate of
release of insulin from the nanoparticles (Fan et al.
2006). In addition to the pH, drug lipophilicity as measured
by the partition coefficient (log P) was shown to play an
important role in drug release (Sawant et al. 2010). For lido-
caine (log P = 2.6; lipophilic) and lidocaine hydrochloride
(log P = 0; hydrophilic), it was observed that when lido-
caine was incorporated into a hydrophilic matrix (hydroxy-
propyl cellulose), the release rate of the drug followed a
slow non-Fickian release rate, whereas the hydrophilic salt
of the drug experienced a burst effect for its release from the
matrix. When lidocaine was incorporated into a hydropho-
bic carrier (Eudragit E100), a burst effect was noted. It
appears that lidocaine interacted with hydroxypropyl cellu-
lose, which resulted in a slow release of the drug from the
matrix. No interaction between Eudragit E100 and lido-
caine was noted, which resulted in a burst effect (Sawant
et al. 2010).
Activation systems that depend on magnetism employ
pellets carrying a tiny magnet within them. The drug–pellet
system is coated with impermeable polymeric membrane to
the drug, except at the center of the pellet, where no coat is
applied. The drug is released from the uncoated center area,
172 11 Formulation Development Concepts
and its rate of release is accelerated using an external mag-
netic field (Chien 1985). Applications of magnet-containing
units may be helpful in situations where an increase in the
drug release rate is desirable at a certain point in time. For
example, insulin delivery through such a device would be
desirable, where the patient can activate delivery of insulin
when needed after meals (Langer 1985). Another possible
external activation energy source is ultrasonic waves.
A biodegradable polymeric matrix containing the drug
may be used, and the rate of polymer degradation is
enhanced by an ultrasonic device, thus facilitating release
of the drug from its matrix (Chien 1985).
11.5 Drug Delivery Technologies
for Improving Oral Delivery
The oral route of administration has been by far the most
convenient method of drug administration. Over 50% of
the drugs available on the market exist in this form (Jain
et al. 2005). The ease of delivery combined with a relatively
rapid onset of action, along with lower cost per dose unit,
makes this route an ideal way for augmenting the therapeu-
tic effects of drugs. One of the recent innovations in enhan-
cing the delivery of drugs via this route is the use of orally
disintegrating tablets (ODTs). While oral delivery of drugs
by conventional dosage forms (tablets, capsules, etc.)
requires the intake of water to facilitate swallowing the
solid dosage form, the rapid disintegration and dissolution
of ODTs in the mouth cavity eliminate the need of water
during administration. Examples of this dosage form are
Ondansetron ODF/Setofilm® (Lavasta Pharma FZ-LLC,
Dubai, UAE; for nausea) and Zolmitriptan Rapidfilm®
(Applied Pharma Research/Labtec, Switzerland/Germany;
for migraine headache). Characteristics that are considered
to be highly desirable for ODT’s bulk excipients are (Joshi
and Duriez 2004) (1) delivers good dispersion and fast dis-
solution (in seconds) upon administration, (2) provides an
agreeable mouthfeel while masking the bad taste of drugs,
(3) remains stable under various storage environmental
conditions (e.g. temperature, humidity, etc.), (4) allows ade-
quate and sufficient loading of the drug within its matrix,
and (5) resists the shocks/stresses imposed on it from
patient’s handling and during transportation. One ODT
technology uses a specialized lyophilization method (i.e.
Zydis, Catalent, New Jersey) in which the entire formula-
tion process is prepared within a blister compartment.
The Zydis fast-dissolving technology (within three seconds
when placed in an aqueous environment) has been shown
to improve the bioavailability of drugs, as well as the onset
of action, comparable to that of an intravenous injection
(Catalent Zydis brochure 2007). ODTs prepared by a direct
compression method produce tablets that disintegrate and
dissolve rapidly in the mouth cavity; however, the final
stage of dissolution and absorption occurs in the gastroin-
testinal (GI) tract after the patient swallows the resulting
“suspension” made of the disintegrated particles
(OraSolv, CIMA Labs, Inc., 2012, MN). ODT technology
can also offer a sustained-release delivery mechanism such
as the tablets developed by Eurand N.V. (2012) (AdvaTab,
Amsterdam, The Netherlands). AdvaTab tablets dissolve
quickly within the mouth cavity (15–30 seconds) and have
the capacity to carry large doses of drug (Eurand N.V. 2012).
In general, an ideal ODT unit should dissolve rapidly in a
matter of seconds within the mouth cavity and should have
a taste-masking property and a smooth mouth feel
(Hamilton and Lutz 2005). In addition to tablets, oral
fast-dissolving films or strips (OFDSs) are also available to
enhance the delivery of poorly water-soluble and permea-
ble APIs. The film is composed of cellulose-based polymers
or poly(ethylene oxide) containing strong flavoring agents
to mask the taste of added ingredients. Examples of these
oral strips are Triaminic, Chloraseptic, and Listerine (Ali
and Quadir 2007). The normal size of an OFDS is about
the dimension of a postage stamp. Upon contact with the
saliva, the film disintegrates instantaneously. Original for-
mulations of OFDSs were introduced on the marker as
breath-freshening films. These original breath-fresheners
were followed by introducing products for delivering vita-
mins, minerals, and dietary supplements. The advantages
of OFDSs are fast disintegration and dissolution, flexibility
of the film, and nonfriability (Hariharan and Bogue 2009).
The permeation of drugs through the GI tract after an
oral administration is crucial for delivering a sufficient
amount of the drug to the circulation in a timely manner.
This becomes even more important if the drug has a poor
permeability profile through the gut mucosa. A system
developed by Merrion Pharmaceuticals (2012), known as
GIPET (gastrointestinal permeation enhancement technol-
ogy), facilitates and enhances the absorption of poorly per-
meable drugs up to 200-fold. GIPET works via a proprietary
micelle–drug membrane transport enhancement method.
This technology is applicable to a wide range of drugs,
including biotechnology products. Because of its good
safety profile, the FDA has included GIPET in its GRAS
(generally regarded as safe) listing. Nanosplode, Lip’ral,
Lip’ral Synchronized Solubilizer Release, and Hydroance
(Lipocine, Inc., 2012, Salt Lake City, UT) are other technol-
ogies that facilitate the absorption of drugs that suffer from
poor absorption and/or a poor solubility profile from the GI
tract. [It is estimated that about 40% of all drugs have poor
solubility in water (Dubin 2005a; Chattopadhyay and She-
kunov 2006).] Following rapid disintegration from solid
dosage form (e.g. capsules), the drug molecules are
engulfed with specialized proprietary lipids that facilitate
drug absorption by the GI mucosa (Lipocine, Inc. 2012).
Another important area for delivering drugs is that to the
brain and the central nervous system (CNS), which is often

limited due to the presence of the blood–brain barrier
(BBB). The BBB is due to the presence of tight junctions
in the blood vessels of the CNS that do not permit drugs
to pass through them, natural protective measure for the
brain. LipoBridge (Genzyme Pharmaceuticals 2012, Massa-
chusetts) is a short-chain oligoglycerolipid compound that
reversibly opens the tight junctions, thus facilitating the
entry of drug molecules into the CNS and brain tissues. This
allows an effective concentration of drugs in the CNS and
brain to increase by a factor of up of 100 above normal.
The system appears to be safe and is excreted unchanged
without being metabolized (Genzyme Pharmaceuti-
cals 2012).
11.6 Drug Delivery Technologies
for Improving Transmucosal Delivery
Transmucosal delivery is seen as somewhat of an extension
to other routes. Via this system, drugs are transported
through a transmucosal pathway by placing them in the
mouth, nose, or another route. The market for transmuco-
sal delivery is estimated to be in billions of US dollars annu-
ally. The transport of drugs through the mucosa of the
mouth cavity is reported to be highly efficient, exceeding
that of the skin manyfold (4–4000 times higher). An exam-
ple of such a delivery system is the OraVescent technology
(CIMA Labs, 2012, Brooklyn Park, MN), which offers
mucosal transport of drugs from the mouth cavity (sublin-
gually or between the buccal and gingival tissues). OraVes-
cent is available in compressed tablet form and is dispensed
in a PakSolv proprietary blister package (Brown and
Hamed 2007). Other advantages of the buccal delivery over
the oral route are avoidance of the gastrointestinal degrada-
tion and the first-pass effect as well as the ability to deliver
the drug in patients who have difficulty swallowing
(Davidson and Rousset 2018).
Sodium alginate is another type of material found useful
through its bioadhesive properties, thus allowing delivery
of APIs through the GI tract and nasal mucosa. Sodium
alginate is a biodegradable natural polysaccharide that
forms viscous solutions upon dissolution in water
(20–400 cP). Its viscosity is pH-dependent, and it decreases
at pH values above 10. It can be used as a matrix in floating
beads and tablets of gastroretentive preparations. Research
on sodium alginate has also revealed its usefulness to pro-
vide or enhance the controlled-release delivery of drugs, as
well as delivering proteins, DNA, and various other carriers
(e.g. liposomes and microspheres) (Patel et al. 2007). With
respect to the mucoadhesive properties, various agents may
be used to provide such action. Among these agents are
chitosan (cationic), carbopol 934 (anionic), and hydroxy-
propyl methylcellulose (HPMC, nonionic). Majithiya
et al. (2008) studied the effect of these agents on the
11.7 Drug Delivery Technologies for Transdermal Delivery 173
mucoadhesive property of tablets using a rat stomach mem-
brane model. Of all the single agents tested, HPMC had the
best mucoadhesive strength. When other agents were
mixed with HPMC, a synergistic mucoadhesive effect was
seen. A combination of chitosan: carbopol (1 : 4) gave the
highest strength for mucoadhesion. Overall, it appears that
combination polymeric mixtures provide a stronger
mucoadhesive force than that of a single agent alone.
11.7 Drug Delivery Technologies
for Transdermal Delivery
The skin is the largest organ of the body. It presents a formi-
dable barrier for foreign substances to enter the body and for
endogenous ones to escape to the outside. The presence of ker-
atin in the stratum corneum prevents water from entering or
leaving the skin (except at sweat pores). For normal skin, the
pH is slightly acidic (pH 5–6), due to the presence of organic
acids on the surface of the skin, such as amino acids and fatty
acids. Transdermal delivery of the drug can be activated by
various physical mechanisms including iontophoresis, sono-
phoresis, electroporation techniques, Magnetophoresis, and
micro-needles (Lopes and Soares 2015; Sirisha and Sailaja
2018) (see Chapter 19). In general, transdermal drug diffusion
may be enhanced by a reduction in the diffusivity resistance
through (1) dissolution of drug in its vehicle; higher concen-
tration leads to higher flux of the drug through the stratum
corneum, (2) disruption of the lipid structure of stratum cor-
neum, which permits local diffusion enhancement of drug
molecules, (3) interaction with intercellular proteins, and
(4) improving drug partitioning into the stratum corneum,
which facilitates the diffusion of drug from the vehicle to
the stratum corneum (Sirisha and Sailaja 2018).
Since the first transdermal product appeared on the US
market in 1979 (Scopolamine for the control of nausea
and vomiting), the transdermal delivery applications of
drug products have gained interest in improving patient
compliance, avoiding hepatic first pass metabolism, and
reducing the occurrence of gastrointestinal side effects of
drugs. Systemic side effects are also reduced by providing
controlled release of the drug while enabling a steady blood
drug-level profile (Sirisha and Sailaja 2018).
Transdermal patches are often used to deliver drugs
through the skin. The solubility of drugs in the adhesive
material of the patch can be difficult to estimate. Myatt
et al. (2008) presented a practical method for estimating
the solubility of solutes in the adhesive matrix. In this
method, the amount of drug diffused over time is obtained
by diffusion experiments, and the data are then plotted to fit
Higuchi’s model:
M = k0 t0 5 11 1
174 11 Formulation Development Concepts
where M is the cumulative amount of drug released per unit
area, t is the time, and k0 is Higuchi’s constant. The value of
k0 is calculated at different starting concentrations of the
solute added to the adhesive matrix. When k0 values are
plotted against the initial concentration of the drug in the
matrix, the graph forms two distinct linear portions that
intersect at a point. The point of intersection corresponds
to the solubility of the drug in the adhesive matrix read
directly from the x-axis (Myatt et al. 2008).
11.8 Special Considerations for
Biotechnology and Protein Delivery
Systems
The biotechnology industry has two main components:
“upstream” processes and other related to “downstream”
processes. The upstream elements deal with materials
and issues related to producing the product, while down-
stream processes handle the product’s purification to its
final active form. Incidentally, downstream processing
comprises 80% of the total cost associated with generating
biotherapeutic agents (Endres and Dabre 2020). Biotech-
nology’s birth is believed to have occurred with the inven-
tion of recombinant DNA by Stanley N. Cohen and Herbert
Boyer in 1973 (Shaffer 2020). The biotechnology applica-
tions are numerous and include immunotherapy,
CRISPR-Cas9 gene editing (CRISPR stands for Clustered
Regularly Interspaced Short Palindromic Repeats, and
Cas9 is an RNA-guided endonuclease found in certain bac-
teria), and gene therapy (Shaffer 2020). CRISPR was ini-
tially discovered by Japanese scientists in bacteria during
the late 1980s and by another scientist from Spain in the
1990s while working on genetic material sequencing in
Archaea (single-celled prokaryotic organisms whose cells
exhibit undefined nucleus) (Behlke 2020). These repeats
in the bacterial genetic code served as a defense mechanism
for the bacteria to protect itself against subsequent viral
infections (i.e. bacterial immunity against viral infections)
(Behlke 2020). Emmanuelle Charpentier and Jennifer
Doudna were awarded the prestigious Nobel Prize in chem-
istry in 2020 for making the genetic editing machinery pos-
sible in vitro using the Cas9 enzyme system (obtained from
Streptococcus pyogenes) as a platform and applying it to
mammalian cells (Jinek et al. 2012). Their work has poten-
tial applications in a variety of areas such as agriculture and
medicine. Gene therapy employs, for the most part, viral
vectors (70% of the cases) to deliver genes to the host cells
(Lingelbach 2020). The types of viral vectors used include
adenoviral, lentiviral, and adeno-associated viruses
(Lingelbach 2020). Concerning the manufacturing of
viral-vectored gene therapy products, biotechnology
companies must consider three critical and essential areas:
(1) safety, (2) purity, and (3) consistency, with the manufac-
turing processes being scalable and reproducible (Washer
and Knop 2020). During pandemics, the pressure exerted
on biotechnology companies to produce billions of doses
rather than millions fast becomes a reality. Companies have
to re-invent operation areas such as quality control to meet
the global demand without compromising safety while
maintaining speed, capacity, and product quality
(Anonymous 2020b). During the COVID-19 pandemic,
the CRISPR technology was implemented to develop
FDA-approved rapid-testing platforms for the SARS-CoV-
2 virus (Straiton 2020).
The future of pharmaceuticals will include a great num-
ber of biotechnology products that are composed of pro-
teins. Sterility, purity, and stability (physical and
chemical) are the obvious challenges in this area of deliv-
ery. Each area should be addressed independently since a
stable product does not guarantee sterility or purity, and
vice versa. Considerations for storage conditions (tempera-
ture, cleanliness) and handling (shaking the container vs.
swirling) are also areas of concern for these delivery systems.
It is known in pharmacy practice that vigorous shaking of a
protein product produces breakage in the chain of the pro-
tein, rendering it denatured (Chan 2008). For biotechnology
products, their solutions may be filtered to remove any living
or nonliving organisms from the product. A 0.22 μm pore fil-
ter can be employed to remove bacteria, and nanofilters are
utilized to remove viruses from the solution. When it comes
to biotechnology agents and products, the term “biosimilar”
is used in place of “generic drug” due to their large molecular
structure, and the different manufacturing processes used
that do not assure consistency in their “identity, purity,
potency, and safety” (see Chapter 24) (Aziz 2020).
Another area of concern is that proteins in formulation
often aggregate in the form of dimers to multimers. The
dimers are usually soluble in pharmaceutical solvent sys-
tems (generally, aqueous in nature), while larger aggregates
may grow to sizes that become visible to the naked eye.
Aggregate formation is believed to give rise to immunoge-
nicity, as well as to create major challenges in the areas of
handling, production yield, and quality (Das and Nema
2008). Detection of particulate matter due to aggregation
of protein molecules can be detected by various measures,
including laser diffraction particle analyzers, dynamic
image analysis, imaging, and Raman spectroscopy (Das
and Nema 2008). Moreover, the pharmaceutical industry
utilizes various techniques for testing the structural integ-
rity of proteins during processing. These include capillary
electrophoresis, high-performance liquid chromatography
methods, mass spectrometry, glycosylation analysis, amino
acid analysis and primary sequencing by Edman chemistry,
spectroscopy techniques, differential scanning calorimetry,
 11.8 Special Considerations for Biotechnology and Protein Delivery Systems
dynamic and static light-scattering methods, and analytical
ultracentrifugation/sedimentation techniques, among
others (Chan 2008). Raman spectroscopy and Raman
microscopy are commonly used to monitor the stability
of proteins during processing, allowing in situ testing, as
well as testing of samples in a rapid manner by matching
Raman fingerprinting of proteins with that of samples.
However, Raman technology does not work well with lyo-
philized material or when the concentration of protein is
too low (Wen et al. 2010).
Routes of administration for biotechnology products are
also limited to the parenteral route, although pulmonary
and nasal delivery may be alternative options. For example,
the delivery of insulin is done primarily by syringes, pens,
jet injectors, and pumps (Shankar 2005) and attempts to
market insulin as a buccal spray or in inhalers have been
seen recently. Generex Biotechnology is already marketing
insulin as a buccal delivery device (Oral-lyn), which is
approved for use in several countries (Amin et al. 2008b).
Inhaled insulin preparations have been introduced to the
market, but for varying reasons (mainly safety issues, effi-
cacy, and cost) they were withdrawn by the manufacturer
within months of being offered (e.g. Exubera, Pfizer). Exu-
bera was a fine powder (particle size 1–5 μm) of bioavailable
human insulin. Novo Nordisk’s AERx Insulin Diabetes
Management System was halted in its development stage
before reaching the market. This system was an inhaled
insulin system in liquid form. Eli Lilly, in cooperation with
Alkermes, has been developing powdered insulin for inha-
lation that could be delivered using a simple inhaler. The
particles are large and have low bulk density, providing
them with good aerodynamic properties. Other devices
for inhaled insulin are also being developed by several com-
panies (Baxter BioPharma Solutions, BioSante Pharmaceu-
ticals, Kos Pharmaceuticals, and MannKind) (Amin
et al. 2008b).
Recent work in the area of oral delivery for peptides and
proteins is also showing some hopeful signs of progress,
although issues related to the reproducibility of delivery
remain a big obstacle. The presence of food in the GI tract,
the availability of active enzymatic processes, and the chal-
lenges facing absorption of high-molecular-mass molecules
by the gut mucosa are just a few factors that render oral
delivery of proteins and peptides the “holy grail” of drug
delivery. Another area of concern is that related to the sol-
ubility of API after being released at the site of administra-
tion. Improvement in the solubility of peptides and proteins
may be facilitated by the inclusion of counterions in the for-
mulation. Acetate and trifluoroacetic acid (TFA) are com-
monly used as counterions in protein formulations.
Higher residual TFA levels are of concern; however, they
are tightly regulated because they can cause undesirable
effects in patients. Also, modifications made to peptide
175
and protein structure can not only improve solubility but
can minimize aggregation and adsorption of the proteins
as well (Dubin 2008).
New approaches, such as Emisphere’s Eligen (2012,
Cedar Knolls, NJ), are promising technologies for oral
delivery of drugs that are otherwise limited to the paren-
teral route. This technology has been tested with many
important drugs, including calcitonin, growth hormone,
heparin, and insulin (Majuru 2005). It utilizes a proprietary
delivery agent that when given along with a drug orally
forms a complex in the GI tract, which changes the physical
characteristics of the drug, allowing it to be absorbed
through the mucosa. Following absorption, the complex,
which is held together by weak intermolecular forces, dis-
sociates, releasing the active drug to exert its pharmacolog-
ical effect.
Emisphere’s Eligen technology has been placed on hold
by the company with respect to developing it further as a
means of delivering insulin orally. In a statement on its
website, the company declared that “Oral insulin could
represent a significant opportunity for Emisphere and, in
theory, would meet unmet market needs. However, realis-
tically, at this stage in its development and based on what
has been and perhaps what has not been done with this
product candidate, it presents significant challenges. At this
time, the Company is exploring its strategic options for the
oral insulin compound.” This illustrates the types of chal-
lenges facing oral delivery of peptides and proteins.
Delivery of insulin via other routes, besides the paren-
teral route, has been investigated fiercely for many decades.
The main reason for this interest is its importance in man-
aging diabetes, a major health issue worldwide. Diabetic
patients are required to self-inject insulin preparations sev-
eral times a day, a painful and inconvenient way to main-
tain a normal lifestyle. Thus, developing alternative routes
of administration has been a priority in this field. The oral
route, in particular, is of greatest interest, as it provides a
way that mimics the body’s own method of insulin “deliv-
ery.” Insulin is a high-molecular-mass hormone of 5800 Da.
It is synthesized by the beta cells in the islets of Langerhans
of the pancreas and transported via the portal vein to the
liver. Similar transport of insulin to the liver occurs via
the oral route, which is the natural way for the body to han-
dle insulin. Unfortunately, the GI tract is an extremely hos-
tile environment for insulin, due to the presence of
enzymes capable of destroying this regulating hormone.
The main enzymes are pepsin, trypsin, and chymotrypsin.
Studies performed in vitro have shown that the effect of
these three enzymes is rapid (seconds to minutes) and
highly effective (80–100% degradation). Pepsin requires
an acidic environment to act, whereas trypsin and chymo-
trypsin need a slightly basic pH. This pH requirement cor-
responds to their location in the gut cavity; pepsin is located
176 11 Formulation Development Concepts
in the stomach, and the other two enzymes are found in the
small intestine. Another obstacle for the oral delivery of
peptides and proteins is the intestinal permeability. By
far the most important area for drug absorption is the duo-
denum. However, the absorbing epithelium is limited in its
action to smaller-molecular-mass drugs and more lipo-
philic solutes. As such, peptides and proteins are not good
candidates for absorption in this region, and thus they have
a very low oral bioavailability (Cheng and Lim 2008).
Recently, the French-based company Gattefossé has intro-
duced self-emulsifying drug delivery systems (SEDDS) that
work by solubilizing the peptides in the gastrointestinal
tract while protecting them from degradation by digestive
enzymes. The SEDDS have been promising applications
for oral delivery of peptides such as desmopressin, insulin,
and leuprorelin. SEDDS are also employed to enhance the
aqueous solubility of poorly soluble compounds, which in
turn render them suitable for oral administration. Solid
SEDDS may also be applicable for processes such as hot
melt extrusion and granulation.
Pharmaceutical companies commonly rely on the concept
of the human maximum absorbable dose (MADhuman) as a
measure of drug bioavailability. MADhuman measures the
oral dose absorbed (mg) (Dabs) that reaches the circulation
as compared to the oral dose administered (Timpe 2007):
Dabs = P S A T 11 2
where P is the apparent permeability coefficient (cm/s), S
the saturation solubility of an API (mg/mL), A the surface
area available for absorption (cm2), and T the transit time
through the small intestine (seconds). In humans, the esti-
mated values for A and T are 800 cm2 and 3.3 hours, respec-
tively. The Dabs desired is often stated as being 40–60% of
the efficacious dose (ED50) (Timpe 2007). It is expected that
the value of Dabs with respect to the oral absorption for
unaided peptides and proteins is very low or nearly zero.
A promising area for delivering proteins orally is the
entrapment of the API in a polymeric mesh, from which
the drug is released slowly overtime, as well as being pro-
tected from enzymatic degradation in the gut. An example
of a system that has produced positive experimental results
in this respect is the polymer system known collectively as a
thiomer. These polymers are essentially thiolated and pro-
vide a zero-order drug release kinetic from their matrix. The
matrix itself is made of a three-dimensional structure that
offers a tightened tablet matrix for the delivery of peptides
and proteins (Langoth and Bernkop-Schnürch 2005).
One area for protein delivery employs an encapsulated
ion of proteins in biodegradable micro- or nanoparticles.
The production of these particles involves a double-
emulsion method, in which the aqueous phase containing
proteins (w1) is emulsified with the polymeric material dis-
persed in an organic phase to form aw1/o emulsion.
A double emulsion (w1/o/w2) is then formed by the
addition of an aqueous solution (w2) containing 2%
poly(vinyl alcohol) (PVAL). The resulting double emulsion
is mixed with a PVAL aqueous phase (0.3%), and the
organic phase is evaporated to form a suspension of nano-
particles. An example of these microparticles is Novartis’s
Sandostatin LAR Depot (composed of a biodegradable glu-
cose star polymer with a copolymer of lactic and glycolic
acids) delivering the chemotherapeutic agent octreotide
acetate, used in the treatment of acromegaly (Bilati
et al. 2005).
Delivery of peptides and proteins through the pulmonary
route is associated with significant challenges regarding the
stability of proteins during transportation, use, and storage.
Metered-dose inhalers (MDIs) contain propellants that are
not compatible with most proteins. Thus, nebulizer solu-
tions containing unbuffered isotonic solutions with a pH
of 5 or higher are preferred over MDIs for delivering protein
drugs. Calcium salts (calcium chloride) are often used in
protein solutions as stabilizers. Surfactants such as Tween
80 are usually added to protein solutions to minimize the
adherence of protein to the container walls and to protect
the protein from aggregation and a rise in temperature dur-
ing nebulization. Instead of solutions, powdered aerosols
may be used where the protein is present in freeze-dried
form. To facilitate the flow of the protein’s powder, an inert
coarse powder carrier is usually mixed with the protein par-
ticles. Protein products should be protected from high tem-
peratures and humidity during storage, as these conditions
lead to denaturing the protein through aggregation reac-
tions. The bioavailability of protein drugs via the lungs
can be restricted, as proteolytic enzymes and the presence
of macrophages act to eliminate the protein and limit its
absorption (Krishnamurthy 1999).
Other areas of biotechnology products are related to gene
delivery. Liposomes are one of the preferred carriers for
genes, due to their potential low immunogenic reaction
(Felgner et al. 1987). In general, liposomes are classified
into three types: multilamellar vesicles (MLVs), small unila-
mellar vesicles (SUVs), and large unilamellar vesicles
(LUVs). With respect to size, MLVs range between 0.1
and 5.0 μm, whereas SUVs and LUVs are 0.02–0.05 μm
and greater than 0.6 μm in size, respectively. There are
two major methods of producing liposomes: by the forma-
tion of emulsions and by hydrating a dried lipid film fol-
lowed by vigorous agitation (sonication or high-shear
impeller) (Chrai et al. 2002). As a carrier for genes, lipo-
somes were first suggested by Felgner et al. (1987). The abil-
ity of liposomes to travel through cytoplasmic and nuclear
membranes is essential for delivering genes to the DNA. In
addition, genes thus delivered should be able to integrate
with the cell’s DNA to become effective. Cationic liposomes
are commonly used to deliver the plasmid gene, because
these liposomes were found to be most stable in vivo
(Bellare and Daga 2005).

Generally speaking, proteins are prepared as lyophilized
dried powders, in solutions, or as frozen preparations. The
latter has its own advantages and disadvantages. Frozen
protein preparations are desirable during batch manufac-
turing of a drug product. They reduce the waste associated
with the manufacture of these products, as well as assuring
that the end product meets its specifications during storage
and throughout its shelf life. In addition, the low freezing
temperature (−20 C) affords a hostile environment for
microorganisms and thus helps in contamination control
of the product. However, freezing and thawing cycles can
cause denaturation of protein. Also, solution heterogeneity,
commonly found in the frozen material, can result in phys-
ical and chemical alterations that have a detrimental effect
on a proteineous drug (Kantor et al. 2011).
11.9 Drug–Excipient and
Excipient–Excipient Interactions
The market for pharmaceutical excipients is expanding at
an accelerated rate and is expected to reach $8.53 billion
by 2023 (it was $6.4 billion in 2018) (Dubin 2019). The
choice of excipients in a formulation is not a marginal
matter. Excipients may contain undesirable contaminants
that need to be identified prior to incorporating them in
formulations. Guidelines for impurities of excipients are
traditionally listed in various pharmacopeias. For example,
the United States Pharmacopeia monograph for
poly(ethylene glycol) lists the limits for major and minor
contaminants such as ethylene glycol (0.25%) and ethylene
oxide (10 ppm) (Erickson 2005). The functions of excipients
in formulations are numerous and include roles as solubi-
lizing agents, stability enhancers, tonicity-adjusting compo-
nents, preservatives, and release-controlling materials,
among others (Dubin 2005a). Interactions among excipi-
ents in the same formulation or between excipients and
an API can lead to altered or diminished bioavailability
and/or product stability (Dubin 2005b). Isothermal stress
methods are often used to test for incompatibilities existing
among ingredients in the same formulation. High tempera-
tures with or without moisture are used as storage condi-
tions to determine how the various ingredients may
affect the stability of an API in a formulation. However,
the use of high temperatures during stress testing has its
limitations if the ingredients melt at elevated temperatures
(e.g. lipid components). For these formulations, extrapola-
tion from high temperatures to ambient temperatures may
be problematic (Holm 2010). Most pharmaceutical compa-
nies that operate on a limited tight budget choose already
acceptable excipient blends by the regulatory agencies for
their formulations. Otherwise, the company has to justify
11.9 Drug–Excipient and Excipient–Excipient Interactions 177
the excipient’s safety before use (Dubin 2005a; Eatmon
and Loxley 2009). The approved list for excipients for cer-
tain formulations may be even more limited, such as those
used in parenteral preparations (Dubin 2005a). Generally
speaking, pharmaceutical companies opt to use an excipi-
ent blend chosen from an FDA-approved excipients list.
Characteristics of a good excipient are compatibility with
the API and other added ingredients in a formulation, being
a nontoxic substance, and its availability on the market
with reasonable purity and an acceptable grade (Eatmon
and Loxley 2009). Some of the excipients used in biophar-
maceuticals may require specific handling. For example,
human serum albumin (HSA; 66,000 Da) is often used as
an excipient in biotechnology formulations. One of require-
ments of its use is that it be free of any viral contamination.
This is accomplished by heating HSA to 60 C for 10 hours.
This excipient has the advantage of being relatively stable at
room or higher temperatures, safe to use due to lack of
immunogenicity, and can be used to eliminate the adsorp-
tion of other proteins from the surface of containers. HSA
is also available from recombinant DNA technology
(Alburex, Mitsubushi Welfarma, Japan), which eliminates
the problem of virus contamination (Chaubal 2005).
Other considerations in drug development are related to
how excipients and drugs interact with each other in the
same formulation. These interactions may affect the phys-
ical characteristics of final products, such as the dissolu-
tion and disintegration time for tablets. For example, an
increase in the mixing time after adding magnesium stea-
rate (a lubricant) to a mixture of ketorolac tromethamine
with spray-dried lactose, cornstarch, pregelatinized
starch, or a combination, resulted in a decrease in the dis-
solution rate of the capsules containing the formulation
with cornstarch but not with that containing pregelati-
nized starch. The disintegration time of the capsules with
cornstarch increased with an increase in mixing time with
magnesium stearate but not with pregelatinized starch.
These effects on dissolution time and disintegration time
were explained by the interaction between cornstarch
agglomerates, magnesium stearate flakes, and drug parti-
cles, as documented by scanning electron microscopy. No
such interactions were seen with pregelatinized starch
(Chowhan and Chi 1985a). When crospovidone was used
in the formula, replacing cornstarch and pregelatinized
starch, similar effects were observed on the dissolution
and disintegration times. Scanning electron microscopy
revealed that drug particles get entrapped in crospovi-
done, and the resulting agglomerates interact with
magnesium stearate flakes as mixing time increased. Lar-
ger drug particles had more efficient entrapment with
crospovidone particles, resulting in a more pronounced
effect on dissolution and disintegration time (Chowhan
and Chi 1985b).
178 11 Formulation Development Concepts
11.10 The Presence of Contaminants
in a Formulation
Chemicals used in formulations are available in various
purity grades. The pharmaceutical industry utilizes those
with characteristics that match those approved by regula-
tory health agencies. In particular, preparations that are
intended for parenteral administration must contain che-
micals of the highest possible purity available on the mar-
ket, with no or an acceptably low content of contaminants
as determined by the health authorities. In certain cases,
such as the formulation of sterile single-use liquid biophar-
maceuticals, the development process requires the use of
stainless steel bulk storage tanks for an API that contains
several elements, such as aluminum, carbon, chromium,
cobalt, copper, iron, manganese, molybdenum, titanium,
tungsten, and zirconium. These elements may leach from
storage tanks and contaminate the product during the
development process (Leveen 2010). Analytical methods
for detecting impurities in APIs or starting materials are
based on various chromatography techniques such as
high-performance liquid chromatography (HPLC) and
supercritical fluid chromatography. The latter method
was found to be much superior over HPLC techniques in
detecting impurities in APIs and other starting additives
(Zelesky 2008).
Particulate matter contamination of sterile products is
another area of concern in the pharmaceutical industry.
Particulates can enter sterile products during processing,
packaging, or use. Glass, plastic, and rubber materials that
are used in vials and ampoules could contribute particulate
matter to the products housed within them. Other sources
of particulates include personnel entering and working in
sterile packaging areas, sealing operation for ampoules,
and gas purging of containers without a filtration step. It
is always easier to prevent contamination from occurring
than to clean a contaminated product. Accordingly, com-
bining multiple steps during manufacturing, such as filling,
sealing, and printing (in that order) under one-time opera-
tion may minimize the risk of introducing particulates into
the product (Dean 1985). Wormuth et al. (2019) conducted
a study on the presence of particulate matter in biopharma-
ceuticals with single-use systems. Their findings indicated
that all bag products tested contained particles within the
visible range. The authors advised that the manufacturer
should implement tightened controls to reduce the level
of particulate matter present to an acceptable level. The
United States, European, and Japanese pharmacopeias
require that the final parenteral product is free from visible
particles. Parenteral manufacturers do not release their
batches until 100% of the units produced are visually
inspected for particulate matter and other defects
(Laskina 2020). However, that does not preclude visible
particles from being present in the products released to
the public. It is estimated that about 50% of the recalls
issued in sterile parenteral products during the period
between 2010 and 2017 were due to this type of concern
(Laskina 2020). Parenteral vials have closures to protect
the product from contamination upon storage and hand-
ling. The United States Pharmacopeia addresses parenteral
container-closure integrity with guidelines found in chap-
ter 1207 (USP <1207>). Other forms of contaminants in
parenteral sterile products are leachables and extractables.
With increased use of newly synthesized polymers [poly-
ethylene, polypropylene, polyester-laminated polymers,
and poly(vinyl chloride)] as container materials for phar-
maceutical and biological products, leachables and extrac-
tables have become of great concern to the pharmaceutical
industry (Northup 2008). This is true not only for sterile
injectables but also for preparations intended for aerosol
inhalations, ophthalmic products, and nasal aerosol sprays
(Northup 2008). Regulatory agencies have published guid-
ing principles to suggest that the best way to test for leach-
ables is with the actual formulation (Scott et al. 2020).
Examples of leachables in orally inhaled and nasal drug pro-
ducts include the polycyclic aromatic hydrocarbons (PAHs),
n-nitrosamines, and 2-mercaptobenzothiazole (2-MBT). All
of these chemicals are considered to be carcinogens. PAHs
and n-nitrosamines appear frequently in the environment,
including in food products, soil, water, and air. 2-MBT resi-
dues are found in rubber products and are known to cause
allergic reactions in some people who wear rubber gloves
(Norwood and Mullis 2009). In general, leachables are
chemical substances (e.g. zinc ions) that migrate from a con-
tainer’s surface into a product concentrate during normal
storage and handling conditions; whereas extractables are
chemical substances (e.g. carbon-based compounds) that
can be obtained from a container’s surface after treatment
with solvents under varying conditions of temperature
and time (Castner et al. 2004). For example, in April 2010,
Johnson & Johnson recalled its children’s formulations, cit-
ing the presence of the pesticide and flame retardant 2,4,6-
tribromophenol in these preparations. This pesticide, which
originated from the shipping pellets, had permeated the
packaging walls, including the actual container of the prod-
uct, and reached the formulation (Lynch 2011). Recommen-
dations to overcome the presence of leachables and
extractables are to obtain information on the raw materials
and equipment used in making the containers, to utilize var-
ious methodologies in their identification, and to be aware of
their sources, which may be from outside the container itself
(Castner et al. 2004). Various analytical techniques can be
utilized in the detection of leachables and extractables.
The list includes single-crystal x-ray spectrometry, Fourier
transform infrared spectrophotometry, diode-array ultravi-
olet spectrophotometry, nuclear magnetic resonance

spectroscopy, and mass spectrometry. The latter is a widely
acceptable method for contaminant detection, due to its
high sensitivity and its ability to interface with gas and liquid
chromatographic techniques (Norwood et al. 2005).
All parenteral preparations must be free from microbial
contamination and their by-products, the endotoxins.
Several tests are available for testing for endotoxins during
in-process and at the final stage before releasing them to
distributors. Among those tests are the Limulus amebocyte
lysate test, monocyte activation test, and recombinant fac-
tor C test (Scott and Komski 2020).
Contamination presents in metered-dose inhalers is
another challenge facing the industry. Although contami-
nation can be traced to ingredients used in the product con-
centrate, the containers themselves may serve as the main
source. Inhaling contaminants into the lungs allows easy
access of potentially toxic materials to the blood circulation
via the alveolar sacs. Thus, proper control of extractables
and leachables from containers is an important safety con-
cern (Hansen 2005). Most important is screening for sub-
stances such as nitrosamines and polynuclear aromatic
hydrocarbons, as their presence is related to serious adverse
health concerns in humans. Quality control methods are
important for detecting the presence of these substances
at concentrations of nanograms per unit weight of the con-
tainer material or lower. The leachables profile over the
shelf life of the product should be determined, and agree-
ment with the level and type of extractables must be estab-
lished (Hansen 2005).
Although all impurities in pharmaceutical preparations
are of concern, those found in biologicals are in particular
worrisome. Biological impurities result from the presence
of undesirable protein components or nucleic acids that
can cause allergic reactions and can lead to mutagenicity
and carcinogenicity reactions. In particular, these concerns
are essential to consider if the product is not intended for
single use (Ciurczak 2020).
11.11 Other Considerations
Certain parameters define the “value” of a product. These
are included in the acronym ESCP: efficacy, safety, conven-
ience, and price (Eatmon and Loxley 2009). It is the efficacy
of a drug that makes it useful clinically. Among drugs with
similar efficacy, the one with the safest profile receives
favorable acceptance among patients and clinicians alike.
If efficacy is more crucial than safety (such as the case with
chemotherapeutic agents), clinicians are willing to use
drugs that are more efficacious, despite pronounced toxic
effect. Although patients prefer the use of products that
are more convenient to take (e.g. once vs. three times a
References 179
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ience. Convenience only becomes important when the
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Finally, pricing the product the right way by matching it
with the consumer’s buying capability is an important fac-
tor, in particular when efficacy and safety profiles are
comparable (Eatmon and Loxley 2009). Another term to
consider beside ESCP is “quality.” This is important when
it comes to healthcare issues, where the market demands
quality pharmaceutical/biopharmaceutical products to be
produced in a most efficient way (Scott 2019). From the
FDA’s point of view, when evaluating product quality,
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product, but rather it should be built into the product”
during development (Dziki 2008), often through clear
communication between R&D and the product’s manu-
facturing unit. For instance, when it comes to the quality
of products intended for pediatric administration, issues
such as heavy metals or residual solvents and the presence
of suspected or known genotoxic substances in the
formulation must be addressed by the manufacturer
(Huang 2020b).
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