Crystal Growth
Therapeutic agents are commonly formed
into crystalline forms for drug delivery.
Three-dimensional surface morphology andcrystal structure have a dramatic effect on
the manufacture, ease of delivery, bioavailability, dissolution rate, and efficacy of
crystalline drug forms. To tailor the growth
process to fit desired behavior, growthparameters such as temperature, pH,concentration, and additive levels need tobe optimized. In situ visualization of
crystallization has been conducted by
AFM to optimize the growth conditions in
producing a desired morphology, as wellas to study growth mechanisms anddefect formation.4–11
For example, researchers Yip and Ward
have used AFM to study in situ
crystallization characteristics of several
insulin forms: bovine insulin, LysB28ProB29
insulin, ultralente insulin, and insulinprotamine complexes. Due to the lowforces needed to image the insulin surface,
they used TappingMode imaging directly inthe crystallization liquor. Figure 1 shows theepitaxial growth of a screw dislocation thatwas observed over 11 hours on LysB28ProB29
insulin.5 The terraces measure approximately
30 angstroms in height, which is consistent
with the c-axis spacing of the insulin
hexamer of rhombohedral LysB28ProB29
insulin. Observing the crystallization
behavior in situ made it possible todetermine growth rates and observe defectformation in real-time. Step advancement
was observed to occur at 2 x 10-6
micrometers per second, which corresponds
to the attachment of approximately five unit
cells (15 Lys B28ProB29 hexamers) per second.
Defects appeared to be caused by large
insulin aggregates that were unable toalign properly with the crystal structure due
to poor mobility, thus forming dislocationsand voids in the growing terraces.
These observations were conducted tostudy differences in growth characteristics
between LysB28ProB29 insulin and wild-type
porcine/bovine insulin. Lys B28ProB29 insulin
differs from wild-type porcine/bovine
insulins due to a sequence inversion at theC-terminus in the B-chain. This sequence

inversion was designed to reduce the insulinmonomer association for better dissolution
properties. However, the sequence inversion
also produced differences in crystallizationbehavior, as observed by AFM. Lys B28ProB29
insulin was shown to have smaller
attachment energies (_G k), more roundedscrew dislocations, larger terrace widths,
and more persistent vacancies in the (001)plane. These differences can havesignificant influence on the crystal quality
and on its behavior as a therapeutic agent.

Polymorphism
The ability of a drug substance to form into
more than one crystalline form is calledpolymorphism. Different polymorphs possess
different physicochemical properties, which
affect solubility, dissolution, adsorption,
melting point, and stability.12 Thus, polymorphic characterization is an importantparameter in maintaining high productquality and reproducibility in thepharmaceutical industry.
In Yip and Ward’s study on insulin
discussed above, the polymorphic forms ofinsulin were identified by using the AFM to
map the crystal structure by imaging themolecular lattice spacings in all three
dimensions.4 At the University of
Nottingham, Danesh and coworkers used
the advanced PhaseImaging technique toidentify and map the distribution ofpolymorphs of the drug cimetidine.13, 14
Force-interaction studies (amplitude-phasedistance relationships) were then conducted
to identify the polymorphs based on
differences in hydrophobicity. In figure 2,
cimetidine polymorphs A and B are noteasily distinguished in the topographic
image, but their distribution is easily
characterized in the phase image. The
contrast in the phase images is most likely
due to the differences in hydrophobicity
between the polymorphs, which produces
a difference in the tip-sample interaction
due to variations in capillary force. This
contrast was investigated by conducting
the experiment with hydrophilic (plasmaetched) and hydrophobic (alkylsilane)functionalized probes.

Figure 2. Distribution of polymorphic forms A andB of the drug cimetidine conducted by
PhaseImaging with hydrophilic probes. The phase
image (right) shows the distribution of the A (dark)
and B (light) polymorphs that is not evident in thetopographic TappingMode image (left). The
phase image contrast is due to differences inhydrophobicity. 4µm scans. Images courtesy ofC. Roberts, University of Nottingham.

(1)

(2)

(3)

1.5
1
0.5
0StartingMicronisedSEDS(4)
Figure 3. Height (left column) and phase (rightcolumn) images of paracetamol formed into drug
particles by micronization and SEDS.
Figure 3-1: raw starting material showing
crystalline lamellae. Figure 3-2: micronizedparticle showing rough, irregular structure.
Figure 3-3: SEDS particles showing regular,smooth structure with 0.9nm crystalline steps.
Figure 3-4: Roughness of starting material,
micronized particle, and SEDS particle. Images
courtesy of Patel, Davies, and Roberts, Molecular
Profiles, UK; and Palakodaty, Gilbert, York,
Bradford Particle Design Ltd., UK. Reference 17.

Figure 4. Surface
morphology of a wetgranule of caffeine,lactose, and polymers.
Roughness measurements
provide information about
the formation process andphysicochemicalproperties. Inset images—Amplitude (left) andHeight (right) image. 5µm
scan. Image courtesy of T.
Li, K.R. Morris, and K.
Park, Purdue University.

Particles
Production of solid dosage forms commonly
begins with the formation of the drug into
particles, typically within the size of 0.1 to10 micrometers. Characterization of theseparticles can be important before drug
formulation since their morphology, size,and shape can provide information about
the manufacturing process. 15 Particle size
has also been shown to influence dissolution
rate, bioavailability, content uniformity,stability, texture, flow characteristics, andsedimentation rates, and thus has asignificant effect on formulation andtherapeutic efficiency.12
There are many methods commonly used toinvestigate particles, such as light-scattering
techniques like dynamic light scattering andlaser diffraction. However, these techniques
sample a large number of particles toprovide a distribution of particle size orcharacteristics.16 There are often cases
where studying the particles individuallybecomes a key step in understanding aparticle system. One common method for
directly studying individual particles istransmission electron microscopy (TEM).
However, often the sample preparation ofsmall particles for TEM is challenging andtime consuming.
AFM has successfully examined
pharmaceutical particles directly to correlate
their morphology to the manufacturing
process and behavioral properties. 15, 17
Figure 3 shows an example of using theAFM to characterize the drug particle
morphology. Drug particles are traditionallyformed by milling a drug crystal to particlesizes less than 10 micrometers by
micronization or spray-drying techniques.
However, problems can result from thesetechniques due to batch-to-batch variations,residual solvent, and statically-charged
particles that can affect powder stability
and flow.17 Another method of particle
formation called solution enhanced
dispersion by supercritical fluids (SEDS)
overcomes many of these problems andprovides more control of the particle size,
shape, and morphology. The top left AFM
image in figure 3-1 shows the starting raw
material of paracetamol in which crystalline
terraces can be seen. Images of particles
formed by micronization and SEDS are
shown in figures 3-2 and 3-3 respectively.

The micronized particles vary in size andare irregular with a significant amount ofsurface roughness, whereas, the SEDS
particles have a regular shape and a sizeof approximately 10 micrometers, andshow a reduction in roughness from the raw
starting material (see the bottom right imageof figure 3-4). As shown through AFM, thesmoother surfaces and regular shapes
produced by SEDS should reduce the batchto-batch variations and static charge
problems encountered with the micronizedmaterials, as well as improve the flow
properties of the particles.

Figure 5. Tablet coating showing complex
pore structure. Surface structure, roughness,and surface area can be easily
characterized. 5µm scan. Sample courtesyof ALZA Corporation.

Coatings
GranulesOnce the drug is in particulate form, it isoften formed into a granule by mixing thedrug particles with binding agents, diluents,
and disintegrating agents. The wet
granulation process consists of adding aliquid binder or adhesive to the mixture,
passing the wetted mass through a screen
sieve of the desired mesh size, and drying
the granules. The resulting granules aretypically in the range of a few millimeters,
and show improvements in flow properties
as well as chemical and physical stability
with respect to particles. AFM has beenvery successful in characterizing themorphology and roughness of granules tocorrelate their surface structure to theunderlying physicochemical and mechanicalprocesses during the manufacturing process(see figure 4). 15

There are many coatings that may beapplied to tablets to serve various
purposes. Common uses ofpharmaceutical coatings consist ofprotecting the drug from air andhumidity, providing a barrier to an
objectionable tasting or smelling drug,and controlling the dissolution behavior.
Sugar coatings are very commonly
applied to tablets, as well as polymer
coatings, which are more durable, lessbulky, and less time-consuming toapply. The polymer coatings are often
designed to rupture in thegastrointestinal tract to avoid stomach
irritation and to improve drug
adsorption. Coating granules and otherdrug substances are also key steps inthe design of controlled-release andmicroencapsulated dosage forms. 12