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Crystallography
Crystallography is the experimental science of determining the arrangement of atoms in the
crystalline solids (see crystal structure). The word "crystallography" derives from
the Greek words crystallon "cold drop, frozen drop", with its meaning extending to all solids with
some degree of transparency, and grapho "I write". In July 2012, the United Nations recognised
the importance of the science of crystallography by proclaiming that 2014 would be the
International Year of Crystallography. X-ray crystallography is used to determine the structure of
large biomolecules such as proteins.
Before the development of X-ray diffraction crystallography (see below), the study of crystals was
based on physical measurements of their geometry. This involved measuring the angles of crystal
faces relative to each other and to theoretical reference axes (crystallographic axes), and
establishing the symmetry of the crystal in question. This physical measurement is carried out
using a goniometer. The position in 3D space of each crystal face is plotted on a stereographic
net such as a Wulff net or Lambert net. The pole to each face is plotted on the net. Each point is
labelled with its Miller index. The final plot allows the symmetry of the crystal to be established.
Crystallographic methods now depend on analysis of the diffraction patterns of a sample targeted
by a beam of some type. X-rays are most commonly used; other beams used
include electrons or neutrons. This is facilitated by the wave properties of the particles.
Crystallographers often explicitly state the type of beam used, as in the terms X-ray
crystallography, neutron diffraction and electron diffraction. These three types of radiation interact
with the specimen in different ways.

X-rays interact with the spatial distribution of electrons in the sample.

Electrons are charged particles and therefore interact with the total charge distribution of
both the atomic nuclei and the electrons of the sample.

Neutrons are scattered by the atomic nuclei through the strong nuclear forces, but in
addition, the magnetic moment of neutrons is non-zero. They are therefore also scattered
by magnetic fields. When neutrons are scattered from hydrogen-containing materials, they
produce diffraction patterns with high noise levels. However, the material can sometimes be
treated to substitute deuterium for hydrogen.

Because of these different forms of interaction, the three types of radiation are suitable for
different crystallographic studies.

Theory
An image of a small object is made using a lens to focus the beam, similar to a lens in a
microscope. However, the wavelength of visible light (about 4000 to 7000 ngstrm) is
three orders of magnitude longer than the length of typical atomic bonds and atoms themselves

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(about 1 to 2 ). Therefore, obtaining information about the spatial arrangement of atoms requires
the use of radiation with shorter wavelengths, such as X-ray or neutron beams. Employing
shorter wavelengths implied abandoning microscopy and true imaging, however, because there
exists no material from which a lens capable of focusing this type of radiation can be
created. (Nevertheless, scientists have had some success focusing X-rays with
microscopic Fresnel zone plates made from gold, and by critical-angle reflection inside long
tapered capillaries.) Diffracted X-ray or neutron beams cannot be focused to produce images, so
the sample structure must be reconstructed from the diffraction pattern. Sharp features in the
diffraction pattern arise from periodic, repeating structure in the sample, which are often very
strong due to coherent reflection of many photons from many regularly spaced instances of
similar structure, while non-periodic components of the structure result in diffuse (and usually
weak) diffraction features - areas with a higher density and repetition of atom order tend to reflect
more light toward one point in space when compared to those areas with fewer atoms and less
repetition.
Because of their highly ordered and repetitive structure, crystals give diffraction patterns of
sharp Bragg reflection spots, and are ideal for analyzing the structure of solids.

Notation

Coordinates in square brackets such as [100] denote a direction vector (in real space).

Coordinates in angle brackets or chevrons such as <100> denote a family of directions

which are related by symmetry operations. In the cubic crystal system for
example, <100> would mean [100], [010], [001] or the negative of any of those directions.

Miller indices in parentheses such as (100) denote a plane of the crystal structure, and
regular repetitions of that plane with a particular spacing. In the cubic system, the normal to
the (hkl) plane is the direction [hkl], but in lower-symmetry cases, the normal to (hkl) is not
parallel to [hkl].

Indices in curly brackets or braces such as {100} denote a family of planes and their
normals which are equivalent in cubic materials due to symmetry operations, much the way
angle brackets denote a family of directions. In non-cubic materials, <hkl> is not necessarily
perpendicular to {hkl}.

Technique
Some materials that have been analyzed using crystallography, such as proteins, do not occur
naturally as crystals. Typically, such molecules are placed in solution and allowed to slowly
crystallize through vapor diffusion. A drop of solution containing the molecule, buffer, and
precipitants is sealed in a container with a reservoir containing a hygroscopic solution. Water in
the drop diffuses to the reservoir, slowly increasing the concentration and allowing a crystal to

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form. If the concentration were to rise more quickly, the molecule would simply precipitate out of
solution, resulting in disorderly granules rather than an orderly and hence usable crystal.
Once a crystal is obtained, data can be collected using a beam of radiation. Although many
universities that engage in crystallographic research have their own X-ray producing
equipment, synchrotrons are often used as X-ray sources, because of the purer and more
complete patterns such sources can generate. Synchrotron sources also have a much higher
intensity of X-ray beams, so data collection takes a fraction of the time normally necessary at
weaker sources. Complementary neutron crystallography techniques are used to identify the
positions of hydrogen atoms, since X-rays only interact very weakly with light elements such as
hydrogen.
Producing an image from a diffraction pattern requires sophisticated mathematics and often an
iterative process of modelling and refinement. In this process, the mathematically predicted
diffraction patterns of an hypothesized or "model" structure are compared to the actual pattern
generated by the crystalline sample. Ideally, researchers make several initial guesses, which
through refinement all converge on the same answer. Models are refined until their predicted
patterns match to as great a degree as can be achieved without radical revision of the model.
This is a painstaking process, made much easier today by computers.
The mathematical methods for the analysis of diffraction data only apply to patterns, which in turn
result only when waves diffract from orderly arrays. Hence crystallography applies for the most
part only to crystals, or to molecules which can be coaxed to crystallize for the sake of
measurement. In spite of this, a certain amount of molecular information can be deduced from
patterns that are generated by fibers and powders, which while not as perfect as a solid crystal,
may exhibit a degree of order. This level of order can be sufficient to deduce the structure of
simple molecules, or to determine the coarse features of more complicated molecules. For
example, the double-helical structure of DNA was deduced from an X-ray diffraction pattern that
had been generated by a fibrous sample.

In materials science
Crystallography is used by materials scientists to characterize different materials. In single
crystals, the effects of the crystalline arrangement of atoms is often easy to see macroscopically,
because the natural shapes of crystals reflect the atomic structure. In addition, physical
properties are often controlled by crystalline defects. The understanding of crystal structures is an
important prerequisite for understanding crystallographic defects. Mostly, materials do not occur
as a single crystal, but in poly-crystalline form (i.e., as an aggregate of small crystals with different
orientations). Because of this, the powder diffraction method, which uses diffraction patterns of
polycrystalline samples with a large number of crystals, plays an important role in structural
determination.

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Other physical properties are also linked to crystallography. For example, the minerals
in clay form small, flat, platelike structures. Clay can be easily deformed because the platelike
particles can slip along each other in the plane of the plates, yet remain strongly connected in the
direction perpendicular to the plates. Such mechanisms can be studied by
crystallographic texture measurements.
In another example, iron transforms from a body-centered cubic (bcc) structure to a face-centered
cubic (fcc) structure called austenite when it is heated. The fcc structure is a close-packed
structure unlike the bcc structure; thus the volume of the iron decreases when this transformation
occurs.
Crystallography is useful in phase identification. When performing any process on a material, it
may be desired to find out what compounds and what phases are present in the material. Each
phase has a characteristic arrangement of atoms. X-ray or neutron diffraction can be used to
identify which patterns are present in the material, and thus which compounds are present.
Crystallography covers the enumeration of the symmetry patterns which can be formed by atoms
in a crystal and for this reason has a relation to group theory and geometry.

Biology
X-ray crystallography is the primary method for determining the molecular conformations of
biological macromolecules, particularly protein and nucleic acids such as DNA and RNA. In fact,
the double-helical structure of DNA was deduced from crystallographic data. The first crystal
structure of a macromolecule was solved in 1958, a three-dimensional model of the myoglobin
molecule obtained by X-ray analysis.[3] The Protein Data Bank (PDB) is a freely accessible
repository for the structures of proteins and other biological macromolecules. Computer programs
such as RasMol or Pymol can be used to visualize biological molecular structures. Neutron
crystallography is often used to help refine structures obtained by X-ray methods or to solve a
specific bond; the methods are often viewed as complementary, as X-rays are sensitive to
electron positions and scatter most strongly off heavy atoms, while neutrons are sensitive to
nucleus positions and scatter strongly even off many light isotopes, including hydrogen and
deuterium. Electron crystallography has been used to determine some protein structures, most
notably membrane proteins and viral capsids.

Crystallization
Although crystallography can be used to characterize the disorder in an impure or irregular
crystal, crystallography generally requires a pure crystal of high regularity to solve the structure of
a complicated arrangement of atoms. Pure, regular crystals can sometimes be obtained from
natural or synthetic materials, such as samples of metals, minerals or other macroscopic
materials. The regularity of such crystals can sometimes be improved with macromolecular
crystal annealing and other methods. However, in many cases, obtaining a diffraction-quality
crystal is the chief barrier to solving its atomic-resolution structure.

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Small-molecule and macromolecular crystallography differ in the

range of possible techniques used to produce diffraction-quality
crystals. Small molecules generally have few degrees of
conformational freedom, and may be crystallized by a wide range
of methods, such as chemical vapor
deposition and recrystallization. By contrast, macromolecules
generally have many degrees of freedom and their crystallization
must be carried out to maintain a stable structure. For example,
proteins and larger RNA molecules cannot be crystallized if their
tertiary structure has been unfolded; therefore, the range of
crystallization conditions is restricted to solution conditions in
which such molecules remain folded.
Protein crystals are almost always grown in solution. The most
common approach is to lower the solubility of its component
molecules very gradually; if this is done too quickly, the molecules
will precipitate from solution, forming a useless dust or
amorphous gel on the bottom of the container. Crystal growth in
solution is characterized by two steps: nucleation of a
microscopic crystallite (possibly having only 100 molecules),
followed by growth of that crystallite, ideally to a diffraction-quality crystal. The solution conditions
that favor the first step (nucleation) are not always the same conditions that favor the second step
(subsequent growth). The crystallographer's goal is to identify solution conditions that favor the
development of a single, large crystal, since larger crystals offer improved resolution of the
molecule. Consequently, the solution conditions should disfavor the first step (nucleation)
but favor the second (growth), so that only one large crystal forms per droplet. If nucleation is
favored too much, a shower of small crystallites will form in the droplet, rather than one large
crystal; if favored too little, no crystal will form whatsoever. Other approaches involves,
crystallizing proteins under oil, where aqueous protein solutions are
dispensed under liquid oil, and water evaporates through the layer

Three methods of preparing

of oil. Different oils have different evaporation permeabilities,

crystals, A: Hanging drop. B:

therefore yielding changes in concentration rates from different

Sitting drop. C: Microdialysis

percipient/protein mixture. The technique relies on bringing the protein directly into the nucleation
zone by mixing protein with the appropriate amount of percipient to prevent the diffusion of water
out of the drop.
It is extremely difficult to predict good conditions for nucleation or growth of well-ordered
crystals.] In practice, favorable conditions are identified by screening; a very large batch of the
molecules is prepared, and a wide variety of crystallization solutions are tested. Hundreds, even
thousands, of solution conditions are generally tried before finding the successful one. The
various conditions can use one or more physical mechanisms to lower the solubility of the
molecule; for example, some may change the pH, some contain salts of the Hofmeister series or

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chemicals that lower the dielectric constant of the solution, and still others contain large polymers
such as polyethylene glycol that drive the molecule out of solution by entropic effects. It is also
common to try several temperatures for encouraging crystallization, or to gradually lower the
temperature so that the solution becomes supersaturated. These methods require large amounts
of the target molecule, as they use high concentration of the molecule(s) to be crystallized. Due
to the difficulty in obtaining such large quantities (milligrams) of crystallization-grade protein,
robots have been developed that are capable of accurately dispensing crystallization trial drops
that are in the order of 100 nanoliters in volume. This means that 10-fold less protein is used per
experiment when compared to crystallization trials set up by hand (in the order of 1 microliter).
Several factors are known to inhibit or mar crystallization. The growing crystals are generally held
at a constant temperature and protected from shocks or vibrations that might disturb their
crystallization. Impurities in the molecules or in the crystallization solutions are often inimical to
crystallization. Conformational flexibility in the molecule also tends to make crystallization less
likely, due to entropy. Ironically, molecules that tend to self-assemble into regular helices are often
unwilling to assemble into crystals. Crystals can be marred by twinning, which can occur when a
unit cell can pack equally favorably in multiple orientations; although recent advances in
computational methods may allow solving the structure of some twinned crystals. Having failed to
crystallize a target molecule, a crystallographer may try again with a slightly modified version of
the molecule; even small changes in molecular properties can lead to large differences in
crystallization behavior.

Radiation Safety
X-rays produced by the diffractometer pose a very real danger. Human
tissue can be
severely damaged if exposed to the x-rays used in x-ray diffraction. Longterm,
chronic exposures at moderate levels can directly cause a variety of skin
disorders, and
chronic relatively low- level exposures may be a factor resulting in
increased cancer
risk in exposed personnel. Although modern XRD equipment includes
safeguards designed to
minimize or eliminate radiation in the work environment, an awareness of
the dangers of
radiation exposure and associated safety issues is required for anyone
using the x-ray
diffractometer.

Interaction of X-Rays with Matter

X-rays are a type of electromagnetic radiation that interacts with matter.
That interaction takes

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place as either absorption (transfer of energy from the X-ray photon to

the absorbing material) or
scattering (in which the X-ray photon is redirected by interaction with
the scattering material).
The process of scattering is primary in producing diffraction data from
test specimens, but both
processes result in the production of a variety of potentially damaging
secondary radiation that
can produce significant short- and long-term health effects in the event of
exposure of human
tissue.
The X-rays produced for diffraction analysis consist of characteristic
radiation dependent on the
anode target plus the continuous spectrum. Recall that the energy of Xrays and their wavelength
are inversely proportional (higher energy = lower wavelength), and that
the continuous spectrum minimum wavelength decreases as the
accelerating voltage (kV) of the X-ray source increases. Also important to
understand is that an increase in filament current (ma) and kV (beyond
the
minimum value required to produce characteristic radiation for the
target)
will result in an increase in the intensity of the generated X-rays but will
not change their energy.

Radiation Sources in the X-Ray Diffraction

Laboratory
The x-ray diffractometer is a source of very intense radiation, and
exposure of any part of the
body to the primary beam can deliver hundreds of times the maximum
permissible yearly dose in
a second, and an hour of exposure to the diffracted beam can result in a
years worth of
permissible exposure. Whereas a primary beam exposure will produce
noticeable skin damage
very quickly, a diffracted beam exposure might not be physically noticed
at all.

Working Safely with X-rays

It is imperative that anyone working with the x-ray diffractometer be
properly trained and
knowledgeable of its use in order to prevent injuries. If you do not feel
you are properly trained
to use the diffractometer, seek appropriate assistance from a competent
and experienced user.
To minimize (and hopefully eliminate) the likelihood of exposed to
radiation in the laboratory,

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our x-ray diffractometer includes many safety-related features, including

Complete shielding of the X-ray source and diffracted beam such that all
radiation is
contained within an enclosed area.
Fail-safe safety interlocks installed on the x-ray chamber that prevent
accidental opening of
the chamber door. If the door is forced open, the X-ray source is turned
off. The x-ray
source will not operate if the door is open.
A fail-safe indicator light should be present which is illuminated when
the x-ray source is
turned on.
Sealed openings of the chamber to eliminate any possible leakage.
Under no circumstances should anyone attempt to override any of the
safety features of the x-ray
diffractometer.
The system should be checked for leakage periodically with the system at
operating conditions
using properly calibrated survey equipment. The system should also be
checked if any of the
following conditions exist:
Any change in the components of the system (with the exception of
minor components such
as scattering and receiving slits).
Prior to and after any maintenance requiring the disassembly or
removal of a component in
the system
Anytime a visual inspection of the system reveals an abnormal
condition.
For routine operations of modern X-ray powder diffractometers, the
radiation exposure in
routine operations should be virtually nonexistent.

Background Information
X-ray diffraction techniques are some of the most useful in the
characterization of crystalline
materials, such as metals, intermetallics, ceramics, minerals, polymers,
plastics, or other inorganic
or organic compounds. X-ray diffraction techniques can be used to
identify the phases present in
samples from raw starting materials to finished product and to provide
information on the
physical state of the sample, such as grain size, texture, and crystal
perfection. Most x-ray
diffraction techniques are rapid and nondestructive; some instruments
are portable and can be
transported to the sample. The sample may be as small as an airborne
dust particle or as large as

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an airplane wing. In general, x-ray analysis is restricted to crystalline

materials, although some
information may be obtained on amorphous solids and liquids. XRD
samples are acceptable in
many forms, depending on the availability of the material and the type of
analysis to be
performed. Single crystals from a few microns to a few inches in diameter
or loose or
consolidated aggregate of many small crystals can be used. Although the
overall size of the
sample may be large, the actual area of the sample examined in a given
experiment rarely exceeds
1 cm2.
The type of information obtained from x-ray diffraction studies ranges
from sample composition
to details of the crystal structure or the state of orientation of the
crystallites. Phase identification
can be conducted on virtually all single crystal or powder samples. Also
useful are measurements
of the physical state of a sample that detect differences from the ideal
crystal (presence of defects,
strain, etc.).

X-ray Powder Diffraction

X-ray powder diffraction techniques are used to characterize samples in
the form of loose
powders or aggregates of finely divided material. These techniques cover
various investigations,
including qualitative and quantitative phase identification and analysis,
determination of
crystallinity, microidentification, lattice-parameter determinations, hightemperature studies, thin
film characterization, and, in some cases, crystal structure analysis. The
powder method is
perhaps best known for its use as a phase characterization tool partly
because it can routinely
differentiate between phases having the same chemical composition but
different crystal
structures (polymorphs). Although chemical analysis can indicate that the
empirical formula for a
given sample is FeTiO3, it cannot determine whether the sample is a
mixture of two phases (FeO
and one of the three polymorphic forms of TiO2) or whether the sample is
the single-phase
mineral FeTiO3 or ilmenite. The ability of x-ray powder diffraction to
perform such
identifications more simply, conveniently, and routinely than any other
analytical method explains

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its importance in many industrial applications as well as its wide

availability and prevalence.
In general, an x-ray powder diffraction characterization of a substance
consists of placing a
powder sample of a crystalline material in a collimated monochromatic
beam of x-radiation.
Chapter 3 of your textbook describes the process of x-ray diffraction, and
how the XRD finds the diffraction pattern
for your material. The diffraction pattern is recorded on film or using
detector techniques, then
analyzed to provide x-ray powder data.
In x-ray powder diffraction analysis, samples usually exist as finely
divided powder (usually less
than 44 m in size) or can be reduced to powder form. The particles in a
sample comprise one or
more independently diffracting regions that coherently diffract the x-ray
beam. These small
crystalline regions are termed crystallites. Consolidated samples, such as
ceramic bodies or asreceived
metal samples, will likely have crystallites small enough to be useful for
powder
diffraction analysis. The size limitation is important in x-ray powder
diffraction because most
applications of powder diffraction rely on x-ray signals from a statistical
sample of crystallites.
Braggs Law (Equation 1) states that the angular position, , of the
diffracted x-ray beam depends
on the spacing, d, between planes of atoms in a crystalline phase and on
the wavelength, , of the
x-ray.
n = 2d sin
The intensity of the diffracted beam depends on the arrangement of the
atoms on these planes.
A powder pattern from a single-phase material will have maxima at
positions dictated by the size
and shape of its unit cell and will increase in complexity as the symmetry
of the material decreases, as shown in Figure 1. For example, many metal
patterns and those from simple compounds that tend to be mostly of
cubic symmetry and have small unit cell edges will produce powder
patterns having fewer lines or maxima than would be expected from a
compound of lower symmetry or one having a very large unit cell. A
pattern of a mixture of phases in which all the individual patterns are
superimposed will produce a complex experimental pattern, especially
when the number of phases present in the mixture exceeds approximately
three or when the phases constituting the mixture are all of very low
symmetry or have very large unit cell dimensions.

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Figure 1. Two superimposed XRD graphs. Note that the CH 4 hydrate has
distinctly more
peaks than Ice (Ih), since it has a much more complex crystalline structure.

Phase identification using x-ray powder diffraction is based on the unique

pattern produced by
every crystalline phase. Much as a fingerprint is unique for each person,
the diffraction pattern can
act as an empirical fingerprint for that phase, and qualitative
identification of phases can be
accomplished by pattern-recognition methods that include established
manual techniques and the
newer methods that use computers, most of which implement programs
based on heuristic
algorithms. All of these methods make use of the database maintained by
the JCPDS
International Centre for Diffraction Data.

The Conventional Powder Diffractometer

Although film techniques have been in use since the inception of x-ray
diffraction, the advent of
powder diffractometer has been more recent. Powder diffractometers
have been in use as a
laboratory tool since the late 1950s and early 1960s.
The Bragg-Brentano geometry is used most commonly for powder
diffractometers. Figure 2
shows this geometry as used for vertical diffractometers. In the vertical
geometry, the axis A is
horizontal.
In the variation of the Bragg-Brentano geometry that the Olin XRD
system uses, termed the -
diffractometer geometry, the sample is held stationary with its surface in
a horizontal plane as the
x-ray tube and the detector move.

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Sample Preparation
Preparation of Powder Samples
Use fine powder if possible. If your powder is coarse (>240 grit or xxx
microns), grind it with
a mortar and pestle or a ball mill.
2. If only a small quantity of powder is available, skip to Step 3. If a large
quantity of powder is
available, fill the recess of an aluminum sample holder with powder and
use a flat object, e.g.,
a glass slide, to level the powder with the top face of the sample holder.
Skip to step 4.
3. If you dont have enough powder to fill the recess of an aluminum
sample holder, youll need
to deposit some of your powder onto a glass slide. Mix a few drops of
amyl acetate, acetone,
or methanol with some of your powder to create a paste. Spread a thin,
even layer of the
paste onto the middle of a glass slide. Its important to maintain a fairly
flat sample surface, so
try to avoid bumps and chunks. When youre satisfied with the deposited
layer, wait for the
solvent to dry.
1.

Preparation of Solid Samples

Prepare a flat sample with approximate dimensions of 1 wide x 2 long
x 3/16 (maximum)
thick.
2. Remove all dirt, grease, etc. from the surface of the solid.
3. Load your sample.
1.

Instrument Operation
This section will walk you through the basic operation of the x-ray
diffractometer. Youll have to
follow a series of tasks that includes
Powering up the instrument,
Loading your sample,
Viewing system parameters and calibrating the instrument,
Setting your testing conditions,
Collecting x-ray diffraction data for your sample, and
Analyzing your results.
It may seem boring at times, but itll be worth it! Just think of yourself as
a mad scientist armed

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with the power to identify completely unknown samples. Is it mullite (kind

of like a fossilized
mullet)? Is it diamond? Is it ice-nine? Only your analysis will tell.

Instrument Startup
Turn on the water chiller. The switch is on the right side of the front
panel.
2. Turn on the XRD main power supply. The switch is on the left side of
the instrument.
3. Turn on the computer and login as guest to the this computer
domain (not
MILKYWAY). You dont need a password.
1.

Sample Loading
Load your sample, as follows:
1. Open the XRD chamber door. Inside the chamber, you will see a
complex hardware system
with a large sample area (in the middle) and two arms that rotate around
the sample stage
area. The x-ray source is mounted on the left arm, and the x-ray detector
is mounted on the
right arm. Unless you know what youre doing, you shouldnt touch
anything connected to
the rotating arms.
2. Locate the sample mounting fixture in the center of the stage assembly.
The mounting
fixture is a half cylinder with two spring clips (on the underside) that
protrudes from the top
center of the stage assembly.
3. With the sample facing up and toward you, slide your sample holder
between the half-cylinder
and the spring clips. If you are using an aluminum sample holder, you
should center the
holder (left and right edges flush with the half cylinder) and slide the
holder all the way back.
4. Be careful not to get any of your powder on the XRD hardware. If you
do, please wipe it off
with a soft tissue or paper towel.