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Sample
Considerations
Fe-concentration
The 57Fe isotope
occurs in 2.2 % natural abundance. A thorough study, involving about 10
spectra taken in various applied magnetic fields over a wide temperature
range, requires either a frozen solution sample containing ca. 1 mM 57Fe in ca. 0.5 mL volume or a solid
sample containing ca. 40 μg/ cm2
57-Fe. Using natural abundance iron corresponds to ca. 50 fold increase in these numbers.
Especially in solvents with low
melting points, 50 mM solution samples pose added
solubility problems. For instance, a compound that is well dissolved at
room temperature may precipitate before the sample freezes, leading to an
inhomogeneous sample, which (to make things more complicated) possibly
contains a mixture of molecules with slow, medium and fast relaxing
electronic spins. To guard against this problem, the sample can be cooled
to just above the solvent freezing point. At this temperature the
supernatant, along with slightly more solvent for good measure, should be
transferred to a Mössbauer cell.
If the species is paramagnetic,
solubility is not the only consideration. For molecules with large spins, a
sample concentration exceeding 5 mM can lead to
intermolecular spin-spin interactions (In a non-glassforming
solvent this can happen even at 100 mM).
Such interactions then have to be overcome by a combination of strong
applied fields (B > 6.0 T) and very low temperature (1.5 K). Similar
problems are often encountered in polycrystalline samples, which initially
seem attractive due to the advantage of high purity and frequently added
structural information. Additional problems include alignment of the
crystallites by the applied magnetic field (This is likely if the electronic
ground state has a very anisotropic magnetic moment, which is the case for
many paramagnetic compounds). Grinding the crystallites in a mortar
generally does not solve the problem; however, grinding the sample in a
powder mill with a corundum ball often reduces the size of the crystallites
sufficiently to prevent texture. Orientation of the resulting crystallites
by the applied field is best prevented by embedding them in a
non-dissolving solvent or mineral oil such as nujol;
the powder should be WELL stirred to provide an absorber with uniform mass
density.
Solvent
Finally, it should be noted
that solvents such as CH2Cl2 or DMSO have very high
electronic extinction coefficients for the 14.4 KeV
Mössbauer radiation. It
is therefore advisable to use solvents containing elements with atomic
number Z < 10. (The extinction coefficient at 14.4 KeV
is 2.2 cm2/g for oxygen and 17.0 cm2/g for sulfur;
the 14.4 KeV γ-beam will be attenuated by
about I = I0 e-2.2 x 0.5 = 0.33 I0 in
passing through a 0.5 cm water sample.) Buffers containing 50 mM phosphate, chloride or sulfur groups do not pose any
problems; it is the high molar concentrations of the pure liquids that
cause trouble. In principle, a solvent such as CH2Cl2
could be used, provided the sample cup was filled only to ca. 1 mm height.
This, however, leads to a serious meniscus problem upon freezing, with the
result being that the central portion of the holder contains no sample
material.
Spectrometer Dependent Considerations
In our laboratory we typically
use cylindrical (12.3 mm OD, 10 mm height) sample holders made of delrin or nylon, for which the optimal filling volume
is 0.5 mL (6 mm high in the cup). If you fill in more, we loose counts by
electronic absorption; if you fill in less, we will loose resonance
absorption (less "effect"). For a very scarce protein sample, the
optimal cup above is exchanged for one which is tapered on the inside. This
cup requires only 0.3 mL of material and yields data as good as the
standard cup when analyzed in zero or small applied fields (i.e. the
"little" dewar). This cup is far from
optimal for high field studies.
We have also frequently designed
alternative cups for specialized experiments. For example, using lucite holders we can perform flash-photolysis studies
in the Mössbauer dewar.
For trouble makers like Miquel we have made more
drastic alterations to either simplify sample preparation or accommodate
subsequent spectroscopies. For example, we have recently used a holder of
12.3 mm diameter and 10 cm height. This unusual sample holder allowed us to
bubble oxygen (in a Schlenk line) through a 0.5
mL acetonitile solution at -60 °C to isolate an Fe(III)-peroxo complex without
spilling sample over the rim.
The main point is when
considering spectrometer dependent variables, it
is always advisable for the chemists to discuss preparation details with
the Mössbauer spectroscopist.
Mössbauer spectrometers can accommodate a variety of sample holders, and we
have always found a way to get around the chemist's problems.
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Sample Labels
As to avoid confusion, samples are to be
labeled with a 6-character code such as EMAS01, where "EM" are the initials of the P.I., "AS" is the
researcher who has made the sample. No two samples should have the same
codes. Send ALWAYS an email describing the samples.
If you are confused or are
missing information, please call us!
It could save everyone involved time and money.
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