Differential scanning calorimetry (DSC) is a thermoanalytical technique in which the difference in the amount of heat required to increase the temperature of a sample and a reference is measured as a function of temperature. Both the sample and reference are maintained at nearly the same temperature throughout the experiment.
Generally, the temperature program for a DSC analysis is designed such that the sample holder temperature increases linearly as a function of time. The reference sample should have a well-defined heat capacity over the range of temperatures to be scanned.
Additionally, the reference sample must be stable, of high purity, and must not experience much change across the temperature scan. Typically, reference standards have been metals such as indium, tin, bismuth, and lead, but other standards such as polyethylene and fatty acids have been proposed to study polymers and organic compounds, respectively.
The technique was developed by E. S. Watson and M. J. O'Neill in 1962,. and introduced commercially at the 1963 Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy.
The first adiabatic differential scanning calorimeter that could be used in biochemistry was developed by P. L. Privalov and D. R. Monaselidze in 1964 at Institute of Physics in Tbilisi, Georgia. The term DSC was coined to describe this instrument, which measures energy directly and allows precise measurements of heat capacity.
Types There are two main types of DSC: Heat-flux DSC which measures the difference in heat flux between the sample and a reference (which gives it the alternative name Multi-Cell DSC) and Power differential DSC which measures the difference in power supplied to the sample and a reference.
Heat-flux DSC With Heat-flux DSC, the changes in heat flow are calculated by integrating the ΔTref- curve. For this kind of experiment, a sample and a reference crucible are placed on a sample holder with integrated temperature sensors for temperature measurement of the crucibles. This arrangement is located in a temperature-controlled oven. Unlike the traditional design, the special feature of heat-flux DSC is that it uses flat temperature sensors placed vertically around a flat heater. This setup makes it possible to have a small, light, and low-heat capacity structure while still working like a regular DSC oven.
Power differential DSC For this kind of setup, also known as Power compensating DSC, the sample and reference crucible are placed in thermally insulated furnaces and not next to each other in the same furnace as in heat-flux-DSC experiments. Then the temperature of both chambers is controlled so that the same temperature is always present on both sides. The electrical power that is required to obtain and maintain this state is then recorded rather than the temperature difference between the two crucibles.
Fast-scan DSC The 2000s have witnessed the rapid development of Fast-scan DSC (FSC), a novel calorimetric technique that employs micromachined sensors. The key advances of this technique are the ultrahigh scanning rate, which can be as high as 106 K/s, and the ultrahigh sensitivity, with a heat capacity resolution typically better than 1 nJ/K.
Nanocalorimetry has attracted much attention in materials science, where it is applied to perform quantitative analysis of rapid phase transitions, particularly on fast cooling. Another emerging area of application of FSC is physical chemistry, with a focus on the thermophysical properties of thermally labile compounds. Quantities like fusion temperature, fusion enthalpy, sublimation, and vaporization pressures, and enthalpies of such molecules became available.
Temperature Modulated DSC When performing Temperature Modulated DSC (TMDSC, MDSC), the underlying linear heating rate is superimposed by a sinusoidal temperature variation. The benefit of this procedure is the ability to separate overlapping DSC effects by calculating the reversing and the non-reversing signals. The reversing heat flow is related to the changes in specific heat capacity (→ glass transition) while the non-reversing heat flow corresponds to time-dependent phenomena such as curing, dehydration and relaxation.
Detection of phase transitions The basic principle underlying this technique is that when the sample undergoes a physical transformation such as phase transitions, more or less heat will need to flow to it than the reference to maintain both at the same temperature. Whether less or more heat must flow to the sample depends on whether the process is exothermic or endothermic.
For example, as a solid sample melts to a liquid, it will require more heat flowing to the sample to increase its temperature at the same rate as the reference. This is due to the absorption of heat by the sample as it undergoes the endothermic phase transition from solid to liquid. Likewise, as the sample undergoes exothermic processes (such as crystallization) less heat is required to raise the sample temperature. By observing the difference in heat flow between the sample and reference, differential scanning calorimeters are able to measure the amount of heat absorbed or released during such transitions. DSC may also be used to observe more subtle physical changes, such as glass transitions. It is widely used in industrial settings as a quality control instrument due to its applicability in evaluating sample purity and for studying polymer curing.
DTA An alternative technique, which shares much in common with DSC, is differential thermal analysis (DTA). In this technique it is the heat flow to the sample and reference that remains the same rather than the temperature. When the sample and reference are heated identically, phase changes and other thermal processes cause a difference in temperature between the sample and reference. Both DSC and DTA provide similar information. DSC measures the energy required to keep both the reference and the sample at the same temperature whereas DTA measures the difference in temperature between the sample and the reference when the same amount of energy has been introduced into both.
DSC curves
The result of a DSC experiment is a curve. The y-axis is the differential heat flux , measured in units of . The x-axis is either time, or the temperature. Both conventions for the x-axis are the same when the temperature is growing linearly with time, as it is usually the case.
For the sign of , there are two different conventions. One convention defines so that for exothermic process in the sample. The other defines the opposite. For example, in the convention where exothermic process corresponds to , the plot would show a negative spike as temperature increases beyond the melting point, since melting is an endothermic process. Conversely, it would show a positive spike as temperature drops below the melting point.
This curve can be used to calculate enthalpies of transitions. This is done by taking the time-integral of peak corresponding to a given transition:where is the enthalpy of transition, is the area under the curve, and is a certain constant called the "calorimetric constant". That particular constant varies from instrument to instrument, and can be measured by performing DSC on a calibration sample whose enthalpies of transition is known accurately.
This curve can also be used to calculate the molar specific heat capacity as a function of temperature. To obtain this, one perform DSC with two samples. One sample is a reference sample whose molar specific heat capacity is known accurately, and the other sample's needs to be measured. Then, after performing the DSC, we havewhere is the mole of the sample, is the mole of the reference, and is the rate at which temperature increases, with units of .
Applications Differential scanning calorimetry can be used to measure a number of characteristic properties of a sample. Using this technique it is possible to observe fusion and crystallization events as well as glass transition temperatures Tg. DSC can also be used to study oxidation, as well as other chemical reactions.
Glass transitions may occur as the temperature of an amorphous solid is increased. These transitions appear as a step in the baseline of the recorded DSC signal. This is due to the sample undergoing a change in heat capacity; no formal phase change occurs.
As the temperature increases, an amorphous solid will become less viscous. At some point the molecules may obtain enough freedom of motion to spontaneously arrange themselves into a crystalline form. This is known as the crystallization temperature (Tc). This transition from amorphous solid to crystalline solid is an exothermic process, and results in a peak in the DSC signal. As the temperature increases the sample eventually reaches its melting temperature (Tm). The melting process results in an endothermic peak in the DSC curve. The ability to determine transition temperatures and enthalpies makes DSC a valuable tool in producing phase diagrams for various chemical systems.
Differential scanning calorimetry can also be used to obtain valuable thermodynamics information about proteins. The thermodynamics analysis of proteins can reveal important information about the global structure of proteins, and protein/ligand interaction. For example, many mutations lower the stability of proteins, while ligand binding usually increases protein stability. Using DSC, this stability can be measured by obtaining Gibbs Free Energy values at any given temperature. This allows researchers to compare the free energy of unfolding between ligand-free protein and protein-ligand complex, or wild type and mutant proteins. DSC can also be used in studying protein/lipid interactions, nucleotides, drug-lipid interactions. In studying protein denaturation using DSC, the thermal melt should be at least to some degree reversible, as the thermodynamics calculations rely on chemical equilibrium.
Blood plasma and serum In addition to purified proteins, DSC has been applied to unfractionated blood plasma and serum; the diluted fluid is measured directly, without labeling. The resulting curve, called a thermogram, approximates the sum of the combined denaturation profiles of the most abundant plasma proteins, weighted by their concentrations. Because these proteins interact, the additivity is not exact. For plasma from healthy donors scanned at 1 °C per minute, three independant research groups reported a fibrinogen transition at about 51 °C, a dominant albumin peak near 62–63 °C, immunoglobulin and other globulin transitions between roughly 68 and 75 °C, and a weak transferrin transition at about 85 °C. Serum exhibits a similar profile, except that the fibrinogen transitions are absent.
Calorimetric measurements of serum and plasma from cancer patients were documented as early as the 1990s by a team in Tbilisi, who observed broadened heat-absorption curves shifted toward higher temperatures compared to healthy controls. From 2007, Chaires, Garbett and colleagues at the University of Louisville reported that plasma from healthy individuals gave a characteristic, reproducible thermogram. Samples from patients with cervical cancer, Lyme disease, rheumatoid arthritis or systemic lupus erythematosus displayed altered profiles. The albumin peak was reduced and the curve shifted to higher temperatures, changes that could not be explained solely by the concentrations of the major proteins; they have been attributed to ligand binding, post-translational modification and altered protein–protein interactions, but the underlying mechanism remains unknown. Independent groups in Poland, Hungary, Bulgaria, and Spain subsequently published thermogram studies. Applied to serum, the method has been termed "thermal liquid biopsy", and it has also been described as a calorimetric form of liquid biopsy.