Anal. Chem. 1988, 8 0 , 747-750
747
Absolute Purity Determination of Thermally Unstable Compounds by Differential Scanning Calorimetry Arnold Ramsland
Pharmaceutical Research Products Section, Quality Control Department, Hoffmann-La Roche, Inc., Nutley, New Jersey 07110
A eutectic purity method Is described which is applicable to many compounds that decompose at the melt. The method Is based upon the coiilgathre properties where minor components lower and broaden the melt of a eutectic comprlsed of the major component and an added second component. The method is applicable to many thermally unstable compounds that cannot be analyzed by the standard differential scannlng calorimetry purity method for two reasons. First, the eutectic melt occurs below the meit of both the major component and the added second component; hence, the eutectic melt can be designed to occur below the onset of decomposltlon by chooslng a low-meltlng second component. Second, only a fraction of the thermally unstable compound wl# actually meH at the eutectlc. The method has an additional advantage in that the sensitivity and range are adjustable by changing the level of added w c h d component. The method Is potentially much more sensitive to impurities than traditional thermal methods. Flnaiiy, the method Is experimentally slmple: only a dmerentlal scanning cakrlmaer and -hart recOrdec are needed. I n the present report, acetaminophen and biotin have been used to demonstrate the utility of the technique.
Thermal analysis has proven to be an accurate method of absolute purity determination for many compounds. The method is based upon the colligative properties where minor components lower and broaden the melt of the major component. An assumption usually made in absolute purity determination is that the minor components are totally soluble in the liquid phase or melt of the major component and totally insoluble in the solid phase of that component. In such a c a s , the van't Hoff equation is applicable (1,2)
where Toand AHf are the experimentally determined values of the melting point and heat of fusion of the pure major component. A plot of sample temperature, T,, versus the reciprocal of fraction melted, 1/F,should yield a straight line with the slope proportional to the more fraction impurity, X,. In addition, other equations have been developed for purity determination which attempt to account for solid solution formation (3, 4 ) . Two experimental methods of absolute purity determination using thermal analysis have traditionally been used: the dynamic and stepwise melting methods. The dynamic method is adequately described in various references (5-7) and is now an official ASTM purity method (E928-83). In the dynamic method, the sample is heated through the melt at a slow rate in order to approximate equilibrium conditions. In the stepwise method (4, €9, the sample is heated through the melt in steps so that equilibrium is achieved at each temperature step. In either method, the sample exists in a partially melted state for a considerable period of time (i.e., 15-30 min). For 0003-2700/88/0360-0747$01.50/0
this reason, absolute purity determination using thermal analysis has been limited to compounds that are stable at the melt. The objective of this work has been to develop a new method of thermal purity analysis which is not limited by this constraint.
THEORY A minor component will lower and broaden the melt of not only the major component but also a two-component eutectic mixture. The new technique, eutectic purity analysis, is based upon the effect of minor components (impurities) upon a eutectic which is purposely formed by adding a pure second component to the major component. An assumption made with this technique is that the minor components are totally soluble in the melt and totally insoluble in the solid phase of the eutectic mixture. Ideally, when a mixture of two pure components is heated, the minor component plus some of the major component will melt sharply at a eutectic. The temperature and composition of this eutectic melt are invariant. The remainder of the major component will continue to melt upon further heating. This thermal behavior is shown in the upper thermal curve in Figure 1. A 9515 molar mixture of acetaminophenlpaminobenzoic acid (PABA) shows a sharp eutectic melt at ca. 140 "C forowed by a broadened melt of the major component, acetaminophen, at 145-170 O C . The lower thermal curves demonstrate the effect of adding a third component, phenacetin, to the two-component mixture. The eutectic melt is significantly lowered and broadened upon increasing the phenacetin concentration from 0 to 1% . Eutectic purity analysis is applicable to many thermally unstable compounds which previously have been excluded from thermal purity methods. First, the eutectic melt occurs below the melt of both the major component and the added second component so that the eutectic melt can be designed to occur below the onset of decomposition by choosing an appropriate low-melting second component. Second, only a small fraction of the thermally unstable compound will actually melt at the eutectic so that decomposition is further minimized. Another advantage of the method is that one can change the extent of lowering and broadening of the eutectic caused by impurities simply by changing the mole fraction of the second component. This flexibility can be used to enhance the sensitivity. For example, a 9911 mixture of acetaminophen/PABA will undergo significantly more lowering and broadening with 1% phenacetin than will a 9515 mixture. Two- and Three-Component Phase Diagrams. The thermal behavior of two- and three-component systems can be more fully described in phase diagrams. Figure 2 shows a theoretical, two-component phase diagram of acetaminophen/PABA which has been constructed by computer using the following previously derived equation (4):
0 1988 American Chemical Society
748
ANALYTICAL CHEMISTRY, VOL. 60, NO. 8, APRIL 15, 1988
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TEMPERATURE tC>
Flgure 1. Effect of Increasing phenacetin upon a 9515 acetaminophen1PABA eutectic.
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for PABAIacetaminophen, PABAIphenacetin,and phenacetinlacetaminophen are designated at polnts 1, 2, and 3, respectively. The calculated three-component eutectic is designated at point 4.
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Flguro 3. Theoretical phase dlagram of acetaminophen, PABA, and phenacetin. The calculated eutectic compositions and temperatures
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Flgure 2. Theoretlcal phase diagram of PABA and acetaminophen. The calculated eutectlc temperature and mde fraction are 412.4 K and 0.555 mol % acetaminophen, respectkely.
Experimentally determined values of the melting point, To, and heat of fusion, A",,for each component have been used to calculate T, as a function of the mole fraction of the major component, X:. In each case, K , the partition coefficient of the minor component between the solid and liquid phase of the major component is set equal to zero so that solid solutions are not considered. The fraction melted, F, has been set equal to one so that the curved, solid lines represent the final melting temperature of the solid phase. The intersection of these lines determines the temperature and concentration of the eutectic. The eutectic temperature represents the initial melting temperature of the solid phase. The eutectic composition represenb the composition where the initial and final melting
temperatures of the mixture are the same. In other words, a mixture of two pure components of eutectic composition will melt sharply at the eutectic temperature. This is observed experimentally where a mixture a t the eutectic composition exhibits a single, sharp melt which appears very similar to the melt of a single pure component. In order to construct theoretical, three-component phase diagrams, the following assumptions are made: 1. The values of To and AHf for each pure compound determine the two-component eutectic temperature and composition. 2. The eutectic also exhibits colligative properties: the lowering and broadening of the eutectic is dependent upon the number rather than the kind of minor component molecules present. 3. Equation 2 is applicable to a pure two-component mixture. Here, Tois the eutectic temperature and A",is a weighted sum of the heats of fusion of each pure compound times the eutectic mole fraction. Figure 3 shows a theoretical, three-component phase diagram of acetaminophen/PABA/phenacetin. The three twocomponent phase diagrams are plotted as solid lines on the faces of the triangle. The lowering of each eutectic by the third component is shown as a broken line in the interior of the triangle. Examination of this phase diagram reveals an internal consistency in that the broken lines intersect at virtually the same temperature, concentration point-the three-component eutectic. This is consistent with the Gibbs phase rule which predicts zero degrees of freedom for four phases and three components a t constant pressure. EXPERIMENTAL SECTION Instrumentation. A Perkin-Elmer DSC-2 differential scanning calorimeter connected to a strip-chart recorder has been used for all experimental work. The thermal curves shown have been constructed with a Hewlett-Packard 9830A computer which receives and stores data from an analog to digital (A/D) converter
(Princeton Applied Research Model 131) and directs a Hewlet-
ANALYTICAL CHEMISTRY, VOL. 60, NO. 8, APRIL 15, 1988
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INCREASE I N SAMPLE S I Z E
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Flgure 4. Construction of extrapolated onsets for a pure and impure eutectic shown in curves A and 6, respectively.
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Packard 9872 plotter to construct labeled thermal curves. Experimental Conditions. All samples are tightly packed into volatile sample pans, evacuated, and then sealed under nitrogen atmosphere. The samples are heated at 2.5 "C/min, starting approximately 60 "C below the expected melt. Sample Preparation. Homogeneous sample preparation is absolutely essential in using the eutectic purity method. Two mixing techniques have been found to give reproducible results. In the first technique, weighed amounts (1-2 g) of the desired components are freeze-ground at liquid nitrogen temperature with a Spex freezer/mill (Model 6702). This technique was used for all doped acetaminophen samples. A second mixing technique is used when sample is limited or when vigorous freeze-grinding causes sample decomposition (as with biotin). In this technique, the second compound and impurity are first mixed at levels necessary to achieve a calibration curve. A few milligrams of this mixture is then weighed into a stainless steel vial (1.2 mm i.d.). Approximately 50 mg of pure major component is then weighed into the vial. A programmable calculator is used to determine the mass necessary to obtain a desired eutectic concentration. The contents are mixed by alternate clockwise and counterclockwisespinning of a spatula which has been cut to contour the inside of the vial. This sample preparation method is designed to minimize powder buildup on inside walls as well as buildup of static charge. Duplicates from the same sample preparation should always be run in order to ensure homogeneity. Temperature Lowering Determination. Traditionally, endothermic transitions are characterized by the extrapolated onset and peak temperatures. In this work, the extrapolated onset has been found to be more useful than the peak temperature in quantitating impurity because of the following: 1. The extrapolated onset reflects not only the lowering but also the broadening of the eutectic peak. 2. The extrapolated onset is not influenced by variations in sample size for a eutectic mixture of two pure Compounds as shown in Figure 4A. This is because the eutectic is an isothermal transition which results in a straight leading edge. If the major component is impure, however, the eutectic is no longer an isothermal transition and the leading edge is curved as shown in Figure 4b. In order to use the extrapolated onset in this case, two experimental procedures have been followed so that an accurate reproducible determination of temperature lowering is achieved. First, the sample size has been maintained relatively constant (5 mg) in order to minimize a possible effect of sample size. Second, in order to avoid ambiguity in determining the extrapolated onset for an impure eutectic, the onset is determined by the intersection of a line drawn through the base line prior to the endothermic peak and a line drawn through the point of inflection. RESULTS AND DISCUSSION Biotin has been chosen to demonstrate the applicability of this technique to thermally unstable compounds. The melt of biotin at 232 OC is accompanied by decomposition which precludes standard thermal purity methods. Biotin forms a eutectic with acetaminophen which experimentally has been found to melt at 159 "C. In order to ensure the accuracy of the experimentally determined eutectic melting temperature, the concentrations of the major component and second com-
454 449 444 439 434 42gL 0. 0
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Figure 5. Theoretical phase diagram of biotin and acetaminophen.
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