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Separation of Active Pharmaceutical Ingredients (APIs) from Excipients in Pharmaceutical Formulations Jerry L. Atwood Cryst. Growth Des., Just Accepted Manuscript • DOI: 10.1021/acs.cgd.5b00317 • Publication Date (Web): 16 Apr 2015 Downloaded from http://pubs.acs.org on April 21, 2015

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Crystal Growth & Design

Separation of Active Pharmaceutical Ingredients (APIs) from Excipients in Pharmaceutical Formulations

Jerry L. Atwood, Department of Chemistry, University of Missouri-Columbia, Columbia, MO 65211

Abstract The active pharmaceutical ingredient (API) may be physically separated from the excipients in a pharmaceutical formulation by making use of the difference in density of the API and that of the excipients. The API may then be fully characterized by standard techniques. In the density separation process, the API is not dissolved, nor is the crystal form of the API changed. As examples, Form I fexofenadine hydrochloride is separated from Allegra Allergy 24 Hour Tablets, and Form I lansoprazole is separated from Equate Lansoprazole Delayed Release Capsules.

Introduction The analysis of formulated drug products has been of increasing importance over the last several years.1,2 The potential for decomposition of the API has always placed emphasis on the analysis of the compound itself. This has routinely been accomplished by techniques such as HPLC.3 However, HPLC is powerless to speak to the crystal form of the API. For crystal form analysis, scientists have placed emphasis on techniques such as XRPD,4-6 infrared and Raman spectroscopy,7,8 and solid-state NMR.7,9 For the characterization of the API, these ‘fingerprint’ techniques are excellent individually, and even more powerful in combination.10 However, the

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API is often a minor component by weight in a tablet or other formulation. The presence of the excipients can interfere with the analysis of crystal forms.8 In the evaluation of crystal form, for best results one must separate the excipients from the API in a pharmaceutical formulation. Dissolution is not an option, since the crystal form is lost in solution. The API has frequently been formulated as a salt if the molecule itself is not readily water-soluble. Most excipients are also water-soluble. The crystallography of five decades past was not based on a high production of single crystal X-ray structures. Indeed, by the early 1970’s a rule of thumb was that a crystallographer with a modern (as of 1970) four-circle X-ray diffractometer and a moderate size research group should strive to do five or six structures per year. Times have changed. Today, moderate sized research groups commonly do more than ten X-ray crystal structures per week. Computer and diffractometer power have consumed much of the art of the crystallography of the 1960’s. In the 1960’s and 1970’s the measurement of the density of single crystals was, depending on the journal, mandatory for publication.11 The density measurement was performed by placing the crystal in a fluid medium in which the crystal was not soluble. The density of the fluid was then adjusted (by adding solute or other miscible fluids) so that the crystal under analysis would float half way up in the column of fluid. At that point, the density of the crystal was equal to the density of the fluid medium

Results and Discussion As noted above, the measurement of crystal density was standard crystallographic practice of the 1960’s and 1970’s and this technique forms the scientific basis for the separation of API from excipients in a pharmaceutical formulation. The essential feature of what is referred


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to herein as the density separation method is that the API be insoluble in the density fluid. A practical consideration is that the fluid be composed of two miscible liquids, one less dense and one, more dense, so that a wide range of densities is available. Mixtures of halocarbons and hydrocarbons work well, and in examples described herein the mixture of iodobenzene and hexane (or toluene or cyclohexane) has been found to work well. Most APIs have densities in the range 1.1 to 1.5 g/cm3, with even more APIs in the more narrow 1.2 to 1.4 g/cm3 range. Common crystalline excipients generally fall into two ranges, those with densities over 1.5 g/cm3 and those with densities less than 1.15 g/cm3. With densities of 1.823 g/cm3 for iodobenzene, 0.865 g/cm3 for toluene, 0.778 g/cm3 for cyclohexane, and 0.655 g/cm3 for hexane, most formulations are amenable to mixtures of iodobenzene and toluene, cyclohexane, or hexane. However, one must always test the target API(s) to make sure no crystal form conversion occurs using the density separation method. This is best ascertained by utilizing a standard sample of the target API under exactly the same conditions as used to density separate the formulation. If the excipients are tightly packed with the API, as is normally the case, one may need to use agitation to free the crystallites of the API from the excipients. A treatment of sonication of the formulation in the density medium works well. Enough sonication must be applied to free the excipient particles from the crystallites of the API, and this variable must be experimentally determined. Note that the API must be stable under exactly the same conditions as those to which the formulation is subjected, including the degree of sonication. Depending on the formulation, some excipient particles or crystals may adhere strongly to the crystals of the API. This has often proved to be case for lactose monohydrate, mannitol and sucrose. If excipients stick to the API, it may be necessary for the density separation method



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to be applied several times to achieve the desired level of separation. The only limitation is the amount of formulated API available. Each pass through the density separation process requires material. It is necessary to dry the density separated API from the iodobenzene/hexane liquid. Washing the API with hexane immediately after removing most of the iodobenzene/hexane by centrifugation and decantation, works well. Generally, the API should be washed twice with hexane, with a centrifugation separation after each washing. The residual hexane may then be removed by 10 minutes in a vacuum chamber at 50 torr and room temperature. As a final check of the density separation procedure, it is appropriate to compare the density separated API with the initial pharmaceutical formulation by XRPD or some other technique of choice. One should find the peaks due to the API are now relatively prominent compared to those of the excipients. The API, thus freed from excipients may then be tested in any desired manner. This procedure also works very well for the determination of the melting point or of the state of hydration of the API.

Experimental Section Separation of Fexofenadine HCl from Allegra Allergy 24 Hour Tablets The commercial product, obtained from WalMart, according to the label consists of tablets containing 180 mg of fexofenadine HCl with colloidal silicone dioxide, croscarmellose sodium, hypromellose, iron oxide blends, magnesium stearate, microcrystalline cellulose, polyethylene glycol, povidone, pregelatinized starch, and titanium dioxide. The total weight of the individual tablets is about 625 mg. Figure 1a shows the XRPD pattern of the crushed and


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lightly ground tablets, while Figure 1b shows the DSC of the crushed and lightly ground tablets. The XRPD pattern in Figure 1a shows peaks attributable to Form I of fexofenadine hydrochloride,12 as well as a broad hump due to the excipients, with the broad microcrystalline cellulose peak in the 21-24o 2θ range. The DSC shows one peak at about 193oC, which corresponds to the melting point range of 193-199oC for Form I fexofenadine hydrochloride.12 Fexafenadine hydrochloride has a density of about 1.25 g/cm3. The crushed and lightly ground tablets were treated successively with an iodobenzene/hexane solution of density ca. 1.36 g/cm3, 1.32 g/cm3, 1.32 g/cm3, and 1.20 g/cm3. The solution of density 1.36 g/cm3 was prepared by mixing iodobenzene and hexane in the volume ratio of 1.5:1. Five mL of the solution was added to ca. 300 mg of the crushed tablets in 13/100 mm culture tube. The mixture was sonicated for 5 min with the use of Crest Tru-Sweep 275 HT sonicator. The culture tube with contents was then centrifuged. The floating white material (that of density less than 1.36 g/cm3) was separated, placed in a culture tube, washed with an equal volume of hexane, and centrifuged. The white solid was separated from the liquid, treated with hexane, separated, and dried for 10 min in a vacuum chamber at room temperature and 50 Torr. The progress of the sequential density separations was monitored by XRPD of the dried powder. The more dense material from the sonication was discarded. The same procedure was followed for successive treatments of the previously density separated material with two solutions of density 1.32 g/cm3. In the final density separation with the solution of density 1.20 g/cm3, after the sonication and centrifugation, the more dense material was retained and the liquid above this material was discarded. The white crystalline material, which resulted from the density separation, produced the XRPD pattern and the DSC thermogram shown in Figure 2a and 2b, respectively. It may be seen that most of the excipients have been removed. Indeed, the XRPD pattern matches that of Form



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I of fexofenadine hydrochloride from the literature, Figure 3.12 The DSC thermogram now shows a single peak for Form I fexofenadine hydrochloride at 194oC. As a check on the stability of Form I fexofenadine hydrochloride, a sample of authentic Form I was treated successively as above with iodobenzene/hexane solutions of density ca. 1.36 g/cm3, 1.32 g/cm3, 1.32 g/cm3, and 1.20 g/cm3. No crystal form change was noted (Figure S1A and B.


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Figure 1a. XRPD pattern of Allegra crushed tablets.

Figure 1b. DSC thermogram of Allegra crushed tablets.



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Figure 2a. XRPD pattern of density separated fexofenadine hydrochloride.

Figure 2b. DSC thermogram of density separated fexofenadine hydrochloride.


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Figure 3. Overlay of the XRPD pattern of density separated fexofenadine hydrochloride with the literature XRPD pattern of Form I fexofenadine hydrochloride.



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Separation of Lansoprazole from Equate Lansoprazole Delayed Release Capsules, 15 mg The commercial product, obtained from WalMart, according to the label consists of capsules containing 15 mg of lansoprazole with hypromellose, low substituted hydroxypropyl cellulose, mannitol, meglumine, methacrylic acid copolymer, polyethylene glycol, polysorbate 80, sodium lauryl sulfate, sugar spheres, talc, and titanium dioxide. The total weight of the contents of the individual capsules (in the form of granules) was about 170 mg. Figure 4a shows the XRPD pattern of the crushed, ground, and powdered granules while Figure 4b shows the DSC of the crushed, ground, and powdered granules. The XRPD pattern in Figure 4a shows a few peaks which may be attributable to Form I of lansoprazole,6 as well as peaks attributable to other crystalline compounds. Additionally, there is also an amorphous hump in the 12-26o 2θ range. The DSC shows at least four peaks, in addition to the broad feature centered at about 70oC. Lansoprazole has a density of about 1.5 g/cm3. The crushed, ground, and powdered granules were treated successively with an iodobenzene/hexane solution of density ca. 1.52 g/cm3, 1.46 g/cm3, 1.51 g/cm3, 1.47 g/cm3, 1.50 g/cm3, and 1.50 g/cm3. The solution of density 1.52 g/cm3 was prepared by mixing iodobenzene and hexane in the volume ratio of 2.8:1. Five mL of the solution was added to ca. 300 mg of the crushed, ground, and powdered granules in 13/100 mm culture tube. The mixture was sonicated for 5 min with the use of Crest Tru-Sweep 275 HT sonicator. The culture tube with contents was then centrifuged. The floating white (the material of density less than 1.52 g/cm3) was separated, placed in a culture tube, washed with an equal volume of hexane, and centrifuged. The white solid was separated from the liquid, treated with hexane, separated, and dried for 10 min in a vacuum chamber at room temperature and 50 Torr. The progress of the sequential density separations was monitored by XRPD of the dried


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powder. The more dense material from the sonication was discarded. The same procedure was followed for the treatment of the previously density separated material with solution of density 1.46 g/cm3. Here, the more dense material was retained and the liquid and floating solids above this material were discarded. The successive density separation treatments were carried out as described above. The white crystalline material, which resulted from the sixth density separation, produced the XRPD pattern and the DSC thermogram shown in Figure 5a and 5b, respectively. It may be seen that most of the excipients have been removed. Indeed, the XRPD pattern matches that of Form I of lansoprazole from the literature, Figure 6.6 The DSC thermogram now shows a single peak for Form I lansoprazole at 167oC. As a check on the stability of Form I lansoprazole, a sample of authentic Form I was treated successively as above with iodobenzene/hexane solutions of density ca. 1.52 g/cm3, 1.46 g/cm3, 1.51 g/cm3, 1.47 g/cm3, 1.50 g/cm3, and 1.50 g/cm3. No crystal form change was noted (Figure S2A and B.



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Figure 4a. XRPD pattern of the crushed, ground, and powdered granules of Equate Lansoprazole.

Figure 4b. DSC thermogram of the crushed, ground, and powdered granules of Equate Lansoprazole.


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Figure 5a. XRPD pattern of density separated lansoprazole.

Figure 5b. DSC thermogram of density separated lansoprazole.



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Figure 6. Overlay of the XRPD pattern of density separated lansoprazole with the calculated literature XRPD pattern of Form I lansoprazole.

Stability of Crystal Morphology under Density Separation Conditions Samples of fexofenadine hydrochloride Form I API and lansoprazole Form I API were observed using a 40x microscope before and after the test density separations as noted above. No crystal morphology was observed. However, the APIs were fine powders and the morphologies were difficult to observe. As a further test, sitagliptin phosphate monohydrate API crystals of elongated plate morphology were treated successively as above with iodobenzene/hexane solutions of density ca. 1.36 g/cm3, 1.32 g/cm3, 1.32 g/cm3, and 1.20 g/cm3. No change in particle size or crystal morphology was noted.


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For Table of Contents Use Only Separation of Active Pharmaceutical Ingredients (APIs) from Excipients in Pharmaceutical Formulations Jerry L. Atwood The active pharmaceutical ingredient (API) may be physically separated from the excipients in a pharmaceutical formulation by making use of the difference in density of the API and that of the excipients. In the density separation process, the API is not dissolved, nor is the crystal form of the API changed. Density Separation

API

Tablet Excipients


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Separation of Active Pharmaceutical Ingredients ... - ACS Publications

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