Black Specks in Tablet Stability Samples

Visual appearance of pharmaceuticals is important in determining how closely patients comply with their dosage requirements. One problem that is often seen in  tablets is discoloration, or black specks, on the surface. 

These specks can form because of contaminants in raw materials or from the tablet manufacturing process, and various interactions that may occur during storage. The development of tablet discoloration during storage is often complicated to investigate, because it could involve a variety of interactions between active ingredients, excipients, and packaging materials. This technical case study describes how black specks in tablets were investigated during a stability study. 

Materials and methods
Materials. The pharmaceutical tablets used in this study were film-coated, controlled-release tablets manufactured at Patheon. All excipients used to manufacture the tablets were commercially available and met the United States Pharmacopeia (USP)/European Pharmacopoeia (EP)/Japanese Pharmacopoeia (JP) standards. Non-active ingredients included:

• Lactose monohydrate

• Hydroxypropylmethylcellulose

• Starch 1500 (pregelatinized starch)

• Sodium lauryl sulfate (SLS)

• Colloidal silicon dioxide

• Magnesium stearate.

• The content of API was low, at either 0.1 or 0.4 mg per tablet.

Methods. The microbial limit test was performed using USP <61> and <62>. Visual inspection was performed on the core, coated tablets, and excipient/active binary mixtures after storage at 40 °C/75% relative humidity (RH) and 60 °C/ambient humidity for 4-6 weeks. Backscattered electron microscopic imaging and elemental composition analysis for the tablets with discoloration were performed using a JEOL JSM 6400 Scanning Electron Microscope (SEM) coupled with Energy Dispersive X-ray Spectroscopy (iXRF EDS/XRF). Mass spectra of SLS were obtained by direct infusion of aqueous solutions of SLS samples into an AB Sciex 3200 LC/MS/MS system with electrospray ionization in both positive and negative modes. Other SLS tests were performed as per the USP/EP/JP monographs for SLS.

Results and discussion
Tablet samples were subjected to a stability study under real time (25 ˚C/60% RH) and accelerated (40˚C/75% RH) conditions. After three months, the samples held under accelerated conditions started to exhibit black specks as demonstrated in Figure 1. Because no observations of black specks were reported when these tablets were initially tested for release, after one month at 40 ˚ C/75% RH, or following storage at 25 ˚C/60% RH, it could be concluded that the specks did not result directly from contamination of tablet ingredients and were not introduced during the manufacturing processes. Instead, they were more likely due to some changes that occurred within the product during the three months when the product was exposed to accelerated storage conditions. An investigation was started to identify a root cause for these observations.

Microbial limit test. Microbial contamination was investigated as a potential cause of tablet discoloration, because the warm, moist conditions within a 40 °C/75% RH stability chamber presented a good environment for accelerating growth of microbes.

Microbial limit tests (MLT) were performed on those tablets whose surfaces showed significant discoloration. Results met all acceptance criteria for total aerobic microbial count, yeasts, molds, and indicator organisms, which ruled out microbial contamination as a root cause of the black specks.

Excipient interactions Because this phenomenon was clearly due to product changes over time, and not some initial contamination, some form of interaction between tablet ingredients was strongly suspected. To study this further, the authors decided to simplify the system by working with binary mixtures. Thus, excipient compatibility tests were performed by compressing binary mixtures (excipient/excipient, excipient/active) of each component of the tablet formulation into tablets and stressing the tablets under 60 °C and 40 °C for up to six weeks. Three different lots of each excipient were tested to evaluate batch-to-batch variability. The core and coated drug product tablets were also stressed to evaluate the compatibility of the coating material with the formulation core. Binary mixture stress results showed that discoloration developed mainly in those samples that contained material from one specific lot of SLS. Further, the extent of discoloration increased with increasing temperature.

In addition, discoloration shown on both core and coated tablets indicated that it was not caused by incompatibility between the formulation core and any coating material ingredients. 

Imaging and elemental analysis by SEM/EDS. Back-scattered electron imaging and elemental composition analysis on the areas of discoloration of the tablets were performed by SEM/EDS. Results are shown in Figure 2; the image is in negative mode so that the white area is actually the black speck. As shown in the figure, discoloured areas on the tablets are enriched with the elements sulfur (S) and sodium (Na). The only excipient in the formulation that contained S and Na was the SLS, which further supported a strong relationship between SLS and tablet discoloration.

Stress study on SLS. Four samples of two lots of SLS were used in this study. Lot A was the same lot used to manufacture tablets that showed discoloration after holding for three months under accelerated conditions. Lot B was a fresh sample from a different supplier. Both lots were held at two different conditions: ambient (as a control) and 60 °C for four weeks. The physical appearance results are summarized in Table I.

Results indicated that discoloration appeared in stressed samples of SLS from Lot A and did not show up in stressed samples from Lot B or the control samples of either lot. A new batch of tablets was manufactured using the SLS from Lot B and then stressed at 60° C/ambient RH for four weeks. No tablet discoloration was reported over this period. Because both lots of SLS were of a pharmaceutical grade that met the requirements of USP/EP/JP, some further investigation was then conducted to try to establish why the different lots performed differently in tablets.

Analysis by mass spectroscopy (MS). Mass spectra of SLS from Lot A and Lot B were obtained by direct infusion of aqueous solutions of each lot into an AB Sciex 3200 MS/MS system. Both positive and negative turbo ion-spray modes were used to collect the MS data. 

Figure 3 displays the mass spectra of negative mode. Peaks at m/z at 60.0 and 212.0 are from blank, and the peak at m/z 265 corresponds to SLS (C12H25SO4). Apart from these peaks, there were two major peaks (m/z 293 and m/z 97) observed only in the SLS sample from Lot A. These two peaks correspond to the C14 tetradecyl sulfate homologue (C14H29SO4) and residual sulfuric acid (HSO4), respectively.

The mass spectra of SLS collected from positive mode were crowded with peaks, which made it difficult to interpret. But, overall, the number of peaks from Lot A was much greater than the number of peaks from Lot B. 

The combined negative and positive mode results indicated that SLS from Lot B has a higher degree of purity than SLS from Lot A, but, more specifically, Lot A showed a significant m/z peak that corresponded to sulfuric acid, which was much less intense in the spectrum of material from Lot B. 

Compendial test results. Table II summarizes the quality control (QC) release test results of the two lots of SLS used in this study. Results showed that, although both lots satisfied the specification requirements of USP/EP/JP, Lot A had a higher level of unreacted alcohols and a lower assay value than Lot B. In addition, the alkalinity and pH value of Lot A are lower than those from Lot B, which again supports the view that this particular lot does contain an excess of sulfuric acid.

Discussion. The investigation data demonstrated that the discoloration of the tablets is closely related to the impurities of the SLS used in the tablet formulation. 

SLS is commercially manufactured by sulfation of lauryl alcohol, followed by neutralization with sodium carbonate. The three most common industrial synthetic processes are:

SO₃ /air process
Step 1 C₁₂H₂₅OH + SO₃ → C₁₂H₂₅SO₄H 
Step 2 C₁₂H₂₅SO₄H + Na₂CO₃ → C₁₂H₂₅SO₄Na + CO₂ + H₂O

Oleum (H₂ SO₄ /SO₃ ) process
Step 1 C₁₂H₂₅OH + H₂SO₄/SO₃ → C₁₂H₂₅SO₄H + H₂O
Step 2 C₁₂H₂₅SO₄H + Na₂CO₃ → C₁₂H₂₅SO₄Na + CO₂ + H₂O

Chlorosulfonic acid process 
Step 1 C₁₂H₂₅OH + ClSO₃H → C₁₂H₂₅SO₄H + HCl
Step 2 C₁₂H₂₅SO₄H + Na₂CO₃ → C₁₂H₂₅SO₄Na + CO₂ + H₂O

For the starting material, lauryl alcohol is usually made from natural products (e.g., coconut or palm kernel oil) followed by a series of hydrolysis and reduction reactions. Due to the natural origin, the lauryl alcohol used for the starting material often contains a minor quantity of C-10, C-14, and C-16 homologues, and they can be further transferred into the final product. Because of the similar chemical/physical property of those homologues, they are not considered to be impurities of SLS. 

From the synthetic routes, the most common synthetic impurities of SLS are the unreacted residual alcohols, residual sulfuric acid (resulting from incomplete neutralization and/or inefficient purification), salts (NaCl and/or Na₂SO₄), and the remaining Na₂CO₃ after the final neutralization reaction. This is probably why compendial monographs of SLS include specification requirements for alkalinity, sodium chloride, sodium sulfate, and unsulfated alcohols content. 

The impurity profile and levels in the SLS final product depend on the actual sulfation process used for SLS manufacturing and how well the process is controlled. For example, the oleum process is an equilibrium process, which usually leaves large quantities of sulfuric acid in the final reaction mixture, and the unreacted sulfuric acid must be separated from the reaction matrix before the final neutralization step takes place. Therefore, SLS manufactured from this process usually contains a relatively high level of sulfate. On the other hand, for the SO3/air or the chlorosulfonic acid process, the reaction is stoichiometric and fast. When reaction conditions are tightly controlled, there is much less sulfuric acid or sulfate formed, resulting in a product of higher quality.

It is possible that during the manufacturing of SLS (Lot A), the neutralization step had not been completed, resulting in the presence of residual sulfuric acid in the final product. This could explain the low alkalinity and pH value reported in Lot A from the QC release test results (Table II). Upon storage at higher temperature, the residual sulfuric acid undergoes dehydration reactions with the carbohydrate excipients (e.g., lactose) in the tablet formulation, resulting in the formation of black specks.

Conclusion
The most likely mechanism of tablet discolouration was related to the dehydration reaction between the residual sulfuric acid (due to incomplete neutralization during SLS manufacture) and the carbohydrate excipients present in the formulation, upon storage at higher temperatures.

Tablet discoloration is closely related to the quality of SLS used in tablet formulation, and the quality of SLS depends on the synthetic route and degree of process control used by the SLS during manufacture.
Current specifications of SLS in compendial monographs are not sufficient to control the quality of SLS, because they do not discriminate between different SLS lots, including those that may cause this type of drug product failures. Therefore, it is recommended to develop a testing requirement in the USP/EP/JP monograph that more tightly controls the formation of residual sulfuric acid and other impurities during SLS manufacturing.

All figures are supplied by author.

Peer-Reviewed Research
Submitted: January 5, 2015.
Accepted: January 26, 2015.

Article DetailsPharmaceutical Technology Outsourcing Resources Supplement
Vol. 39, No. 8
Pages: 40–44, 48

Citation: When referring to this article, please cite it as G. Carr, "Black Specks in Tablet Stability Samples," Pharmaceutical Technology 39 (8) 2015.