QUALITY CONTROL - Testing Methods | Overcome OTC Challenges

Jeff Layne, PhD, and Sky Countryman

Refine testing to meet FDA concerns about over-the-counter formulations In September, the FDA and U.S. Pharmacopeial Convention held a joint meeting to discuss limitations of the testing procedures for over-the-counter drug substances and products. Reviews of testing protocols for OTC products found that many are not adequate for testing for such things as impurities, which would indicate expired products. Developing suitable testing methods can be more challenging than it might initially seem. Many OTC cough and cold medicines, for example, contain more than one API, which, due to differences in efficacy, are dosed at different concentrations. Thus, developing a single test for all degradants in one analysis can be challenging. High performance liquid chromatography—more specifically, reversed-phase HPLC—is the technique of choice for quantitation of drug products and impurities. Other techniques, such as ion exchange, do not provide the efficiency necessary to quantitate impurities at the required 0.1% level. Fortunately, there are many different RP-HPLC stationary phases and particle types available today. When dealing with formulations containing multiple active components, it is often beneficial to utilize high-efficiency particles that will give improved resolution between critical components. Kinetex (Phenomenex) core-shell particles were engineered to provide increased efficiencies and improved performance compared to traditional fully porous particles. This improved performance typically leads to much faster separations and higher sensitivity. click for larger view FIGURE 1. Analysis of Cough and Cold Medicine (NyQuil) on Kinetex 2.6μμm C18 100 x 4.6mm ID. The high-efficiency nature of the Kinetex particle allows for the accurate quantitation of all drug substances in less than four minutes. Comparing results from complementary stationary phases such as the C18, XB-C18, and PFP can help identify the best separation. In the case of the well-known cough and cold formulation, Vicks NyQuil, the Kinetex C18 provided the best resolution and fastest analysis time for all three active components (Figure 1). Difficult Formulations Adding more complications are the various formulations for the same medicine: tablet, gel capsule, liquid syrup, and suspensions. In any of these dosage forms, the API or APIs usually only make up a small fraction of the total mass, with the bulk being made up of various excipients such as stabilizers, fillers, flavoring agents, and preservatives. It is the fillers and excipients that are often the source of many of the chromatographic challenges. Oral drug suspensions can be especially problematic for HPLC analysis due to the fact that, by definition, a suspension is a fluid that contains undissolved solids, which are large enough to undergo sedimentation. When injected onto an HPLC, the undissolved solids can physically clog the inlet frits of the column, the pores of the silica, and the interstitial space between the silica particles. Physical clogging of the media and/or frits most often manifests as an increase in system backpressure, which, if it gets high enough, can shut down an HPLC run and also render the HPLC column unusable. click for larger view FIGURE 2. A comparison of the Effectiveness of Centrifugation Versus a Two-Step Filtration for A Drug Suspension Sample. Filtering a drug suspension sample through a 0.45 µm followed by a 0.2 µm PTFE filter is much more effective at protecting the analytical HPLC column from clogging due to suspended particulates in the suspension matrix than is centrifugation alone. Depending upon the nature of these undissolved solids, they may be readily removed using centrifugation. Centrifugation of a diluted aliquot of a drug suspension, using a small benchtop centrifuge for five to 10 minutes, can remove suspended particulates. However, Figure 2 gives an example of a centrifuged OTC suspension where injection of the supernatant led to an increase in system backpressure with each sequential injection. Upon removal of the HPLC column and replacement with a new column, the system pressure returned to normal, indicating that the pressure buildup was within the column itself and not in the HPLC system plumbing. Back-flushing of the column using a highly aqueous solution and a 100% organic solvent (acetonitrile) for about one hour resulted in only a partial return to the initial pressure, indicating that, for all practical purposes, the clogging was irreversible. The use of syringe filters for removing particulates from samples prior to injection is well established, and there are many materials to choose from. In the case of the previous OTC suspensions, we have found that filtration using a Phenex 0.45 µm polytetrafluoroethylene filter followed by a second filtration using a Phenex 0.20 µm PTFE filter was a more effective way to remove the undissolved microparticulates. After this process, even relatively large sample volumes (50-100 µL if necessary) can be injected without significantly affecting system backpressure. Filtering with a Phenex 0.45 µm filter alone will not usually protect the column from clogging. Likewise, skipping the first crude filtration step and attempting to filter the initial drug suspension (or even a 10-fold dilution of drug suspension) directly through a Phenex 0.2 µm filter will typically result in the filter itself rapidly becoming clogged before a sufficient volume of the drug suspension has been clarified. click for larger view FIGURE 3. Effectiveness of Security Guard Ultra at Protecting an Analytical Column from Physical and/or Chemical Degradation. In this accelerated longevity test using sequential injections of an unfiltered matrix, the guard cartridge system is able to effectively absorb the majority of the contaminants that would otherwise foul the analytical column. Filters are designed to be inert, but they can interact with the drug, causing problems with recovery. If a filter material is to be used for an assay, a filter absorption study must be done to verify recovery of the API or its associated impurities. If recovery is too low, it may be necessary to explore alternative membrane types such as nylon. Please note that, while filtration and centrifugation are effective at removing undissolved solids from drug suspensions, neither is effective for removing the surfactants and emulsifiers that are often an integral part of the suspension formulation. Thus, while filtration will often protect that HPLC column from exhibiting increases in backpressure due to clogging, it is still quite common to see a gradual shift in chromatography (retention, selectivity, and so on) that can be attributed to the gradual buildup of surfactants from the sample matrix in the HPLC column. An additional sample preparation step is necessary to deal with these intractable components of the suspension matrix. Surfactants and Emulsifiers In order to maintain the relatively homogenous composition of the drug suspension and prevent, as much as possible, the components in the formulation from settling out, drug manufacturers must add a variety of surfactants and/or emulsifiers. Surfactants, which display a high affinity for the C18 phases that are typical in RP-HPLC methods, can be very difficult to wash off under normal mobile phase conditions and tend to build up over time, gradually changing the nature of the stationary phase surface and leading to a shift in chromatographic retention and selectivity. This gradual shift in chromatography as a result of surfactants from the sample matrix is problematic in that it may cause an HPLC column to fail system suitability testing if a monitored parameter, such as retention time or resolution, shifts outside of an acceptable limit. Ionic surfactants can be removed using ion-exchange SPE resins that selectively retain the surfactant, allowing the API and related substances to pass through. However, many non-ionic surfactants such as Tween (polysorbate) cannot be removed by this or any other mechanism. In such cases, the use of a guard column can be highly effective in preserving column performance. click for larger view FIGURE 4. A Guard Column Can Protect the Analytical HPLC Column from Shifts in Chromatography Due to Injection of Drug Suspension Samples that Contain Emulsifiers and/or Surfactants. Injection of a drug suspension sample containing the surfactant Tween results in an obvious and rapid shift in chromatographic retention for several of the peaks (top overlay; red trace = initial rejection, blue trace = second injection of drug suspension containing Tween-80). However, with the C18 guard cartridge system in place, the shift in retention is not apparent. Guard columns protect analytical columns in two ways: 1) by physically capturing suspended particulates that would otherwise clog the analytical column and 2) by trapping chemicals from either the mobile phase or the injected sample that may have a detrimental effect on HPLC column performance (Figure 3). Cartridge-based systems such as the SecurityGuard and SecurityGuard ULTRA are a cost-effective way to protect the analytical column. When changed out at appropriate time intervals, the same analytical column can be used for a longer time than would be possible without the guard column. When working with drug suspensions, guard columns are particularly important. We have found guard columns to be vital in preserving column performance. SecurityGuard C18 appears to absorb some of the emulsifier and/or surfactants present in a sample matrix, preventing them from accumulating in the analytical column and preserving chromatography (Figure 4). In the absence of a SecurityGuard C18 guard, a single injection of the placebo matrix triggers a substantial change in chromatography, which continues to degrade performance on subsequent injections. However, when the guard column is included, the injections of placebo have little if any effect on chromatography. We must attribute this protective effect to the guard column “capturing” some surfactant contained in the injected sample matrix. Of course, the ability of the guard to absorb these detrimental components will be determined by the relative amount of media present in the guard and the amount of contaminant loaded with each injection. To prevent saturation of the guard column, we recommend changing it at regular intervals, a practice facilitated by the use of disposable cartridge-type guard column systems. The FDA is heavily scrutinizing OTC formulations. The USP and other organizations are working hard to develop standard testing methods for companies to use. In the meantime, robust analytical methods for common OTC formulations are needed. We have found guard columns to be vital in preserving column performance. SecurityGuard C18 appears to absorb some of the emulsifier and/or surfactants present in a sample matrix. OTC drug formulations present an extreme challenge for the HPLC analyst due to the presence of multiple active components and the complex nature of the sample matrix, which make it difficult to separate active ingredients and their impurities from excipients. The use of high-efficiency Kinetex core-shell materials provides increased resolution that can help resolve critical impurities. When working with challenging formulations like suspensions, the presence of particulates and emulsifiers in the sample matrix can lead to poor column lifetime and reproducibility. When working with these types of formulations, analytical HPLC column performance and lifetime can be extended significantly with the use of Phenex syringe filters (0.45 µm and 0.2 µm PTFE), which physically capture the suspended particulates before they can clog up the analytical column. The use of the SecurityGuard and SecurityGuard ULTRA guard cartridge systems (C18 cartridge-type preferred) can also chemically absorb surfactants and emulsifiers that can alter HPLC column performance over time. Jeff Layne, PhD, is technical manager for PhenoLogix at Phenomenex, and Sky Countryman manages PhenoLogix and applied technology for the firm. Editor’s Choice Aicher B, Peil H, Peil B, Diener HC. Pain measurement: Visual Analogue Scale (VAS) and Verbal Rating Scale (VRS) in clinical trials with OTC analgesics in headache. Cephalalgia. 2012;32(3):185-197. Friedland S, Kothari S, Chen A, Park W, Banerjee S. Endoscopic mucosal resection with an over-the-counter hyaluronate preparation [published online ahead of print Feb. 29, 2012]. Gastrointest Endosc. U.S. Food and Drug Administration. Drug applications for over-the-counter drugs. FDA website. Available at: www.fda.gov/drugs/developmentapprovalprocess/howdrugsaredevelopedandapproved/approvalapplications/ over-the-counterdrugs/default.htm. Accessed March 13, 2012.