Protecting the patient

Regulatory authorities demand the evaluation of foreign particles in all types of respiratory drugs. The technique described in this paper is well suited to detect and even chemically characterise particles that adhere to the capsule shell and which might be inhaled (size ca. <10µm) by the patient. The particles are sourced from the capsule shell itself as well as from the environment. The combination of image analysis and Raman spectroscopic opens access to the morphology, the size and shape, as well as the chemical structure of the identified particles: e.g. particles having their origin on the surface of the capsule shell can be discriminated from foreign particles sourced from the environment. Respiratory drugs are subject to high requirements with regard to purity. Amongst other analytical tasks necessary for proof of the quality of powder formulations administered in dry powder inhalers, the identification and characterisation of inhalable particles in particular which are foreign particles and particles not being part of the powder blend are troublesome. For the characterisation of these particles, several techniques such as SEM, IR microscopy or Raman microscopy are principally suited. By these methods, only a limited number of particles without a significant statistical base can be analysed. Automation of these spectroscopic methods would solve the problem getting material information of a larger number of particles. Automated SEM/EDX [1-3] systems are available for particle analysis, but provide only data on elemental composition of the investigated particle. This data is not sufficient for the characterisation of organic material. Limited by the wavelength of the used light with IR, only particles with a lower size limit of ca. 10µm can be characterised, so that the characterisation of the inhalable fraction size of particles in particular is not feasible. The combination of an image analysis system and a Raman microscope opening the access to particles with a minimum size of 2µm should also be very well suited for the characterisation of organic particles.The instrument setup and the investigation of foreign particles and particles from capsule shell used for administration in a Dry Powder Inhaler of the customer are discussed in this paper.

Reliable and qualified identification rap.ID Particle Systems manufactures analytical systems for the automatic chemical analysis of particles on the basis of Raman spectroscopy ã. The measuring device identifies all organic (except dark and/or fluorescent) and inorganic (except pure metals) particles of organic and inorganic nature larger than 2µm. The Liquid Particle Explorer was developed for the reliable identification of particles according to the high standards of measuring devices in a cGMP-controlled environment ä. In analogy to the method of membrane evaluation described in the United States Pharmacopoeia (USP), microscope images of the entire membrane surface covered with particles are automatically recorded and evaluated [6,7]. The position, length and width of the particles are exactly determined to the micrometer. After recognition, the particles are analysed with Raman spectroscopy. The resulting spectra are automatically attached to the pharmaceutical and customer-specific database. The database is created with Raman spectra of material samples. The system can therefore effectively recognise mixtures of materials, such as rubber stoppers or dyed polymers, due to their characteristic spectra. An automatically created report provides the size, identification and spectrum quality as well as the result for each individual particle in the form of hypertext.

Patented filtration device The product sample is opened and within a few minutes it is sucked through the gold coated and patented filter membrane. The handling is easy due to the aluminium ring setting. Depending on the viscosity of the solution and on the size of the particles to be examined, the size of the pore is selected. The contrast between particles and membrane is optimised for particle recognition through automatic image analysis. The membrane itself – as with all pure metals - produces no Raman signal. As a consequence, the particles on the membrane surface can be analysed without troubling metal signals using the Liquid Particle Explorer. All steps for the preparation are carried out under a laminar flow bench in a particle free environment. Prior to preparation of a sample and sample measurement, a blank test is performed. The system works in a highly automated fashion: only three parameters need to be set. 1) Size of the particle loaded area of the membrane. 2) Laser exposure time 3) Minimum particle size to be analysed

Foreign particle investigation The example described here was intended to provide information about ethanol insoluble foreign particles with a minimum size of 2 µm in empty gelatine capsules (for administration in dry powder inhalers). Per definition "Foreign Particles" differ from capsule components (capsule material). The capsule consists mainly of Gelatine, Titandioxide and Indigocarmine. The particles were investigated with regard to their number and chemical characteristics as well as their dimensions and appearance. The empty capsule is washed out with 2ml ethanol using a syringe equipped with a 0.2µm filter. The particle containing solution is filtrated (pore size 0.8µm) by means of a gold coated polycarbonate membrane. Thereafter the vessel is rinsed out twice with 3ml ethanol. The residue on the membrane is washed twice with 3ml water. The filter is then allowed to dry for 12 hours in a clean environment. The particle-loaded area is then automatically analysed with the Liquid Particle Explorer. Each particle bigger than 2µm is identified by the system. The following parameters are used: 1. exposure time per particle: 60s 2. automated picture analysis of 100 Fields (500 µm x 500 µm) 3. minimum particle length >2 µm

Results summary In the first step, high resolution photographs of the filter surface were generated and analysed by image recognition. In Fig. 1 one can clearly observe the particles on the surface of the membrane. Each of the identified particles gave a recognisable spectra which was identified by the APSys - Identifier database. An example of a Raman spectrum is shown in Fig. 2. The investigated particle could be clearly identified as polypropylene which could not be attributed to the ingredients of the gelatine capsule. To eliminate errors due to sample handling and sample preparation, a blank value was prepared and analysed in the LPE. In Table 1 the results of the blank measurement are presented: the investigation leads to the identification of 23 particles giving recognisable Raman spectra of different materials. The chemical structure of only three particles (UPOs) could not be elucidated by the system. In the investigation of ethanol-insoluble particles of the gelatine capsules, 78 particles were identified and the chemical structure of 76 could be elucidated (two UPOs). Six of the identified materials were also found in the blank value measurement. The main difference, the appearance of Protein and Titandioxide with a total number of 21 particles can be attributed to the composition of the gelatine capsule. Therefore, no major foreign particulate contamination can be found if the result of the blank value is taken into account. The identification of particles can be clearly distinguished between foreign particles and particles from the capsule shell, or as we found in another investigation, from the drug product. This kind of information is not accessible using conventional particle counters and microscopic measurements. The examinations with the Liquid Particle Explorer enable a statistically relevant and comparable conclusion about the chemical composition of the particles in a short time frame. The main contamination sources can be detected on the basis of the particle spectrum of a product sample. Another potential application field for the LPE in pharmaceutical development is the analysis of particulate drug delivery systems. Some of these drug delivery systems consist of a drug susbstance containing a core with the drug substance and a polymeric shell. The thickness of the shell is responsible for the time dependence of the drug release. For quality assurance purposes, the analysis of the thickness of the shell can be performed with an accuracy of less than a micrometer. At the same run, the content of the drug substance and excipients can also be determind. These parameters provide relevant information about the quality of the drug delivery system which is not accessible directly with another analytical method.