WO2005099404A2

WO2005099404A2 - Method for assaying chemicals deposited by a chemical delivery system

Info

Publication number: WO2005099404A2
Application number: PCT/US2005/012049
Authority: WO
Inventors: Charles Bryden
Original assignee: Aradigm Corporation
Priority date: 2004-04-08
Filing date: 2005-04-08
Publication date: 2005-10-27
Also published as: WO2005099404A3

Abstract

An assay method is disclosed which comprises depositing a compound on a surface of a material and obtaining an NIR spectrum of compound deposited on the surface. The compound may be dissolved in a solvent resulting in a solution which is aerosolized and moved through a filter which is permeable to the gas of the aerosol but substantially impermeable to the particles of the aerosol. An NIR spectrum is obtained of the solution deposited on the filter and the resulting spectrum is used to generate a calibration making it possible to use NIR spectra of samples to routinely assay chemicals deposited on a surface such as a filter in any manner such as an aerosol generating device.

Description

METHOD FOR ASSAYING CHEMICALS DEPOSITED BY A CHEMICAL DELIVERY SYSTEM

FIELD OF THE INVENTION [0001] The invention relates generally to spectroscopic methods for assaying chemicals, and specifically to Near-Infrared Spectroscopy (NIRS) methods for determining the amount of chemicals deposited in a container or on a surface.

BACKGROUND OF THE INVENTION

[0002] Current assay methods for chemicals delivered by a drug or other chemical delivery system typically require a significant amount of labor to obtain desired results. As a first step delivered chemical components may be collected in a container or collected on a surface such as a glass plate surface. Often one or more steps are required to process the sample before an assay is carried out, e.g. washing the chemical components from the collection surface into a container, extraction, evaporation, dilution to a specific volume and then transferring the resultant solution to a container suitable for assay. These steps are frequently time-consuming and labor intensive.

[0003] Further, the assay carried out after these sample processing steps can also be time- consuming and frequently requires highly skilled personnel to operate, maintain and trouble- shoot instruments and interpret results. Some of the assay techniques that fall in this category are HPLC, GC, Mass Spectrometry and UN Spectrometry.

[0004] An advantage of ΝIRS is that it can assay chemicals with few or no sample processing steps. A further advantage of ΝIRS is that the instrumentation is robust, requiring only infrequent calibration, maintenance or trouble-shooting. Thus, a highly skilled operator is not required to operate, maintain, and repair a Νear-Infrared (ΝIR) spectrometer.

[0005] The present invention uses ΝIRS to assay chemicals delivered by a chemical delivery device and collected in a container or deposited on a surface.

SUMMARY OF THE INVENTION [0006] In this invention, drugs, solvents, excipients or other chemical components of a solid or solution delivered by a delivery system are transferred to a collection material and analyzed using an NIR spectrometer. The chemical(s) delivered include but are not limited to the categories of inorganic compounds, small organic molecules, or large organic molecules such as proteins, DNA, RNA and synthetic polymers. Any collection material can be used, including but not limited to containers such as glass beakers or collection surfaces such as metal disks, glass fiber filters, or plastic filters. The container or surface can contain or be coated with a liquid or other material that aids in the retention of the delivered chemical(s) (e. g. a solvent or polymeric coating). The NIR data for one or more of the chemical component(s) of the delivered solid or solution are correlated to data obtained from a reference method to generate a calibration model using measurement of the area or height of NIR absorption bands, chemometrics techniques or other calibration techniques.

[0007] For routine analysis, the NIR spectrum of a sample is obtained and the content of the chemical component(s) in the sample is computed according to the calibration model. Because there is no wet chemistry processing, the NIR assay method is a highly specific method that is much faster and less costly than other specific methods such as HPLC, GC, UN Spectrometry and Mass Spectrometry techniques.

[0008] In accordance with a method of the invention any type of compound can be applied to a surface and thereafter an ΝIR spectrum taken of the compound on the surface. In accordance with the invention the surface may be a surface such as a glass fiber, glass container or glass sheet such as a slide which is transparent to the wavelengths generated by the ΝIR spectrameter. The compound can also be placed on material such as Teflon® or other perfluronated polymer surface that does not absorb in the near infrared wavelength region. Such a polymer reflects all light back into the integrating sphere rather than transmitting all light in a manner such as a transparent surface as glass does. Glass is a suitable surface when using a transmittance accessory. However, a reflecting surface is necessary when using a reflectance accessory. When using a reflectance accessory, a Teflon® cylinder is placed ontop of the glass fiber filter to reflect all of the near infrared light back through the filter and into the integrating sphere. Those skilled in the art will understand that a variety of surfaces which reflect all light such as providing a mirror surface can be used as a surface for depositing the compound. Further, in accordance with the invention after the first scan is taken with the surface in the first position, the surface may be moved such as being rotated 45°, 90°, 270°, or various amounts therebetween and another scan with the ΝIR spectrameter taken. The results of the plurality of scans can be combined to obtain additional inforamtion with respect to the compound deposited on the surface.

[0009] In accordance with the invention the compound may be deposited on the surface by any number of means. However, it is desireable to consider depositing the compound on the surface by the use of devices generally used in the field of medicine for the delivery of drugs. Accordingly, the compound can be deposited on the surface by creating an aerosol with a pulmonary drug delivery device or deposited on the surface by the use of a needleless drug delivery device which injects a solution of a pharmaceutically active drug into a patient without the use of a needle. [0010] An aspect of the invention is a method aerosolizing a solution comprising a solvent and a solute so as to create aerosolized particles in a gas, directing the particles in the aerosol onto a surface, obtaining an NIR spectrum of the solution deposited on the surface (in particular the solute within that solution) and calibrating the NIR spectra. [0011] In accordance with another aspect of the invention a method is provided wherein an NIR spectrum is obtained for a compound deposited on one or a plurality of surfaces by a drug delivery system, calibrating the NIRS spectra using a reference and employing the calibrated NIRS method to routinely assay chemicals deposited in a container or on a surface by a delivery device. [0012] In accordance with another aspect of the invention NIR spectra are obtained a plurality of times after rotating by X° the surface each time before taking the NIR spectra wherein X is in a range of from 1° to 180°. [0013] These and other objects, advantages, and features of the invention will become apparent to those persons skilled in the art upon reading the details of the formulations, methods and systems as more fully described below.

BRIEF DESCRIPTION OF THE DRAWINGS

[0014] The invention is best understood from the following detailed description when read in conjunction with the accompanying drawings. It is emphasized that, according to common practice, the various features of the drawings are not to-scale. On the contrary, the dimensions of the various features are arbitrarily expanded or reduced for clarity. Included in the drawings are the following figures:

[0015] Figure 1 is a schematic representation of a filter collection apparatus positioned downstream of a mouthpiece with the components labeled, which components may be used in connection with the method of the invention.

[0016] Figure 2 is a schematic representation of an Antaris™ ThermoNicolet integrating sphere used for acquiring diffuse reflection NIR spectra. 0017] Figure 3 is an NIR spectrum graph taken of a protein formulation using a method of the invention wherein a band at about 7100 cm^"1 is due to water in the filter and the band at about 5200 cm^"1 is due to water in the filter combined with the protein. The remaining bands are due to the protein. 0018] Figure 4 is a graph of an NIR spectrum of a protein formulation demonstrating the effect of interference fringes wherein the frequency of the interference fringes, the gap in the glass fiber filter was calculated to be 58 micrometers. 0019] Figure 5 is a graph of an NIR spectrum of a cromolyn formulation showing large bands at about 6900 cm^"1 and 5200 cm^"1 which are due to water in the aerosol and the filter whereas a band at about 6050 cm^"1 is due to the cromolyn. 0020] Figure 6 is a graph showing the results of the calibration experiment described in Example 1 showing the relationship between the calculated NIR value and the actual HPLC emitted dose using a spinning cup accessory with PLS calibration. [0021] Figure 7 is a table showing a comparison of NIR and microtiter plate assays of cromolyn filters.

DETAILED DESCRIPTION OF THE INVENTION

[0022] Before the present invention is described in detail, it is to be understood that this invention is not limited to particular variations set forth and may, of course, vary. Various changes may be made to the invention described and equivalents may be substituted without departing from the true spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation, material, composition of matter, process, process act(s) or step(s), to the objective(s), spirit or scope of the present invention. All such modifications are intended to be within the scope of the claims made herein.

[0023] Methods recited herein may be carried out in any order of the recited events which is logically possible, as well as the recited order of events. Furthermore, where a range of values is provided, it is understood that every intervening value, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the invention. Also, it is contemplated that any optional feature of the inventive variations described may be set forth and claimed independently, or in combination with any one or more of the features described herein.

[0024] All existing subject matter mentioned herein (e.g. , publications, patents, patent applications and hardware) is incorporated by reference herein in its entirety except insofar as the subject matter may conflict with that of the present invention (in which case what is present herein shall prevail). The referenced items are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such material by virtue of prior invention. 0025] Reference to a singular item, includes the possibility that there are plural of the same items present. More specifically, as used herein and in the appended claims, the singular forms "a," "and," "said" and "the" include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as "solely," "only" and the like in connection with the recitation of claim elements, or use of a "negative" limitation. Unless defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

DEFINITIONS

0026] "Aerosol" means a suspension of particles in a gaseous medium, e.g., aqueous particles suspended in air. An "aqueous aerosol" is an aerosol formed from an aqueous solution (i.e., a solution containing water as a solvent).

0027] "Dosage form" or "DF" is a container closure system that is used to hold a dose (or partial dose) of a formulation prior to aerosolizing it.

0028] "Emitted dose" or "ED" is the amount of aerosolized particles of the active ingredient that is emitted from an aerosol drug delivery device.

[0029] "Gas Chromatography" or "GC" is a means of separating chemical components in a mixture, e. g. a drug formulation, and quantifying some or all of the chemical components by partitioning the chemicals between a stationary phase and a gaseous mobile phase.

[0030] "High Performance Liquid Chromatography" or "HPLC" is a means of separating chemical components in a mixture, e. g. a drug formulation, and quantifying some or all of the chemical components by partitioning the chemicals between a stationary phase and a liquid mobile phase.

[0031] "Mass Spectrometry" is a means of separating chemical components in a mixture, e. g. a drug formulation, and quantifying some or all of the chemical components by measuring the mass or mass-to-charge ratio of the components or fragments of the components. [0032] "Partial Least Squares" or "PLS" is a chemometrics technique used to derive the concentration of an analyte in a mixture using spectral data. During calibration, spectra are decomposed into factors. The factors are used to compute analyte concentration from the spectra of samples. PLS output parameters during calibration include the Root Mean Squared Error of Calibration (RMSEC) and the Root Mean Squared Error of Prediction (RMSEP).

[0033] "Specificity" is the ability of an analytical technique such as NIR or HPLC to assess unequivocally the analyte in the presence of components which may be expected to be present.

[0034] "Wavenumber" or "cm^"1" is the reciprocal of the wavelength in centimeters of the light (e. g. NIR radiation) passed through a sample.

[0035] "Diffuse reflectance" is light scattered from a reflecting surface in multiple directions.

[0036] "Integrating sphere" is a sphere coated with a reflective surface that collects light from a diffusely reflected surface and delivers it to a detector.

SPECIFIC EMBODIMENTS

[0037] A compound to be analyzed is first dissolved within a solvent. The solvent may be any suitable solvent useful in dissolving the compound. The solvent may be pure water or pure ethanol. Once the compound is completely dissolved in the solvent a solution is loaded into an aerosol generating device. The aerosol generating device may be any device, e.g. any device generally used in connection with pulmonary drug delivery. The device may create aerosolized particles which have a diameter in a range of from about 0.1 micron to 50 microns, or 1 micron to 25 microns or 2 microns to 10 microns.

[0038] Once the solution is created and loaded into the aerosol generating device an aerosol is created. The aerosol is generated and for purposes of conducting the assay the aerosol is forced through a filter. In general the aerosol is created in air and the filter is designed so as to allow the air to pass through the filter while collecting the particles of the aerosol on the filter. After the particles are collected on the filter an NIR spectra is obtained. The spectrum is examined to determine information. The examination and determining of information may require comparing the spectrum with other known reference spectra in order to determine the composition of the compound present within the solution.

[0039] In one embodiment, aerosols are generated using a pulmonary drug delivery device. The aerosols are collected onto a filter that is permeable to the air flowing through the device. NIR spectra are obtained in the diffuse reflectance mode using a NIR Spectrometer. Reference assays are performed by washing the same filters used for the non-destructive NIR method and determining the amount of one or more chemical components of the aerosol using a reference technique such as HPLC. A number of calibration methods may be employed. In an exemplary embodiment, calibration is performed using a PLS algorithm to develop a model that predicts the reference result using the second derivative of the NIR spectrum.

[0040] In another embodiment, the collection container or surface is coated with a material and/or has a solvent to aid in the delivery and/or collection of the delivered chemical(s).

[0041] In another embodiment, the NIR spectrum is acquired using other techniques than diffuse reflectance, including but not limited to specular reflectance, internal reflectance, transmittance, and fiber optic probes.

[0042] In another embodiment, there are multiple collection surfaces or containers for the purpose of obtaining the particle size distribution of the aerosol.

[0043] In another embodiment, the delivery is achieved by other means than pulmonary drug delivery devices, including but not limited to nasal delivery devices, buccal delivery devices, transdermal delivery devices or systems, syringe injectors, pen injectors and needleless injectors.

[0044] In another embodiment, the delivery is to other parts of the body, including but not limited to the eye, skin, nose, mouth, lungs, and any other bodily orifice.

[0045] In another embodiment, the delivered drug is for another area than the respiratory/pulmonary therapeutic area, including but not limited to the oncology, hematology, rheumatology/arthritis, cardiovascular, CNS/neurology, dermatology, endocrinology, immunology and gastrointestinal therapeutic areas.

[0046] When the NIR beam has a smaller area than the area of the collection material, sampling methods must be employed for the NIR assay. In one embodiment of a sampling method, referred to as manual sampling, glass fiber filters are placed between two square glass slides, then placed aerosol side down on top of the window of the integrating sphere of the diffuse reflectance NIR accessory. A reflecting material is placed on top of the slides to reflect near infrared light back into the integrating sphere. Several positions of the filter are scanned and the spectra merged into one spectrum. Several NIR scans per position are collected to provide an adequate signal-to-noise ratio for the NIR spectrum. The fraction of the filter surface area that the method samples depends on the diameter of the filter.

[0047] The presence of interference fringes in a spectrum is observed occasionally when scanning a filter using the manual method. These interference fringes are caused by small air gaps in the glass fiber filter (see Fig. 4). Whenever interference fringes are observed, the glass filter may be moved and re-scanned to improve the quality of the NIR spectrum.

[0048] In another embodiment of a sampling method, referred to as spinning cup sampling, a procedure is employed that allows most of the collection material to be sampled in one or more concentric rings. To accomplish this, an NIR spinning cup accessory is positioned so that the top edge of the NIR beam is at the center of the collection material. A reflecting material is placed on top of the collection material in the cup to reflect near-infrared light back into the integrating sphere. The spectrum is acquired while the accessory rotates the cup, so that a ring is sampled. If necessary, the accessory can then be positioned so that a second larger concentric ring, just touching the first one, is sampled for the second spectrum. This process can be repeated for a third concentric ring if desired. If more than one ring is sampled, all of the spectra obtained for each sample are merged into one spectrum.

[0049] An advantage of this sampling method is that it is easier to sample a large fraction of the collection material when the diameter is larger than the NIR beam diameter. Another advantage is that no interference fringes are observed when the collection material is a glass fiber filter or other material that may give rise to interference fringes.

EXAMPLES

[0050] The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the present invention, and is not intended to limit the scope of what the inventors regard as their invention nor is it intended to represent that the experiments below are all or the only experiments performed. Efforts have been made to ensure accuracy with respect to numbers used, but some experimental errors and deviations should be accounted for.

EXAMPLE 1

[0051] Aqueous aerosols were generated using an electronic AERx™ pulmonary drug delivery device equipped with a temperature controller to extrude protein formulation from dosage forms (See U.S. Patent 5,660,166 and related patents in the series). The aerosols were collected onto glass fiber filters contained in a plastic collection apparatus (see Fig. 1). Glass fiber filters were chosen because the glass is transparent to NIR radiation.

[0052] Dosage forms were extruded at dose levels nominally 20%, 40%, 60%, 80% and 100% of the full dose. Nine emitted doses were generated per dose level for a total of 45 filters. Filters were assayed first by the NIR method then by an HPLC method. A ThermoNicolet spinning cup accessory was used with an Antaris™ Fourier Transform NIR spectrometer from ThermoNicolet to acquire NIR spectra from aerosols deposited on the glass fiber filters. The spectra were collected from 4000 to 10,000 cm^"1 at 8 cm^"1 resolution using the Antaris diffuse reflectance accessory (see Fig. 2). A Teflon cylinder was placed on the filter in the cup to reflect the NIR beam back into the integrating sphere of the diffuse reflectance accessory. To improve the spectral signal-to-noise ratio, 30 scans were collected per spectrum. Fig. 3 shows an example spectrum of the protein formulation on the filter. [0053] Results of the HPLC assays were entered into the NIR computer and PLS calibration was performed on the second derivatives of the spectra using 30 of the filters for calibration and 15 for validation (3 at each dose level). The results of the calibration are shown in Figure 6. Only two factors were required in this calibration model. The correlation coefficient was 0.996, the RMSEC was 1.15, and the RMSEP was 0.87. No interference fringes were observed in any of the spectra. These results demonstrate that the accuracy and variability of the NIR ED method for this protein is comparable to the accuracy and variability in the HPLC method. The NIR method can thus replace the HPLC method with no loss in accuracy or precision.

EXAMPLE 2

[0054] Aqueous aerosols were generated using a mechanical AERx™ pulmonary drug delivery device to extrude cromolyn formulation from dosage forms. The experimental procedure was the same as for Example 1, except that filters were assayed first by the NIR method, then by a method employing a microtiter plate reader with UN detection. Fig. 5 shows an example spectrum of the cromolyn formulation on the filter. The water bands in Fig. 5 are larger than in Fig. 3 because the mechanical device does not contain a temperature controller to heat the aerosol.

[0055] Calibration was performed as in Example 1, except that all dose levels were nominally the same. The RMSEP was 2.4. No interference fringes were observed in any of the spectra. Figure 7 shows the results for the NIR and microtiter plate (MTP) assays of the samples.

[0056] A paired t-test showed no statistically significant difference between the NIR and MTP assay results presented in Figure 7. These results demonstrate that the accuracy and variability of the NIR ED method for cromolyn is comparable to the accuracy and variability in the microtiter plate method. The NIR method can thus replace the microtiter plate method with no loss in accuracy or precision. While the present invention has been described with reference to the specific embodiments thereof, it should be understood by those skilled in the art that various changes may be made and equivalents may be substituted without departing from the true spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation, material, composition of matter, process, process step or steps, to the objective, spirit and scope of the present invention. All such modifications are intended to be within the scope of the claims appended hereto.

Claims

CLAIMS That which is claimed is:

1. A method, comprising the steps of: aerosolizing a solution comprising a solute in a solvent so as to create aerosolized particles in a gas; directing the particles of the aerosol onto a surface; obtaining an NIR spectrum of the solution deposited on the surface; and comparing the NIR spectrum with reference data to obtain a calibration model.

2. The method as claimed in claim 1, wherein the calibration model is a calibration curve.

3. The method as claimed in claim 1 , wherein the surface is comprised of a material which is chosen from a material substantially transparent to frequencies used by an NIR spectrometer and a material which reflects substantially all frequencies used by an NIR spectrometer.

4. The method as claimed in claim 3, wherein the surface is comprised of a material chosen from a glass fiber filter and a mirror glass surface.

5. The method of claim 1 wherein the obtaining the NIR spectrum is repeated a plurality of times after the surface has been translated or rotated to present another portion of the solution deposited on the surface to an NIR beam.

6. The method of claim 1 wherein obtaining the NIR spectrum is carried out continuously while the surface is rotated and/or translated.

7. The method of claim 1 wherein the NIR spectrum is obtained using a reflectance chosen from specular reflectance, internal reflectance and diffuse reflectance.

8. The method of claim 1 wherein the NIR spectrum is obtained using transmittance.

9. The method of claim 1 wherein the NIR spectrum is obtained using a fiber optic probe and wherein the aerosolizing is carried out with an aerosol delivery device.

10. The method of claim 1 wherein the surface is comprised of a plurality of collection surfaces or containers and an analysis made of particle size distribution of the aerosol on the surfaces and wherein the aerosolizing is carried out with a pulmonary drug delivery device.

11. The method of claim 10, wherein the aerosol delivery device is chosen from a metered dose inhaler, a dry powder inhaler, a nebulizer and a liquid aerosol delivery system.

12. The method of claim 9 wherein the aerosol delivery device is a nasal delivery device.

13. A method, comprising the steps of: depositing a compound onto a surface of a material; and obtaining an NIR spectrum of the compound deposited on the surface held in a first position.

14. The method of claim 13, wherein the surface is a surface of a material chosen from a material substantially transparent to energy frequencies emited by an NIR spectrometer and a material which reflects substantially all frequencies emmitted by an NIR spectrometer.

15. The method of claim 14, further comprising : comparing the NIR spectrum with reference data to obtain a calibration model.

16. The method as claimed in claim 15, wherein the compound is deposited from a device used for the delivery of a pharmaceutical drug.

17. The method as claimed in claim 13, further comprising: rotating the surface relative to its first position to a second position and obtaining a second NIR spectrum; and comparing the NIR spectrum taken in the first and second positions to reference data and obtaining a calibration model.

18. A method, comprising the steps of: applying a pharmaceutically active drug from a delivery device onto a surface of a material in a first position; obtain a first NIR spectra of the drug on the surface while the surface is in the first position; moving the surface to a second position different from the first position relative to a NIR spectrometer used to obtain the NIR spectrum; obtain a second NIR spectrum of the drug while the surface in in the second position; and comparing the spectrum obtained in the first and second positions to reference data to obtain a calibration model.

19. The method of claim 18, wherein the material is chosen from a material substantially transparent to energy generated by an NIR spectrometer and a material comprising a surface which reflects substantially all energy generated by an NIR spectrometer.

20. The method of claim 19, wherein the drug is applied to the surface from a drug delivery device which creates an aerosol.