JPH0389146A - Evaluating apparatus of percutaneous absorption - Google Patents

Abstract

PURPOSE:To measure the emitting process of a medicine from a basic agent on the skin and the permeating process of the medicine through the skin respectively by the acoustooptic measurement and absorption photometry simultaneously and in real time by applying an acoustooptic measuring apparatus as a vertical diffusion cell. CONSTITUTION:A sensor part of this evaluating apparatus is comprised of an acoustooptic sensor part a vertical diffusion cell, an absorption photometry measuring part and a processing part. An acoustooptic sensor has a microphone arranged at an upper end thereof. The neighborhood of a pressing part at a lower end of the acoustooptic sensor is made of light-permeable material. Laser beams are introduced via an optical fiber to the acoustooptic sensor to irradiate a sample on the skin. An acoustooptic signal generated at this time is received and processed by the microphone, whereby the emitting process of a medicine from a basic agent on the skin is measured. At the same time, the permeating process of the medicine through the skin is simultaneously measured. The vertical diffusion cell is formed of silica to make part to be irradiated a rectangular parallelepiped. The absorption photometry part measures the light passing through the diffusion cell by a photocell. The laser beams are spectrally separated at this time, one intaraoduced to the diffusion cell to obtain a measuring signal of the absorption and the other introduced to the photocell as a reference signal.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of Application] The present invention relates to an apparatus for evaluating percutaneous absorption of drugs, which is indispensable for the development of pharmaceuticals and cosmetics.

[Prior Art] Appropriate evaluation experimental methods are essential for the development and design of transdermal absorption preparations. Numerous transdermal absorption experimental methods have been reported over the past 30 years, but the most commonly used method, which has been developed by many researchers, is the in vitro absorption method using a diffusion cell.
-This is an in vitro experiment. In-
The vitr○ experiment is a method in which the amount of drug that has permeated through the membrane is quantified at regular intervals, and the release rate of the drug from the base and the membrane permeation rate of the drug are determined from this change over time. The diffusion cells used can be divided into two types: parallel type and vertical type.

Parallel cells sandwich the skin between the donor and receiver sides, and both cells are agitated at a constant rate. The sample is placed as a solution on the donor side, and physiological saline or the like is placed on the receiver side. The vertical cell does not have a stirring device on the donor side and only stirs on the receiver side, and in this case it has the advantage of being able to apply not only a solution but also a base to the donor side, and can be said to be closer to the clinical situation.

In the skin permeation experiment method, the excised skin is first placed between a donor and receiver cell, and both sides of the cell are filled with physiological saline or a solvent. Next, a drug sample is applied to the cell on the stratum corneum side (time t=0), and samples (drugs) collected from the receiver at regular intervals are subjected to quantitative analysis using HPLC (high performance liquid chromatography) or the like. The drug concentration in the receiver is proportional to time as a function of time and after a lag time to steady state. Data commonly used are flux and transmission coefficient. These values are obtained experimentally by the following equations.

J=PAΔC=■・dc/dt J is steady state f lux (total 5tead
y-state f lux), P is the transmission coefficient, and A is the diffusion area. ΔC is the concentration gradient of the drug in the skin, and in actual experiments, the dissolved 'fi concentration in the donor cell is used. d c /d t is the slope of the straight line of the obtained data, and ■ is the volume of the receiver cell. Therefore, the permeability coefficient of the drug can be determined experimentally using the following equation.

P= (V ・d c/d t) / (A ・ΔC)
A release experiment from a base is performed by placing a base such as an ointment on the donor side of the cell used in the permeation experiment. 5i on receiver cell
Add purified water etc. as nk. In addition, if the base is water-soluble and
When eluting into an ink solution, there are methods such as sandwiching a membrane with very fast permeation between the base and the 5 ink solution, or using a gel such as agar in place of the 5 ink solution. The amount of drug released is quantified using the same method as in the skin permeation experiment described above. The data obtained is based on Iguchi's diffusion theory (T
, Higuchi, J., Soc., Cosmet.
, Chem, 1U, 375 (1986),), it is proportional to the square root of time.

Therefore, it is necessary to determine experimental conditions depending on the purpose of each experiment, whether it is a drug permeation experiment through the skin or a drug release experiment from a base.

[Problems to be solved by the invention] However, in-vitro studies using this diffusion cell
Transdermal absorption experiments pose the following problems.

One is that bubbles are likely to be generated at the interface between the membrane and the solvent because an appropriate amount of the solvent (physiological saline) in the diffusion cell is extracted during HPLC measurement. This affects the reproducibility and precision of results in long-term measurements. Second, the purpose of the experiment is to measure the release rate of the drug from the base and the membrane permeation rate of the drug, and it is necessary to perform separate measurements under different conditions. Therefore, it is not possible to measure the release process from the drug base to the skin and the skin permeation process of the drug actually applied (10 drug bases) using the same system. If these two problems are solved, information regarding the base, drug, and skin permeation can be easily obtained in just one measurement.

The present invention was made in view of the problems of the prior art described above, and its purpose is to use a vertical diffusion cell, which is a system closer to clinical conditions than a parallel cell, to remove blood from the base on the donor side. The object of the present invention is to provide a transdermal absorption evaluation device that can simultaneously measure the drug release process and the skin permeation process of the drug on the receiver side with high sensitivity in real time without sampling.

[Means for Solving the Problems 1] In order to achieve the above object, the transdermal absorption measuring device according to the present invention includes a vertical diffusion cell made of quartz processed into a cubic shape.
It includes a photoacoustic measurement section and an absorbance measurement section.

Here, in a special feature in which a laser beam is used as a light source for measuring the photoacoustic W signal and absorbance, one laser beam is split into two directions and used for photoacoustic measurement and absorbance measurement, respectively. It is characterized by having been.

The photoacoustic W measurement unit consists of a cylindrical body that is open at one end and can be inserted almost airtight onto the sample surface, and a microphone is installed at the other end. When inserted, the sample surface can be irradiated with light by a light guide provided in the cylinder.

Here, the vicinity of the insertion part of the photoacoustic measuring part with the sample is made of a light-transmitting material, and is characterized in that this part is provided with a light guide.

Moreover, in the above-described apparatus, it is preferable that the light-transmitting material used in the photoacoustic measuring section be made of quartz glass.

The part formed of the light-transmitting material is about 10 to 10 minutes from the insertion part.
It is also suitable that it is 50 mm.

It is also preferable to make the capacity of the cylinder forming the photoacoustic measuring section variable.

The absorbance measurement section performs measurements by transmitting laser light through a vertical diffusion cell made of processed quartz.

At this time, the light irradiation part of the cell is characterized in that it is perpendicular to the light.

Further, it is also preferable that the light irradiation part be located near the center of the cell.

Therefore, in the present invention, by splitting the modulated laser beam into two directions, it has become possible to perform photoacoustic measurement and absorbance measurement simultaneously in real time.

[Function] The percutaneous absorption evaluation device according to the present invention has the above-mentioned means, so it can be used in an in-house using a vertical diffusion cell similar to a clinical system.
In vitro transdermal absorption evaluation, it becomes possible to simultaneously measure in real time the process of drug release from the base on the skin by photoacoustic measurement, and the process of drug permeation through the skin by spectrophotometry.

Therefore, compared to conventional methods, it is possible to measure the drug release process from the base and the skin permeation process of the N agent in an actually used formulation in a system that is closer to clinical conditions in one go.

[Embodiment 1] Hereinafter, a preferred embodiment of the present invention will be described based on the drawings.

FIG. 1 shows a portion of a sensor of a transdermal absorption measuring device according to an embodiment of the present invention.

The sensor portion of the percutaneous absorption evaluation device shown in the same figure includes a portion of a photoacoustic sensor, a vertical diffusion cell made of processed quartz, an absorbance measuring section, and a processing section.

In the photoacoustic sensor, a microphone is placed at the upper end in the figure, and the lower end is open to form a skin insertion part. A skin contact rubber (〇-ring) is installed in the skin insertion part,
Keeps the space between it and the skin almost airtight.

Further, the microphone is formed into a block, and the outer periphery of the block is threaded into a male thread. and,
The photoacoustic sensor is screwed into the female-threaded part at the top of the sensor so that its vertical position can be adjusted freely.

The characteristic feature of the present invention is that the vicinity of the part where the photoacoustic sensor is inserted into the skin is made of a light-transmitting material,
For this reason, in this embodiment, the photoacoustic sensor is made of quartz glass over a distance of <40 mm from the tip.

Note that the space between the upper end of the quartz glass portion and the microphone is made of a material such as brass, as in the prior art.

The vertical diffusion cell is made of quartz instead of glass, and the light irradiation area is made into a rectangular parallelepiped so that absorbance can be measured.

Note that during measurement, the physiological saline in the diffusion cell is stirred using a rotor and a magnetic stirrer in the same manner as in the conventional method.

The absorbance measurement section is configured with a system that measures light transmitted through a diffusion cell using a photocell.

At this time, the laser beam for absorbance measurement is roughly split into two directions, one of which is guided to a diffusion cell to obtain an absorbance measurement signal, and the other is guided to a photocell as a reference signal.

The processing section is the microphone and the photocell.

Here, the entire device system including a part of the sensor of the percutaneous absorption evaluation device is shown in FIG.

The argon laser beam used as a light source is modulated by an optical chopper, and then split into two directions by a beam splitter, which are used for photoacoustic and absorbance measurements, respectively.

The photoacoustic measuring device according to this embodiment is roughly configured as described above, and its operation will be described below.

Increasing the sealing degree of the gas inside the cell, which has been a conventional measure to improve sensitivity, causes problems in usability, and it is physically impossible to completely seal the gas inside the cell.

Therefore, in this example, by utilizing the semi-sealed state of the gas inside the cell, a high frequency (
Helmholtz resonance is generated at a frequency of 3 kHz or higher), and the resonance frequency unique to the photoacoustic cell is measured. Therefore, the signal is maximum, and by minutely changing the capacitance of the cell, it is possible to optimally increase the signal at the resonant frequency.

Moreover, since only the resonant frequency components of the early signal are extracted by the lock-in amplifier, in-situ
In the Il determination, only the desired photoacoustic signal can be measured.

That is, the photoacoustic sensor is inserted onto the sample surface via an O-ring.

In the photoacoustic sensor, modulated laser light is guided from a laser light source through an optical fiber, and a sample (
Locally irradiate the drug (10 drug bases).

By the way, when the sample to be measured is a strongly scattering substance such as skin or ointment, the influence of reflected light and scattered light from the sample cannot be ignored.

Therefore, as mentioned above, in this example, by providing a quartz glass part of an appropriate length (40 mm) in the part of the photoacoustic cell that is inserted into the sample, reflected light and scattered light from the sample is directed to the outside without any influence. The aim is to improve accuracy and sensitivity.

Note that by setting the length of the quartz glass portion to 10 mm to 50 mm, a sufficient performance improvement can be achieved in practice.

The photoacoustic signal generated within the sample is converted into a gas compressional wave, and the sound is received by a microphone attached to the other side of the cell.

In addition, the position of the microphone attached to the threaded block can be adjusted using a screw, and the volume inside the cell can be adjusted.

By changing the volume inside the cell, Helmholtz resonance optimal for the resonance frequency at the time of measurement can be caused.

At this time, when the volume changes by 10 μL, the resonance frequency changes to 3H.
Since z changes, a thread cutter that allows minute adjustments is required.

Then, as shown in FIG. 2, the signal S1 detected by the photoacoustic sensor is input to a lock-in amplifier, and the lock-in amplifier converts only the modulation frequency of the irradiation light and the same frequency component generated by the optical chopper into 81 Take it out and output it. The signal is then recorded using a chart recorder and analyzed using a computer.

As described above, the process of drug release from the base on the skin can be measured by the action of the photoacoustic sensor.

Next, we will explain the function of the absorbance method, which simultaneously measures the skin permeation process of the drug in real time and is structured as outlined above.

Conventionally, in the skin permeation process of drugs, a solvent (such as physiological saline) in a diffusion cell is periodically sampled and quantitatively analyzed using HPLC or the like. This method tends to generate bubbles at the skin-solvent interface, which affects the accuracy of the data obtained and is difficult to operate.

Furthermore, since laser light has excellent wavelength characteristics, measurements can be made with higher sensitivity than conventional methods.

Therefore, in this example, in order to quantify the drug that permeates the skin without collecting the solvent, absorbance measurement is performed using laser light.

One of the modulated laser beams (the other beam is used for photoacoustic measurement) is roughly split into two directions, one of which is sent to a vertical diffusion cell made of quartz that can measure absorbance, and the other is sent to a reference signal (■2 ) is guided to a photocell, which is a detector.

The measurement signal (11) and the reference signal (Step 2) are guided to an operating amplifier, and the computer determines the absorbance based on this difference signal (■3).

Simultaneously at this time, analysis of the photoacoustic signal and changes in absorbance over time is performed.

By simultaneously measuring the photoacoustic signal and the absorbance in real time as described above, it is possible to easily and quickly measure the drug release process from the base and the skin permeation process of the drug in one measurement.

Next, an actual measurement example will be explained.

An in-vitro transdermal absorption evaluation experiment was conducted using the skin of a hairless mouse using the apparatus shown in FIGS. 1 and 2. 6 mg of 3% shikonin ointment (equivalent to 180 μg of shikonin) was applied to a 5 mm diameter area of the skin of a hairless mouse (up to the dermis layer, the fat area was removed), and physiological saline was used as the solvent in the diffusion cell, and photoacoustic Simultaneous signal and absorbance measurements were performed. The results are shown in FIGS. 3, 4 and 5.

In FIG. 3, the vertical axis represents absorbance and photoacoustic signal intensity, and the vertical axis represents time.

As the optical sound W signal decreased, an increase in absorbance was confirmed.

In other words, a decrease in the amount of shikonin on the skin and an accompanying increase in the amount of shikonin that permeated the skin were simultaneously observed. The drop in absorbance is due to floating matter derived from the skin.

At the end of the measurement after 25 hours, the interface between the skin and the solvent was observed, but no bubbles were observed.

In FIG. 4, the vertical axis represents the quantification of absorbance, and the horizontal axis represents time. Hence, the skin permeation rate of the drug.

The amount of shikonin that permeated the skin increased in proportion to time after a 7-hour lag time, and after 25 hours, it was 3.7% of the applied amount.
It was observed that shikonin permeated through the skin. The reason why the lag time was longer than usual is probably because the temperature of the physiological saline in the receiver was measured at room temperature (200C).

In Figure 5, the vertical axis indicates the decrease value of the photoacoustic signal (the decrease value of the signal from the initial photoacoustic signal), and the horizontal axis (the decrease value of the signal from the initial photoacoustic signal) indicates time. The decrease value of the photoacoustic W signal corresponds to the amount of decrease in shikonin on the skin.Therefore, if we consider the change in the obtained photoacoustic signal over time as the process of drug release from the base, Higuc
It showed a good correlation with the diffusion theory of hi.

From the above results, by simultaneous measurement of photoacoustic signal and absorbance,
In-vitro transdermal absorption percutaneous absorption evaluation method It has been found that the drug release process from the base and the drug permeation process through the skin can be simultaneously measured in real time in a system similar to bed conditions.

As explained above, according to the percutaneous absorption evaluation device according to the present example, in an in-vitro percutaneous absorption experiment using a vertical diffusion cell, which is a system close to clinical conditions, skin absorption was measured by photoacoustic measurement. It is possible to measure the drug release process (see above) and the skin permeation process of the drug by absorbance.

Then, apply BB○ to a visible laser such as an argon laser.
By combining nonlinear optical crystals such as , it is possible to oscillate ultraviolet light and measure many drugs.

Therefore, by varying the wavelengths of the light source, including infrared, visible, and ultraviolet light, this device can be expected to have a wide range of applicability as a device that can obtain information regarding the penetration of base materials, drugs, and skin.

Note that the percutaneous absorption evaluation device according to the present invention uses a photoacoustic sensor that can set the optimal volume at the resonance frequency, and as long as it has a structure that simultaneously measures absorbance, the shape of the diffusion cell, etc. The size, length, etc. are not limited in any way. [Effects of the Invention] Since the present invention is configured as described above, it produces the following effects.

According to the transdermal absorption evaluation device according to claim (1), the photoacoustic method, which is effective for measuring strongly scattering substances such as ointments, can be applied to in-vitro transdermal absorption using a commonly used diffusion cell. By applying this method to experiments for the first time, it becomes possible to develop a new percutaneous absorption evaluation device.

According to the apparatus according to claim (2), by using a laser beam as a light source, highly sensitive drug analysis becomes possible.

According to the device according to claim (3), in-vivo,
By using a photoacoustic measurement device capable of in-situ measurement, an in-vitr○ transdermal absorption evaluation device that is closer to clinical conditions can be achieved.

According to the device according to claim (4), the optical sound W8 can be produced with high sensitivity.
! It becomes possible to measure the absorbance in real time at the same time as II constant.

According to the device according to claim (5), it becomes possible to newly measure the drug release process from the base on the skin in a system similar to clinical conditions.

According to the device according to claim (6), it is possible to quantify the amount of drug that has permeated the skin without collecting the solvent in the diffusion cell.

According to the device according to claim (7), the process of drug release from the base can be determined by -times of measurement using a preparation in actual use, and
The process of drug permeation through the skin can be measured. Therefore, information on bases, drugs, and skin permeation can be obtained easily and quickly.

[Brief explanation of drawings]

FIG. 1 is an explanatory diagram of a part of the sensor of a percutaneous absorption evaluation device according to an embodiment of the present invention, FIG. 2 is an explanatory diagram of the circuit configuration of the device shown in FIG. 1, and FIG. , is an explanatory diagram of the measurement results of an in-vitro transdermal absorption evaluation experiment using the skin of a hairless mouse using the apparatus shown in FIG. 1. FIG. 4 is an explanatory diagram of the results of quantifying the change in absorbance over time in the measurement results of FIG. 3. FIG. 5 is an explanatory diagram of the measurement results of FIG. 3, in which the change over time in the photoacoustic signal is expressed as a decrease value from the initial signal.

Claims (7)

[Claims] (1) A transdermal absorption evaluation device characterized in that a photoacoustic measuring device is applied to a vertical diffusion cell commonly used in in-vitro transdermal absorption experiments. (2) A transdermal absorption evaluation device according to claim (1), characterized in that a laser beam is used. (3) The device according to claim (1) or (2), which uses a photoacoustic measurement device (patent application filed on January 27, 1988) capable of in-vivo, in-situ measurement. A transdermal absorption evaluation device characterized by: (4) A percutaneous absorption evaluation device according to claim (1), characterized in that a vertical diffusion cell made of cubic quartz is used. (5) A transdermal absorption evaluation device according to claim (1), characterized in that the device according to claim (3) measures changes over time in the amount of decrease in the amount of drug on the membrane. (6) A transdermal absorption evaluation device according to claim (4), characterized in that the change over time in the amount of drug that has permeated through the membrane is measured by absorbance. (7) A transdermal absorption evaluation device capable of simultaneously performing the measurements according to claims (5) and (6) in real time.