WO2020228871A1

WO2020228871A1 - Apparatus for measuring the permeability of membranes and a method for measuring the same

Info

Publication number: WO2020228871A1
Application number: PCT/CZ2020/050013
Authority: WO
Inventors: Martina VRABLOVA; Ivan Koutnik
Original assignee: Vysoka Skola Banska - Technicka Univerzita Ostrava
Priority date: 2019-05-15
Filing date: 2020-03-17
Publication date: 2020-11-19
Also published as: DE212020000628U1; CZ2019301A3; CZ308388B6

Abstract

An apparatus for measuring the permeability of membranes (5) comprising an optical prism (1), at the face (101) of which a metal coating (11) is applied that is provided with at least one detection spot (12), where outside of the body of the optical prism (1), a source (2) of electromagnetic beams (21), a mirror (22), and a sensor (3) are arranged, which are arranged with respect to one another in such manner that after reflection from the mirror (22), the electromagnetic beams (21) emitted from the source (2) pass through the body of the optical prism (1) towards its face (101) and, after reflection from the metal coating (11), fall on the receiving area of the sensor (3). On the metal coating (11), a measuring cell (4) is arranged that consists of a detection member (41) abutting the surface of the metal coating (11), a flow-through member (42), and a pressure module (43), where the detection member (41) as well as the flow-through member (42) are formed in the shape of frames, between which at least one membrane (5) is placed, and where the detection member (41) is provided with at least one cut (412), the geometry of which determines the perpendicular projection of the effective area of the membrane (5) on the area of the face (101) of the prism (1), and the flow-through member (42) is provided with a window (422), thereby forming in the measuring cell (4), after assembling the apparatus, at least one measuring chamber (411) and one gastight flow-through chamber (421) separated from one another by the measured membrane (5), wherein each measuring chamber (411) is arranged above at least one detection spot (12), and the pressure module (43), which closes the flow-through chamber (421), is provided with an inlet channel (431) and an outlet channel (432), which are situated above the peripheral regions of the flow-through chamber (421).

Description

Apparatus for measuring the permeability of membranes and a method for measuring the same

Technical Field

The present invention falls within the field of examination and analysis of the physical and chemical properties of materials and relates to an apparatus for measuring the permeability of membranes and a method for measuring the same.

Background of the Invention

Structures that make up interfaces between two environments separated from one another are called membranes. A property typical of membranes is selective permeability to different substances with different properties. The permeability rate of a given substance may be conditioned by a number of physicochemical properties, typically the size of the particles comprising the substance, polarity, electric charge. The ratio between the permeabilities of two substances through a membrane is called selectivity. As mentioned, for example, in the publication PLAWSKY, Joel L, Transport phenomena fundamentals, 3rd ed., CRC Press, 2014, ISBN 978-1 -4665-5533-4, one of the processes mediating the transport of substances through the membrane is diffusion, which is the relative motion of one component of the mixtures with respect to the rest of the mixture. This motion is most often due to the difference in the concentration of the substance between two locations within the mixture’s volume. Diffusion of the substance within the liquid mixture depends on time, temperature, viscosity, and mutual miscibility. Therefore, diffusion of the substance through the membrane particularly depends on the shape of the membrane’s pores and the proportion of their size to the size of the particles of the diffusing substance and on the polarity rate of pore surface relative to the polarity of the diffusing substance.

At present, measuring the permeability of semi-permeable membranes is usually performed using several commonly known methods. One of the most widely used methods is based on the use of the Franz diffusion cell, as described in a number of publications, such as Danha, Liu, et. al., Permeation measurement of gestodene for some biodegradable materials using Franz diffusion cells, Saudi Pharmaceutical Journal, Volume 23, Issue 4, September 2015, 413-420, or Mauricio Espinal-Ruiz, et.al., Effect of pectins on the mass transfer kinetics of monosaccharides, amino acids, and a corn oil-in-water emulsion in a Franz diffusion cell, Food Chemistry, Volume 209, 15 October 2016, 144-153, or Alves, Ana Catarina, On-line automated evaluation of lipid nanoparticles transdermal permeation using Franz diffusion cell and low-pressure chromatography, Talanta, Volume 146, 1 January 2016, 369-374. The basic model of a Franz cell consists of two chambers located on top of each other, separated by a measured membrane of a known area. The upper chamber is filled with a solution containing a solvent and a substance, diffusion of which is observed through the membrane, and the lower chamber is filled with a pure solvent. The permeability rate is assessed based on the time dependencies of concentration changes in both of the chambers. The concentrations may be determined by discontinuous sampling through a sampling port of each of the chambers. The taken samples are subsequently analysed using an analytical method with an appropriate sensitivity and detection limit. The methods commonly used today include chromatography, conductivity detection, or spectrometry in the ultraviolet, visible, and infrared region. Modified models of the Franz diffusion cell are usually equipped with additional input and output channels of the individual chambers, which may either serve for the application of cross-flow techniques, i.e. techniques of cross flow, or for connecting an analyser of a flow-through type allowing for continuous measurement of concentrations with an online output. Another option how to continuously monitor concentrations with an online output is to introduce probes of a suitable type into the chambers. To eliminate the effect of free diffusion of substances in the solutions, homogenisation of the contents of the chambers using a stirrer is provided. Measuring cells in a so-called side-by-side arrangement, where the chambers are located next to each other and the separating membrane is oriented vertically, represent a slightly different variant. Said methods are also used to characterise membranes of biological origin, as mentioned in, for example, the publication Viktoria Valeska Zeisler-Diehl, Britta Migdal and Lukas Schreiber, Quantitative characterization of cuticular barrier properties: methods, requirements, and problems, Journal of Experimental Botany, Vol. 68, No. 19, 2017, 5281-5291 , or Remus-Emsermann, M. N., de Oliveira, S., Schreiber, L., Leveau, J. H., Quantification of lateral heterogeneity in carbohydrate permeability of isolated plant leaf cuticles, Frontiers in microbiology, 2011 , 2, 197.

One of the rapidly developing optical detection techniques is surface plasmon resonance with imaging using CCD sensors, or Surface Plasmon Resonance imaging, SPRi, described in, for example, the paper by Homola,

Surface plasmon resonance sensors for detection of chemical and biological species, Chemical reviews 108 (2), 462-493. It is an optical method that uses the incidence of polarised light through an optical prism on a thin gold layer, where resonance of the surface plasmon occurs, and the reflected light is subsequently detected in the CCD detector. At a certain incidence angle, the intensity of the reflected light is reduced. Solvent solution, or solvent with an analyte flows on the other side of the metal layer. In the presence of the analyte, the optical properties of the flowing solution are changed, changing the index of refraction near the gold layer and shifting the resonance angle of the reflected light. The shift of the resonance angle, or, in case of a fixed angle, the shift of the light intensity, is detected in time as an SPR signal corresponding to the concentration of the analyte. In the SPRi measurement, the area of the prism with the size of approximately 0.5 x 0.7 cm is imaged, which may be divided into tens or hundreds of spots using software; the presence of the analyte may thus be detected not only with regard to time but also with regard to space. The analysis may be performed in flow-through mode with a selectable liquid flow rate, most often, the solution flow rate of 50 mI/min, or static state is selected. In case of implementation of a membrane into the system, the measurement may be performed on a liquid-liquid or gas-liquid interface, where the liquid must be in contact with the prism.

The use of the SPRi method to determine the concentrations of an aqueous solution of raffinose and methanol using an agar membrane created in situ on the surface of an SPR chip is described in the publication Menchie E. Montecillo, Toshifumi Yoshidome, Toshiyuki Yamagata, Tatsuya Yamasaki, Masaru Mitsushio, Brian John Sarno, Morihide Higo, Concentration Determination of Individual components in Methanol-Raffinose Mixtures Using Diffusion through Agar Membrane Attached to a Surface Plasmon Resonance Sensor, Bull. Chem. Soc. Jpn., Vol. 83, No. 12, 2010, 1531-1533. Based on the different diffusivity of raffinose and methanol in the agar layer and in combination with the dependence of the SPR resonance angle on the refractive index, the concentration of raffinose and methanol is calculated using a mathematical model. However, the described solution does not allow for the characterisation of membranes. The preparation itself of the in-situ created hydrogel membrane on the surface of the SPR biochip is discussed in the publication Khulan Sergelen, Christian Petri, Ulrich Jonas, Jakub Dostalek, Free-standing hydrogel-particle composite membrane with dynamically controlled permeability, Biointerphases 12, 051002 (2017). The created membrane is used for the pre-separation of substances based on the size of their particles, which creates new options how to determine the components of mixtures of low-molecular-weight and macromolecular substances. However, the characterisation of the membranes is not described here. The design of the apparatus in which separation of substances with different molecular weights takes place on a membrane in the form of a hollow fibre and its connection to an SPR detector are described in the article Richard C. Stevens, Scott D. Soelberg, Steve Neara, Clement E. Furlong, Detection of cortisol in saliva with a flow-filtered, portable surface plasmon resonance biosensor system, Anal Chem., 2008, September 1 , 80(17), 6747-6751. The separation extends the applicability of SPR and allows for the determination of cortisol directly in blood plasma. Said embodiment does not allow for either the use of planar membranes or their characterisation. The publication Julien Breault-Turcota and Jean- Francois Masson, Microdialysis SPR: diffusion-gated sensing in blood, Chem. Sci., 2015, 6, 4247-4254 mentions the use of a membrane in conjunction with SPR detection for the separation of particles based on their size. In the publication, this method is applied to the analysis of blood plasma, and it is shown that if the membrane is selected properly, the differences in the diffusivities of the substances may be utilised for their “pre-separation” and subsequent detection at different times depending on the size of the particles of the given substances. However, the described solution is not equipped with a CCD sensor, and the detection does not take place on individual spots, instead, it takes place throughout the area of the channels, thereby determining the necessary shape and area of the membrane. Furthermore, it is discontinuous measurement that does not allow for maintenance of constant concentration on the inlet side of the membrane. The membrane characterisation itself is not described here.

At present, no method of utilising the SPRi technique to determine the permeability of solid, so-called free-standing, membranes, is known.

The object of the present invention is to present a method for measuring the permeability of membranes using techniques of surface plasmon resonance using a CCD detector. It is a further object of the invention to present an apparatus for carrying out this method.

Summary of the Invention

The object is achieved with an invention that is an apparatus for measuring the permeability of membranes comprising an optical prism, at the face of which metal coating is applied that is provided with at least one detection spot, where outside of the body of the optical prism, a source of electromagnetic beams, a mirror, and a sensor are arranged, which are arranged with respect to one another such that after reflection from the mirror, the electromagnetic beams emitted from the source pass through the body of the optical prism towards its face and, after reflection from the metal coating, fall on the receiving area of the sensor. The invention is characterized in that on the metal coating, a measuring cell is arranged, which consists of a detection member abutting the surface of the metal coating, a flow-through member, and a pressure module, where the detection member as well as the flow-through member are formed in the shape of frames, between which at least one membrane is placed, and the detection member is provided with at least one cut, geometry of which is determined by the perpendicular projection of the effective area of the membrane to the area of the face of the prism, and the flow-through member is provided with a window, thereby forming in the measuring cell, after assembling the apparatus, at least one measuring chamber and one gastight flow-through chamber separated from one another by the measured membrane, wherein each measuring chamber is arranged above at least one detection spot, and the pressure module, which closes the flow-through chamber, is provided with an inlet channel and an outlet channel, which are situated above the peripheral regions of the flow-through chamber.

In a preferred embodiment, the detection member is formed in the shape of a frame provided with cuts, the number of which corresponds to the number of the measuring spots, wherein each of the measuring chambers is separated from the flow through chamber by a different membrane. The invention is also characterized by a method for measuring the permeability of membranes using the said apparatus, where, after assembling the measuring cell, when the membrane is inserted between the detection member, the measuring chamber of which is filled with the liquid selected, and the flow-through member, while a fluid medium with a different concentration of the dissolved substances than that of the liquid in the measuring chamber is flowing through its flow-through chamber, the time concentration response of the substance diffusing through the membrane, based on which the value of the permeability of the membrane is subsequently determined, is measured using monitoring and assessment of the changes in signals of electromagnetic beams passing through the body of the prism, incident on the detection spots of the metal coating, and captured in the sensor.

With the present invention, a new and improved effect is achieved, by the characterisation of the membranes using the detection capability of the surface plasmon resonance technique which allows for measurement of the permeability of membranes at a miniaturised scale, also, simultaneous characterisation of multiple membranes is further possible, and, finally, areal characterisation of the permeability of membranes is also possible. The principle of the SPRi technique, i.e. the use of a CCD detector, thus provides for the possibility of simultaneous online specific detection of a larger number of substances on a large number of measuring spots on the surface of the metal coating.

Description of the Drawings:

Specific exemplary embodiments of the invention are schematically shown in the appended drawings, where:

Obr.1 is an overall schematic cross-section of the apparatus for measuring the permeability of membranes,

Obr.2 is a schematic drawing of a cross-section of the measuring cell of the apparatus of fig. 1 ,

Obr.3 is an exploded view of parts of the measuring cell of the apparatus with a set of measuring spots in one measuring chamber, Obr.4 is an exploded view of an alternative arrangement of a part of the measuring cell of the apparatus with a single measuring spot in the measuring chamber,

Obr.5 is an exploded view of an alternative arrangement of a part of the measuring cell of the apparatus with a set of measuring cells,

Obr.6 is an exploded view of an alternative arrangement of a part of the measuring cell of the apparatus with two measuring cells and two membranes,

Obr.7 is an exploded view of another alternative arrangement of a part of the measuring cell of the apparatus,

Obr.8 is a record of increase of urea concentration over time when measuring according to example 1 ,

Obr.9 is a record of increase of fructose concentration over time when measuring according to example 2,

Obr.10 is a record of increase of sucrose concentration over time on individual detection spots when measuring according to example 3, and

Obr.1 1 is an arrangement of the record detection spots of fig. 10.

The drawings showing the presented invention and the subsequently described examples of particular embodiments do not, in any way, limit the scope of protection stated in the definition, instead, they merely clarify the essence of the invention.

Exemplary Embodiments of the Invention

The apparatus for measuring the permeability of membranes is, in the basic embodiment shown in figs. 1 to 3, consisting of an optical prism 1, on a face 101 of which a metal coating 11_, which is provided with a set of detection spots 12 and on which a measuring cell 4 is seated, is applied. Outside of the body of the optical prism 1, a source 2 of electromagnetic beams 21_, a mirror 22, and a sensor 3 are arranged, which are arranged with respect to one another such that after reflection from the mirror 22, the electromagnetic beams 21_ emitted from the source 2 pass through the body of the optical prism 1 towards its face 101 and, after reflection from the detection spots 12 of the metal coating 11_, fall on the receiving area of the sensor 3. The measuring cell 4 consists of a detection member 41_, a flow-through member 42, and a pressure module 43. where the detection member 44. as well as the flow-through member 42 are formed in the shape of frames, between which a membrane 5 is arranged, wherein the detection member 41_ is provided with a cut 412 and the flow-through member 42 with a window 422, thereby forming, in the measuring cell 4^ after assembling the apparatus, a measuring chamber 441. and a flow-through chamber 421 separated from one another. The pressure module 43 is provided with an inlet channel 431 and an outlet channel 432, which are situated above the peripheral regions of the flow-through chamber 421. Therefore, the flow-through chamber 421 is, as is obvious from fig. 1 and fig. 2, designed as gastight, as the fluid medium may be a gas as well. Alternatively, only one detection spot 12 is situated in the measuring chamber 411 , as shown in fig. 4.

Another alternative is the embodiment shown in fig. 5, where a set of cuts 412 is formed in the detection member 44_., which, after assembling the measuring cell 4, are used to form a corresponding number of measuring chambers 411 , wherein one detection spot 12 is situated in each measuring chamber 411.

Furthermore, another alternative is the embodiment shown in figs. 6 and 7, where the detection member 44_. comprises two cuts 412. which are, after assembling the measuring cell 4, used to form two measuring chambers 411. of different shapes, wherein each of the measuring chambers 441. is separated from the flow-through chamber 421 by a different membrane 5 and one detection spot 12 is situated in each measuring chamber 411.

In the method of measuring the permeability of membranes 5 using an SPRi measuring device equipped with an apparatus according to the invention, a membrane 5 is inserted between the detection member 44_. and the flow-through member 42, the measuring chamber 441 is filled with the liquid selected, and in the flow-through chamber 421 , a flow of a fluid medium of a different concentration of the dissolved substance than that in the measuring chamber 441 is set. The response of the signal obtained on the detection spot 12 or on multiple spots 12 is recorded in sensor 3. Example 1

In the embodiment of fig. 4, a polyestersulfone membrane 5 of the pore size of 50 kilodaltons was inserted between the detection member 41 and the flow-through member 42. The measuring chamber 411 was filled with water treated with reverse osmosis with a resistance of at least 18 megaohms. Urea solution with the concentration of 8 g.dm^-3 was fed into the flow-through chamber 421 above the membrane 5 at the rate of 50 ml.min^-1. The detection took place in a non-specific manner on the detection spot 12. From the record of increase of urea concentration at a time shown in fig. 8, a diffusion coefficient was determined at time t = 16 min, being 0.1616.

Example 2

In the embodiment of fig. 7, two different membranes 5 of a natural origin, Ficus cuticles, were inserted between the detection member 41 and the flow-through member 42 in such manner that one membrane 5, an adaxial cuticle with a lower permeability, separates one measuring chamber 411 from the flow-through chamber 421 and the other membrane 5, an abaxial cuticle with a higher permeability, separates the other measuring chamber 411 from the flow-through chamber 422. The measuring chambers 411 were filled with water treated with reverse osmosis with a resistance of at least 18 megaohms. A fructose solution with the concentration of 8 g.dm^-3 was fed into the flow through chamber 421 at the rate of 50 ml.min^-1. The detection took place in a non specific manner on the detection spots 12. From the record of increase of the fructose concentration at a time shown in fig. 9, a diffusion coefficient was set for the abaxial cuticle at time t = 68 min, being 1.177 x 10^-3, and for the adaxial cuticle at time t = 85 min, being 1.611 x 10^-4.

Example 3

In the embodiment of fig. 3, a membrane 5, which was the adaxial cuticle, was inserted between the detection member 41 and the flow-through member 42. The measuring chamber 411 was filled with a sucrose solution with the concentration of 8 g.dm^-3. Air was fed into the flow-through chamber 421 at the rate of 50 ml.min^-1. The detection took place in a non-specific manner on nine detection spots 12 arranged in a 3x3 square matrix. From the record of increase of the sucrose concentration at time on individual detection spots 12, which is shown in fig. 10, diffusion coefficients were determined at time t = 12 min, and these were arranged in the following matrix such that for each coefficient, the row and column indexes correspond to the detection spots 12 on which they were measured.

Industrial Applicability

The invention is usable in the examination of the interaction of plants with the external environment, including the transport of substances through the plant cuticles and receiving substances by the plant through the surface of the plant body, and, furthermore, in the research of biomechanisms of living organisms, and also in the development of industrial separation methods.

Claims

C L A I M S

1. An apparatus for measuring permeability of membranes (5) comprising an optical prism (1 ), on a face (101 ) of which a metal coating (11 ) is applied, which is provided with at least one detection spot (12), where outside of the body of the optical prism (1 ), a source (2) of electromagnetic beams (21 ), a mirror (22), and a sensor (3) are arranged, which are arranged with respect to one another such that the electromagnetic beams (21 ) emitted from the source (2) pass, after reflection from the mirror (22), through the body of the optical prism (1 ) towards its face (101 ) and, after reflection from the metal coating (11 ), fall on the receiving area of the sensor (3), characterised in that on the metal coating (11 ), a measuring cell (4) is arranged that consists of a detection member (41 ) abutting the surface of the metal coating (11 ), a flow-through member (42), and a pressure module (43), where the detection member (41 ) as well as the flow through member (42) are formed in the shape of frames, between which at least one membrane (5) is placed, and where the detection member (41 ) is provided with at least one cut (412), the geometry of which is determined by the perpendicular projection of the effective area of the membrane (5) on the area of the face (101 ) of the prism (1 ), and the flow-through member (42) is provided with a window (422), thereby forming in the measuring cell (4) after assembling the apparatus at least one measuring chamber (411 ) and gastight flow-through chamber (421 ) separated from one another by the measured membrane (5), wherein each measuring chamber (411 ) is arranged above at least one detection spot (12), and the pressure module (43), which closes the flow-through chamber (421 ), is provided with an inlet channel (431 ) and an outlet channel (432), which are situated above the peripheral regions of the flow-through chamber (421 ).

2. An apparatus of claim 1 , characterised in that the detection member (41 ) is formed in the shape of a frame provided with cuts (422), the number of which corresponds to the number of the measuring spots (12), wherein each of the measuring chambers (411 ) is separated from the flow-through chamber (421 ) by a different membrane (5).

3. A method for measuring permeability of membranes using the apparatus of claim 1 or 2, characterised in that after assembling the measuring cell, when the membrane is inserted between the detection member, the measuring chamber of which is filled with the chosen liquid, and the flow-through member, through flow through chamber of which a fluid medium with a different concentration of the dissolved substances than that of the liquid in the measuring chamber flows, the time concentration response of the substance diffusing through the membrane, based on which the value of the permeability of the membrane is subsequently determined, is measured using monitoring and assessment of the changes in signals of electromagnetic beams passing through the body of the prism, incident on the detection spots of the metal coating, and captured in the sensor.