Systems and associated methods for producing mixtures of fluids in microfluidic channels, particularly for the formulation of liposome-based drugs through the alternating injection of two liquid phases - Patents.com

Abstract

The invention relates to a system (10) for producing a mixture of fluids, comprising a pressure source (11), a pressure regulator (12), a first container (16.1) containing at least a first fluid (15.1) and a second container (16.2) containing a second fluid (15.2), a microfluidic mixer (20) and a control unit (40) which controls the pressure level of the first fluid (15.1) and the pressure level of the second fluid (15.2) and controls the opening and closing of a first valve (23.1) and a second valve (23.2) to sequentially inject the first fluid and the second fluid into the microfluidic mixer (20) and generate a profile of fringes (F1, F2) of the first fluid (15.1) and the second fluid (15.2) in a common outlet channel (34).

Description

[Technical field]

[0001] The present invention relates to a system and associated methods for producing mixtures of fluids in microfluidic channels, in particular for the formulation of liposome-based drugs through the alternating injection of two liquid phases. [Background technology]

[0002] Several systems for mixing organic solvent-lipid solutions and aqueous solutions in microfluidic circuits have been described in the state-of-the-art literature. Existing methods most commonly use passive mixers, such as hydrodynamic focusing or chaotic mixers, whose mixing performance is sensitive to the fluctuations in the flow rate of the mixing phases. This limits the scale-up of liposomal pharmaceuticals for mass production. Moreover, existing devices suffer from aggregation issues, which can foul the microchannels. Summary of the Invention

[0003] The object of the present invention is to effectively remedy these drawbacks by proposing a system for producing a mixture of fluids, the system comprising: A pressure source; a pressure regulator to which the pressure source is connected; At least a first container containing a first fluid and a second container containing a second fluid, wherein pressurization of the first fluid and the second fluid is controlled by the pressure regulator; a microfluidic mixer comprising at least a first inlet orifice and a second inlet orifice associated with at least a first valve and a second valve; the first container is connected to the first inlet orifice via the first valve, and the second container is connected to the second inlet orifice via the second valve; the microfluidic mixer further comprises at least a first microfluidic conduit and a second microfluidic conduit, the first microfluidic inlet conduit in fluid communication with the first inlet orifice and the second microfluidic inlet conduit in fluid communication with the second inlet orifice, the first microfluidic inlet conduit and the second microfluidic inlet conduit intersecting at a non-zero angle at an intersection opening into at least one common outlet channel; The system further comprises a control unit for controlling a pressure level of the first fluid in the first container and a pressure level of the second fluid in the second container, as well as opening and closing of the first valve and the second valve, to perform sequential injection of the first fluid and the second fluid into the microfluidic mixer so as to generate a fringe profile of the first fluid and the second fluid in the common outlet channel.

[0004] The present invention allows the production of rapid homogeneous mixtures, which are necessary especially for the formulation of small size nanoparticles below 100 nm. Due to the absence of static liquid-liquid interfaces and ultrafast pressure fluctuations, the present invention dramatically reduces nanoparticle aggregation and accumulation in the microfluidic channels. It also reduces the formation of fluidic instabilities by avoiding fouling of the microfluidic circuits. Furthermore, the present invention allows the dilution time of the organic-lipid phase in the aqueous phase to be reduced to less than 1 ms by optimizing the generation of alternating fringes of the organic-lipid phase and the aqueous phase in a pulsed manner, thereby allowing the formulation of liposomes with high size monodispersity. Preferably, the polydispersity index PD1 is less than 0.1.

[0005] According to one embodiment of the invention, the control unit is configured to generate a fringe profile with alternating fringes of the first fluid and fringes of the second fluid.

[0006] According to one embodiment of the invention, the control unit is configured such that the fringe of the first fluid is narrower than the fringe of the second fluid.

[0007] According to one embodiment of the invention, the ratio of the volume of the fringe of the second fluid divided by the volume of the fringe of the first fluid is between 2 and 20, in particular between 8 and 15, preferably between 9 and 11.

[0008] According to one embodiment of the invention, the first container contains a solution of lipids and / or polymers diluted in an organic solvent corresponding to the first fluid, and the second container contains an aqueous solution corresponding to the second fluid.

[0009] According to one embodiment of the present invention, the first valve and the second valve are solenoid valves with a small dead volume, in particular less than 5 μL, and a high response time, in particular less than 5 ms.

[0010] According to one embodiment of the invention, the common outlet channel is extended by a channel having a cross-sectional width greater than a cross-sectional width of the common outlet channel.

[0011] According to one embodiment of the invention, the system comprises an exchangeable flow sensor for measuring the fluid flow rate at the outlet of the microfluidic mixer.

[0012] According to one embodiment of the invention, the first and second microfluidic inlet conduits intersect at an angle of less than 90 degrees.

[0013] According to one embodiment of the invention, the first and second microfluidic inlet conduits each have a cross-sectional height between 150 μm and 300 μm, preferably on the order of 200 μm, and a width between 150 μm and 300 μm, preferably on the order of 200 μm.

[0014] According to an embodiment of the invention, the pressure level of the first fluid in the first container and the pressure level of the second fluid in the second container are each between 0 and 8000 mbar.

[0015] According to one embodiment of the invention, the injection frequency, which is equal to the reciprocal of the sum of the injection duration of the first fluid and the injection duration of the second fluid, is between 0.1 Hz and 200 Hz, preferably between 10 Hz and 100 Hz.

[0016] The invention will be better understood and other features and advantages will appear on reading the following detailed description, which includes embodiments given by way of illustration and with reference to the accompanying drawings, presented as non-limiting examples, which will serve to complete the understanding of the invention and the description of its implementations, and ultimately contribute to its definition. [Brief description of the drawings]

[0017] [Figure 1] FIG. 1 is a schematic diagram of a system for producing a mixture of two fluids according to the present invention.

[0018] [Diagram 2] FIG. 2 is a cross-sectional view of a microfluidic conduit for use in a system according to the present invention.

[0019] [Diagram 3] FIG. 3 illustrates a method for generating alternating fringes in a static microfluidic mixer according to the present invention.

[0020] [Figure 4] FIG. 4 shows the flow rates of the first and second fluids as a function of time during the injection phase of alternating fringes of the first and second fluids. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0021] It should be noted that structural and / or functional elements that are common to the different embodiments are given the same reference numerals and therefore, unless otherwise specified, such elements have the same structural, dimensional and material characteristics.

[0022] FIG. 1 shows a system 10 for producing a fluid mixture comprising a pressure source 11 which may be in the form of, for example, an air compressor or a bottle containing pressurized gas such as air or nitrogen.

[0023] The pressure source 11 is connected via a conduit 13 to a pressure regulator 12 for controlling the pressurization of the first and second fluids 15.1 and 15.2 contained in a first and second container 16.1 and 16.2, respectively. The pressure regulator 12 is connected to the first container 16.1 via a conduit 17. The pressure regulator 12 is connected to the second container 16.2 via a conduit 18. The pressure regulator 12 may be a PID (Proportional, Derivative, Integral) type regulator based on the use of a sensitive piezoelectric sensor.

[0024] Advantageously, the first container 16.1 contains a solution of lipids and / or polymers diluted in an organic solvent, which corresponds to the first fluid 15.1 (organic phase), and the second container 16.2 contains an aqueous solution, which corresponds to the second fluid 15.2 (aqueous phase).

[0025] The microfluidic mixer 20 comprises a first inlet orifice 21.1 to which a first container 16.1 is connected via a first valve 23.1 and a second inlet orifice 21.2 to which a second container 16.2 is connected via a second valve 23.2. The first valve 23.1 and the second valve 23.2 are preferably solenoid valves (also called electromagnetic valves) with a small dead volume, in particular less than 5 μL, and a high response time, in particular less than 5 ms.

[0026] To this end, the first container 16.1 is in fluid communication with an inlet of a first valve 23.1 via a conduit 25. The outlet of the first valve 23.1 is in fluid communication with the first inlet orifice 21.1 via a conduit 26. The second container 16.2 is in fluid communication with an inlet of a second valve 23.2 via a conduit 27. The outlet of the second valve 23.2 is in fluid communication with the first inlet orifice 21.1 via a conduit 28.

[0027] The static microfluidic mixer 20 further comprises a first microfluidic inlet conduit 30.1 in fluid communication with the first inlet orifice 21.1 and a second microfluidic inlet conduit 30.2 in fluid communication with the second inlet orifice 21.2. As shown in Fig. 2, the first microfluidic inlet conduit 30.1 and the second microfluidic inlet conduit 30.2 each have a cross-sectional height h between 150 μm and 300 μm, preferably on the order of 200 μm, and a width l between 150 μm and 300 μm, preferably on the order of 200 μm. "On the order of" means a variation of plus or minus 10% relative to the indicated value.

[0028] The first and second microfluidic inlet conduits 30.1, 30.2 intersect at a relatively non-zero angle at an intersection 33 which opens into a common outlet flow channel 34. Advantageously, the first and second microfluidic inlet conduits 30.1, 30.2 intersect, i.e., cut off from each other, at an angle of less than 90 degrees. The outlet orifices of the first and second conduits 30.1, 30.2 open at the intersection 33.

[0029] The common outlet channel 34 is extended by a channel 35 having a cross-sectional width greater than the cross-sectional width of the common outlet channel 34. According to an exemplary embodiment, the common outlet channel 34 may have a width l approximately equal to twice the width l of the microfluidic inlet conduits 30.1, 30.2, i.e. of the order of 400 μm. The common outlet channel 34 is extended by another channel 35 having a cross-sectional width l of between 1 and 5 mm, preferably of the order of 3 mm. The heights h of the conduits 30.1, 30.2, 34, 35 may be identical to each other. Alternatively, the height h may be different for each of the conduits 30.1, 30.2, 34, 35. The height h may also be variable within the same microfluidic conduit / channel 30.1, 30.2, 34, 35.

[0030] For measuring the flow rate of the fluid at the outlet of the microfluidic mixer 20, an exchangeable flow sensor 37 is provided.

[0031] The control unit 40 is capable of controlling the pressure level of the first fluid 15.1 in the first container 16.1 and the pressure level of the second fluid 15.2 in the second container 16.2, as well as the opening and closing of the first valve 23.1 and the second valve 23.2 for carrying out the sequential injection of the first fluid 15.1 and the second fluid 15.2 in the microfluidic mixer 20, so as to generate fringe profiles F1, F2 of the first fluid 15.1 and the second fluid 15.2 in the common outlet channel 34. The pressure levels of the first fluid 15.1 and the second fluid 15.2 are controlled by the control unit 40 via the pressure regulator 12. The control unit 40 is electrically connected to the valves 23.1, 23.2 and controls their opening and closing.

[0032] Figure 3 shows how to generate alternating fringes F1, F2 in the common outlet channel 34 of the microfluidic mixer 20 and an experimental model made with phosphors diluted in ethanol (corresponding to the first fluid 15.1) and water (corresponding to the second fluid 15.2). In the figure, the phosphor diluted in ethanol appears darker than the water. The mixing system 10 multiplexes the liquid-liquid interfaces of the organic and aqueous phases, facilitating mixing between the two liquids.

[0033] It is observed that the control unit 40 is configured to generate a fringe profile F1, F2 in which the fringes F1, F2 of the first fluid 15.1 alternate with the fringes F1, F2 of the second fluid 15.2, i.e. a fringe F1 of the first fluid 15.1 (phosphor+ethanol) is followed by a fringe F2 of the second fluid 15.2 (water), which is followed by a fringe F1 of the first fluid 15.1, etc. The fringes F1, F2 correspond to the amount of fluid passing through the valves 23.1, 23.2 during their opening time. By adjusting the opening time of the valves 23.1, 23.2 and the pressure levels of the corresponding fluids, it is possible to adjust the width of the fringes F1, F2.

[0034] Advantageously, the control unit 40 is configured such that the fringe F1 of the first fluid 15.1 is narrower than the fringe F2 of the second fluid 15.2 to facilitate dilution of the first fluid 15.1 in the second fluid 15.2.

[0035] 4 shows the flow rates D of the first and second fluids expressed in microliters / second as a function of time (seconds) during the phase of injection of alternating fringes of the first and second fluids. Preferably, the ratio of the volume V2 of the fringe F2 of the second fluid divided by the volume V1 of the fringe F1 of the first fluid 15.1 is between 2 and 20, in particular between 8 and 15, preferably between 9 and 11.

[0036] The injection frequency f, which is equal to the reciprocal of the sum of the injection duration dt1 of the first fluid and the injection duration dt2 of the second fluid, is between 0.1 and 200 Hz, preferably between 10 Hz and 100 Hz.

[0037] The injection duration dt1 of the first fluid and the injection duration dt2 of the second fluid correspond to the opening times of the first valve 23.1 and the second valve 23.2, respectively. When the injection flow rates of the first and second fluids are equal, the ratio dt2 / dt1 is equal to the ratio V2 / V1.

[0038] The pressure level of the first fluid 15.1 in the first container 16.1 and the pressure level of the second fluid 15.2 in the second container 16.2 are each between 0 and 8000 mbar, in particular between 500 mbar and 7500 mbar.

[0039] 3, the intensity of fringe F1 decreases rapidly with distance from the outlet of channel 34. A graph showing pixel intensity level as a function of distance from the outlet of channel 34 indicates that the organic phase is diluted after passing just over 5 mm within channel 35.

[0040] The present invention allows the dilution time of the organic phase of lipids (and / or polymers) in the aqueous phase to be reduced to less than 1 ms. The present invention further allows the formulation of liposomes with high size monodispersity (PDI < 0.1). The present invention also allows the nucleation rate of lipid nanoparticles to be optimized.

[0041] These data (fluid pressure level, fluid injection frequency and the corresponding duty cycle) are provided as input parameters to the control unit 40. For this purpose, a human machine interface such as a keyboard, a touch screen or other device adapted to the application can be used.

[0042] The control unit 40 may include memory for storing software instructions for controlling the pressure regulator 12 and the valves in accordance with received input parameters. The control unit 40 may take the form of, for example, an application specific computer or microcontroller.

[0043] By precisely controlling the time and amplitude of the pulse injection in this way, the fringe profiles F1, F2 of the organic solvent-lipid and / or polymer solution and the aqueous solution are determined, and therefore the final size of the liposomes.

[0044] The present invention also relates to a method for producing a mixture of fluids implemented by the system 10.

[0045] Alternatively, system 10 does not include conduit 35 and is comprised only of conduit 34 .

[0046] Alternatively, the system 10 may be used to generate a gas mixture.

[0047] Alternatively, the microfluidic mixer 20 may comprise more than two inlet orifices 21.1, 21.2, in particular N inlet orifices associated with N microfluidic conduits and N valves, N being an integer. The fringe profile in the common outlet channel 34 may be a combination of the N fluids injected one after the other, or according to any possible combination type of the fluids present. Also, the number of vessels may be greater than two.

[0048] Alternatively, multiple common outlet channels may be used, each corresponding to a particular fringe profile.

[0049] Alternatively, a 3 / 2 splitter, rotary valve, or other means for alternately injecting fluids into a common outlet flow path may be used.

[0050] Of course, different features, variants and / or embodiments of the invention can be associated with each other in various combinations, unless they are mutually exclusive or incompatible with each other.

[0051] Furthermore, the present invention is not limited to the above-described embodiments, which are provided only as examples, and which encompass various modifications, alternative forms and other variations that may occur to those skilled in the art in the context of the present invention, in particular any combination of the various operating modes described above may be taken separately or in combination.

Claims

1. 1. A system for producing a mixture of fluids, comprising: a pressure source; a pressure regulator to which the pressure source is connected; at least a first container containing a first fluid and a second container containing a second fluid, wherein pressurization of the first fluid and the second fluid is controlled by the pressure regulator; a microfluidic mixer including at least a first inlet orifice and a second inlet orifice associated with at least a first valve and a second valve; the first container is connected to the first inlet orifice via the first valve, and the second container is connected to the second inlet orifice via the second valve; the microfluidic mixer further comprises at least a first microfluidic conduit and a second microfluidic conduit, the first microfluidic inlet conduit in fluid communication with the first inlet orifice and the second microfluidic inlet conduit in fluid communication with the second inlet orifice, the first microfluidic inlet conduit and the second microfluidic inlet conduit intersecting at a non-zero angle at an intersection that opens into at least one common outlet channel; The system further comprises a control unit that controls a pressure level of the first fluid in the first container and a pressure level of the second fluid in the second container, and controls opening and closing of the first valve and the second valve to perform sequential injection of the first fluid and the second fluid into the microfluidic mixer so as to generate fringe profiles of the first fluid and the second fluid in the common outlet channel.

2. 2. The system of claim 1, wherein the control unit is configured to generate a fringe profile that alternates between fringes of the first fluid and fringes of the second fluid.

3. 3. The system of claim 2, wherein the control unit is configured to cause the fringe of the first fluid to be narrower than the fringe of the second fluid.

4. 4. The system according to claim 3, characterized in that the ratio of the volume of the fringes of the second fluid divided by the volume of the fringes of the first fluid is between 2 and 20, in particular between 8 and 15, preferably between 9 and 11.

5. 4. The system according to claim 1, wherein the first container contains a solution of lipids and / or polymers diluted in an organic solvent corresponding to the first fluid, and the second container contains an aqueous solution corresponding to the second fluid.

6. 5. The system according to claim 1, wherein the first valve and the second valve are solenoid valves having a low dead volume, in particular less than 5 μL, and a high response time, in particular less than 5 ms.

7. 5. A system according to claim 1, wherein the common outlet channel is extended by a channel having a cross-sectional width greater than the cross-sectional width of the common outlet channel.

8. 5. The system according to claim 1, further comprising an exchangeable flow sensor for measuring the fluid flow rate at the outlet of the microfluidic mixer.

9. 5. The system of claim 1, wherein the first microfluidic inlet conduit and the second microfluidic inlet conduit intersect at an angle of less than 90 degrees.

10. 5. A system according to any one of claims 1 to 4, characterized in that the first and second microfluidic inlet conduits each have a cross-sectional height of between 150 μm and 300 μm, preferably of the order of 200 μm, and a width of between 150 μm and 300 μm, preferably of the order of 200 μm.

11. 5. The system according to claim 1, wherein the pressure level of the first fluid in the first container and the pressure level of the second fluid in the second container are each between 0 and 8000 mbar.

12. 5. The system according to claim 1, wherein the injection frequency, which is equal to the reciprocal of the sum of the injection duration of the first fluid and the injection duration of the second fluid, is between 0.1 and 200 Hz, preferably between 10 Hz and 100 Hz.

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