MICROFLUIDIC SYSTEM AND METHOD FOR CONTINUOUS MONITORING OF METABOLITES AND/OR PROPERTIES OF BIOFLUIDS

Abstract

The present invention refers to a microfluidic system based on passive capillary valves and pumps, that allows a discontinuous and autonomous measurement process for extensive periods of time. The microfluidic system comprises: at least one measuring chamber, at least one inlet for the input of a biofluid, a microfluidic intake channel fluidly communicating the inlet with the measuring chamber, at least one sensor suitable for measuring a parameter of an analyte of a biofluid. A passive fluid pump is fluidly communicated with the measuring chamber, and it is adapted to generate a capillary pressure greater than the biofluid generation pressure. A retention valve is interposed between the measuring chamber and the fluid pump, and the retention passive valve is configured to stop flow of biofluid for a certain period of time, when the measuring chamber if filled with biofluid. The invention provides a robust device, capable of collecting and conveying a biofluid, preferably sweat, for repetitive and electrochemical measurements.

Claims

1. Microfluidic system for continuous monitoring of metabolites and/or properties of biofluids, the system comprising: one measuring chamber, at least one inlet for the input of a biofluid, a microfluidic intake channel fluidly communicating the inlet with the measuring chamber, at least one sensor suitable for measuring a parameter of an analyte of a biofluid, and arranged to measure the parameter of a biofluid contained in the measuring chamber, a passive fluid pump fluidly communicated with the measuring chamber, and adapted to generate a capillary pressure greater than the biofluid generation pressure, a retention valve interposed between the measuring chamber and the fluid pump, wherein the retention passive valve is configured to stop flow of biofluid for a certain period of time, when the measuring chamber if filled with biofluid.

2. The system according to claim 1, further comprising: a secondary microfluidic channel connected to the microfluidic intake channel and connected to the atmosphere, and a stop passive valve interposed at the secondary microfluidic channel, and adapted to impede fluid flow out of the secondary microfluidic channel towards the atmosphere.

3. The system according to claim 2, wherein the stop valve is configured such that its bursting pressure is greater than the maximum biofluid generation pressure.

4. The system according to claim 2, wherein the bursting pressure of the retention passive valve is lower than the bursting pressure of the stop passive valve.

5. The system according to claim 1, wherein the retention valve is configured to feature a bursting pressure that retain the measuring chamber filled with biofluid and stop biofluid flow for a period within the range 1 minute to 5 hours.

6. The system according to claim 1, wherein the retention passive valve is configured as a sudden enlargement of the cross-section area of the microfluidic intake channel.

7. The system according to claim 1, wherein the retention passive valve is configured as a chemical modification of the surface of the microfluidic intake channel.

8. The system according to claim 1, wherein the fluid pump is a micro- machined capillary pump capable of forcing fluid circulation by capillary action.

9. The system according to claim 1, wherein the fluid pump is a porous material pump.

10. The system according to claim 1, further comprising a cycle detector adapted to detect when the measuring chamber is filled with biofluid, and to monitor changes on cycle frequency.

11. The system according to claim 10, wherein the cycle detector comprises two electrodes arranged to measure an electric parameter at the microfluidic circuit between the inlet and the fluid pump inlet, so as to detect whether there is a biofluid or air in the microfluidic circuit.

12. The system according to claim 1, wherein the inlet is adapted to collect sweat from the skin of a subject.

13. The system according to claim 1, adapted to monitor sweat metabolites and/or properties, and wherein the sweat properties include conductivity and/or sweat rate and/or sweat pH, and/or ions.

14. The system according to claim 1, further comprising an electronic device electrically communicated with the sensor, and adapted for processing data generated by the sensor.

15. A wearable device for sweat monitoring incorporating the system according to claim 1, and wherein the system is configured as a disposable cartridge detachably coupled with the wearable device.

16. The system according to claim 4, wherein the bursting pressure of the retention valve is within the range 2.4-6 kPa.

17. The system according to claim 5, wherein the retention valve retains the measuring chamber filled with biofluid and stop biofluid flow for a period within the range the range 1 to 5 minutes.

18. The system according to claim 8, wherein the passive fluid pump is adapted to generate a capillary pressure higher than 6 kPa.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0059] Preferred embodiments of the invention are henceforth described with reference to the accompanying drawings, wherein:

[0060] FIG. 1.—shows schematically in a top plan view, the structure of a geometrical passive valve according to the prior-art (height constant) where all relevant parameters are noted.

[0061] FIG. 2.—shows schematically a first embodiment of the microfluidic system of the invention at five sequential stages (a-e) of the measurement process. Q is biofluid flow. In the microfluidic system, dark areas represent air, and striped areas represent liquid flow.

[0062] FIG. 3.—shows schematically the chronological steps of the system of FIG. 2, with a graphical representation of the theoretical fluid flow evolution over time.

[0063] FIG. 4.—shows schematically the working principle and the different steps of a second embodiment of the microfluidic system, that includes an air vent and a secondary channel.

[0064] FIG. 5.—shows a practical implementation of the embodiment of FIG. 2, wherein Figure A shows the entire system, Figure B is an enlarged detail of the measuring chamber and retention passive valve including three sequential stages (I, II, III) of the process. FIG. C shows the measuring chamber of the second embodiment of FIG. 4 also including three sequential stages (I, II, III) of the process, and areas for detecting air bubble formation. In the figure, dark areas represent fluid flow.

[0065] FIG. 6.—shows a wearable device integrating the proposed microfluidic system, wherein Figure A is an exploded view; and Figure B is a schematic representation of the arrangement of layers to implement the system.

[0066] FIG. 7.—shows a sequece of captions (A-F) of different stages of the cycle carried out in a practial implementation of the microfluidic device.

[0067] FIG. 8.—shows more captions (A-D) of different stages of the cycle carried out in a practial implementation of the microfluidic device.

PREFERRED EMBODIMENT OF THE INVENTION

[0068] FIG. 2 shows a first embodiment of the microfluidic system of the invention at five sequential stages (a-e) of the measurement process. The microfluidic system comprises: [0069] one measuring chamber (2), [0070] one inlet (1) for the input of a biofluid, for example adapted to collect sweat from the skin of a subject, [0071] a microfluidic intake channel (8) fluidly communicating the inlet (1) with the measuring chamber (2), [0072] a passive fluid pump (4) fluidly communicated with the measuring chamber (2), and adapted to generate a capillary pressure greater than the biofluid generation pressure, [0073] at least one sensor (not shown) suitable for measuring a parameter of an analyte of a biofluid, and arranged to measure the parameter of a biofluid contained in the measuring chamber. Preferably, one sensor is enclosed inside the measuring chamber, [0074] a retention passive valve (3) interposed between the measuring chamber (2) and the fluid pump (4).

[0075] The retention passive valve (3), preferably is of the geometrical type, and it is configured to stop flow of biofluid for a certain period of time (preferably 1 to 5 minutes for sweat applications), when the measuring chamber is filled with biofluid.

[0076] The passive pump (4) drains the fluid at a high flow rate, and it only works when the retention valve (3) is surpassed. It could be a naturally porous material (like paper) or a microfabricated pillar-like capillary pump. The relevant characteristic of the passive pump, is that its wicking pressure provides a flow rate greater than the one at the inlet of the microfluidic system. For sweat applications, pump fluid generation rate varies between 1 to 20 nL/min.Math.gland, which can be traduced (accounting 100 glands/cm.sup.2 and a collection area of 0.5 cm.sup.2) to a 50-1000 nL/min range.

[0077] FIG. 3 shows the chronological steps of the operation of the embodiment of FIG. 2 with a graphical representation of the theoretical fluid flow evolution over time, which is as follows: [0078] Filling, (stage A in FIGS. 2 & 3). Fluid enters through the inlet (1) and flows through the microfluidic system at the rate of fluid generation. [0079] Measurement, (stage B in FIGS. 2 & 3). When the fluid front arrives at the retention passive valve (3), it is stopped for a few minutes, because the dominant pressure governing fluid flow is lower than the bursting pressure (BP) of the retention valve (3). With no fluid flow and with the measuring chamber (2) filled, then measurements are taken while fluid pressure (P3) at the fluid-air interface increases due to the fluid generation pressure at the inlet. [0080] Bursting, (stage C in FIGS. 2 & 3). After a certain period of time, the retention passive valve (3) bursts due to the increased fluid pressure, and fluid flows through it (at a high flow rate (note first flow peak in FIG. 3), and reaches the passive pump (4) flowing through the pump. [0081] Next measurement, (stage D in FIGS. 2 & 3). Then, after a few seconds due to the high flow rate forced by the pump (4), the flow disconnects, that is, the fluid flow is interrupted and an air-fluid interface is regenerated at the passive valve (3), and the fluid is stopped again and measurements in the measuring chamber are taken again. [0082] Bursting, (Stage E in FIGS. 2 & 3). After several minutes, bursting occurs again (second flow peak), repeating the cycle described above.

[0083] In FIG. 3, P.sub.3 is the fluid pressure at position 3 while P.sub.fluid refers to the dominant pressure governing fluid flow, BP is bursting pressure of the retention (or delay) passive valve (3), and CP is capillary pressure of the pump (4).

[0084] FIG. 4 shows a second embodiment in which the following elements are incorporated to the first embodiment of FIGS. 2 and 3: [0085] an air inlet (7) open to the atmosphere, allowing the pass of air into the intake channel, [0086] a secondary microfluidic channel (5) connected to the microfluidic intake channel (8) and connected to the atmosphere by the air inlet (7) allowing the entrance of air. This secondary channel (5) is placed before the measuring chamber (2), and it is connected with the intake channel (8) at a microfluidic junction (T-junction). [0087] a stop passive valve (6) interposed at the secondary microfluidic channel (5), and adapted to impede the fluid to flow out of the secondary microfluidic channel (5) towards the atmosphere. For that, the stop valve (6) is configured to feature a bursting pressure (BP) greater than any other pressure of the microfluidic system to avoid being surpassed, specially its bursting pressure is greater than the maximum biofluid generation pressure.

[0088] In the embodiment of FIG. 4, the secondary channel (5) opens to the atmosphere allows the creation of air bubbles in the middle of the channel in a passive manner. The entrance of air comes from the air vent (7) when the fluid in the main intake channel (8) is drained due to the high pressure generated by the passive pump (4), creating an air bubble at the microfluidic junction of the channels (5,8). As the pump (4) is still connected to the fluid, the air bubble travels along the microfluidic main channel (8) (embodiment 1), restoring the functionality of the retention passive valve (3) when the interface arrives there. This way, a similar cycle as the one described for the embodiment of FIGS. 2 and 3, is created.

[0089] FIG. 4 additionally represents the scheme of the working principle and the different stages of the microfluidic system proposed in this second embodiment. At first (Stage A), fluid coming from the inlet (1) fills the measuring chamber (2) at a given flow rate, while the secondary channel (5) is also filled but up to the stop valve (6), where it is stopped. When fluid arrives at the retention passive valve (3), it is temporarily stopped (Stage B). When the valve (3) bursts (Stage C), the passive pump (4) is connected to the fluid (fluid reaches the pump) and drains it rapidly, generating an air bubble at the microfluidic junction (Stage D).

[0090] As the passive pump (4) is still connected to the remaining fluid, it keeps draining making the air bubble to travel along the measuring chamber (2) until the retention passive valve (3) (Stage E). Once there, the retention passive valve (3) has again an air-fluid interface and stops the flow as described previously for a few minutes (Stage F).

[0091] In FIG. 4, P.sub.3, P.sub.5, P.sub.6 are the fluid pressures at each location of the microfluidic circuit, whereas P.sub.fluid refers to the dominant pressure governing fluid flow.

[0092] The capillary pump must provide a capillary pressure greater than the maximum biofluid generation pressure. In the case of sweat applications, it has been noted that the maximum secretory pressure is below 6 kPa. Therefore, capillary pressures of the papers/capillary pumps used should be designed to be at least higher than 6 kPa. In the case of the second embodiment (FIG. 4) the capillary pressure of the pump must be greater than inlet (in order to drain air to the junction), and below a maximum capillary pressure that avoids disconnection before air bubble travels to the retention valve.

[0093] In addition, the retention valve bursting pressure (BP) should be designed to be within the biofluid generation pressure. Preferably, the bursting pressure (BP) of the retention valve is within the range 2.4-6 kPa. This way, it will be able to burst eventually, but also stopping flow temporarily. The retention time will be a factor that would depend on the BP (proportionally), but subject to other system conditions. It should be remembered than once it is surpassed by the interface, the retention valve just behaves as a fluid resistor.

[0094] Furthermore, the stop valve bursting pressure should be designed to be greater than the maximum biofluid generation pressure. This way, the pressure barrier created will never be surpassed by the fluid pressure.

[0095] As it can be noted, all the above conditions are related to the biofluid generation pressure (secretory pressure for sweat glands, in the case of sweat). There is not a concrete relationship between the BP of the valves and the capillary pressure of the pump by design. They are conditioned by their relationship with the fluid generation pressure. This relationship generates the phenomenon that has been found, that is why, in FIGS. 3 & 4, flow rate was chosen to represent each state of the cycle, as it is the observable variable. However, we can define the pressures involved at each state of the cycle as described for FIGS. 3 & 4.

[0096] At first, fluid flow is directed by the biofluid pressure as it fills the measuring chamber. When fluid encounters the passive valves, the pressure at each valve is not enough to overcome the pressure barrier generated by their geometry (BP). However, as the BP of the retention (delay) valve is found within its range, eventually the liquid will burst (when P.sub.3 is greater than delay, BP.sub.delay) and when this occurs, the fluid connects with the passive pump and its great capillary pressure dominates now the flow rate that is increased in the microfluidic circuit.

[0097] As commented previously, the pressure mismatch between pump and inlet produces a disconnection of the fluid flow at the retention valve, regenerating the air-liquid interface there and stopping the flow, as the fluid pressure now is limited to the inlet (pump is not connected to fluid). Then, the cycle starts again. In the case of the second embodiment, pump disconnection is achieved by the air arrival to the delay valve, restoring its functionality as pressure barrier.

[0098] Preferably, the system of any of the two embodiments described above, further includes a cycle detector adapted to detect when the measuring chamber is filled with biofluid, and to monitor changes on measuring cycle frequency. This cycle detector can be embodied as two electrodes arranged to measure an electric parameter at the microfluidic circuit between the inlet and the fluid pump inlet, so as to detect whether there is a biofluid or air in the microfluidic circuit.

[0099] For example, for first embodiment of FIGS. 2 and 3, the cycle detector can be located preferably just after the retention passive valve (3), and in the case of the second embodiment of FIG. 4, it could also be located in the microfluidic junction or at the sensing chamber, as the air bubble would travel through them.

[0100] FIG. 5 shows in FIG. 5A a practical design of the embodiment 1 (FIGS. 2 & 3), and in FIG. 5B it shows (as an enlarged view of the measuring chamber) several amperometric sensors (9) (CE,WE,RE) arranged inside the measuring chamber (2) for a metabolite such as lactate, and the two electrodes (10) of the cycle detector arranged just downstream the retention passive valve (3), at the zone where fluid connects to the passive pump (4). In FIG. 5B a cycle to be detected is represented as the sequence I, III and III.

[0101] In FIG. 5C a similar device as the one in FIG. 5B is represented, but corresponding to the second embodiment (FIGS. 2 & 3), wherein the cycle detector is adapted to detect air bubble formation, such that the two electrodes (10) are arranged between the microfluidic “T” junction and the entrance of the measuring chamber (2).

[0102] Since the cycle detector monitors the electrical resistance between the electrodes, it is possible to know at a certain moment in which cycle state is the process. A high resistance would imply that there is air in that position, meaning, in a configuration such as FIG. 5B, that the liquid is stopped at the delay valve, and it is time to perform the amperometric measurement. In addition, cycle frequency can be calculated from the time difference between this resistance change. In another configuration, FIG. 5C, a similar system could be used to detect the presence of the air bubble generated at the junction, being able to extract also the cycle state and frequency.

[0103] Given a known system, variations on cycle frequency could be attributed solely to the biofluid generation rate, as the valve and pump parameters would be designed beforehand. This way, cycle frequency could be related to biofluid generation rate, a parameter of interest in sweat applications.

[0104] FIG. 6 shows a representation of a wereable device integrating the microfluidic system of the invention, wherein the system is configured as a disposable cartridge detachably coupled with the wearable device. In particular, the exploded view of FIG. 6A shows two layers: a first layer (11) including the microfluidic system with sensors, and a second layer (12) on top of the first one, incuding electronics for processing data generated by the sensors, and the two layers enclosed inside a two parts casing (13).

[0105] FIG. 6B shows an schematic representation of the two layers arrangement, wherein the bottom later (14) is an adhesive layer suitable to be adhered to the skin (15) of a subject, and having an opening (16) for sweat collection from the skin. The first layer (11) includes the microfluidic system with the structure defined above, and a second layer (12) that integrates the electronics and the sensors (9), and that together with the first layer (11) forms the measuring chamber (2).

[0106] Regarding fabrication, the microfluidic system could be constructed by assembling two or three layers. In a two-layer structure, the microfluidic channels would be fabricated on one by any convenient microfabrication procedure (soft lithography, injection moulding, . . . ) and closed by the second layer. In a three-layer structure, the microfluidic channels would be fabricated on the middle layer by any convenient microfabrication procedure (laser cutting, . . . ) and the other layers would close the microfluidic system.

[0107] Regarding fluid collection and stimulation, and more specifically for sweat dedicated applications, sweat generation could be induced by exercise or thermal action or stimulated iontophoretically either by pilocarpine or carbachol.

[0108] Regarding skin integration, the system described could add a double-sided adhesive layer (acrylic preferably) to conformally bond to skin and the microfluidic system with the convenient openings to allow collection of sweat generated. This adhesive layer could be part also of the microfluidic system as one of the layers described before.

Experimental Results

[0109] For the first experiments, soft lithography was used as fabrication procedure since it provides the needed resolution to fabricate the structures needed. The material used was PDMS, a common material in the field of microfluidics, whose transparency allowed to characterize fluid behaviour by simply visualizing a coloured fluid. Paper was used as passive pump due to its simplicity and easy to change properties by simply changing the type of paper. Two different papers have been used successfully, but the cycle period changed keeping the rest of conditions constant, showing the tunability of this parameter by design. Fluid is injected continuously at the entrance of the microfluidic system at a constant flow rate (200 nL/min) which is found within the range of perspiration rates.

[0110] FIG. 7 captions of the different stages of the cycle of the microfluidic device in the first embodiment. Fluid arrives at the retention valve (A) and, after some minutes stopped, it bursts (B) connecting to the paper (C). After some seconds, the fluidic connection start to disappear (D) until it eventually does (E). After several minutes, the retention valve bursts again (F) repeating the whole cycle over again.

[0111] FIG. 8 shows images of different stages of a cycle of the microfluidic device in the second embodiment. Fluid arrives at the retention valve and is stopped for several minutes (A). When it bursts, it connects with the paper (B). During the connection, the fluid that has gone into the side channel returns back, due to the wicking force of the paper, creating an air bubble in the main channel (C). Then, the fluidic connection with paper disappears (D) and the air bubble remains pinned in the main channel.

[0112] Table 1 shows characteristics of the passive valve tested once fabricated. As described previously, the bursting pressure (BP) of the stop passive valve (6) should be greater than the maximum sweat secretory pressure achieved. In particular, the BP of a tested passive valve are comprehended in Table 1:

TABLE-US-00001 TABLE 1 Description of the different parameters used for the passive valves (both retention valve and stop valve) tested wherein BP is calculated from Equation 1, considering h = 100 um, sweat surface tension of 0.072 N/m and a PDMS contact angle of 110°). Type of Valve Width (um) Betha (°) BP (Pa) Retention Passive Valve 75  90 2406 Stop Passive Valve 25 110 6235

[0113] Other preferred embodiments of the present invention are described in the appended dependent claims and the multiple combinations of those claims.