(73d) Specific Development of Ewod Lab-on-a-Chip for Chemical Processor

In recent years, lab on a chip (LOC) systems dedicated to biotechnological and chemical applications have attracted growing interest. Compared to traditional batch reaction, miniaturized systems allow the use of very small quantities of reagents while optimizing the reactions and producing the desired products faster and in greater yield and purity. In channel microfluidic devices, fluids are manipulated in a network of micron-dimension channels etched in a matrix of glass or polymer. An other approach is the digital microfluidic, where simple droplets are displaced on a two dimension chip. This droplet microfluidic system has several advantages, such as easy fluidic connection and multiplexing [1, 2].

Digital microfluidic coupled with EWOD appears very promising for LOC application, since all the fluidic functions can be achieved by electostatique actuation, simplifying integration of complex protocols, without pumps or any other moving parts [3]. After a brief presentation of the state of the art, this presentation will focus on validation of the platform that was developed in our lab. Different functionalities were demonstrated and characterized: fast droplet motion, reproducible on-chip droplet dispensing, or quick mixing? Chips design can be drastically simplified by using a processor architecture inspired from micro-electronic engineering using an assembly of elementary fluidic modules. Ease of integration of complex fluidic protocol is demonstrated by performing on chip serial basic micro fluidic function. To illustrate the ease-of-use for end users such as biologist, chips and compact associated instrumentation was performed. The system is now validated for biological application like PCR multiplex analysis and enzymatic DNA repair. Protocols performed in a few nanoliter droplets show good reactions yield and no cross contamination.

To increase the field of applications, we are currently working on a chemical processor [2]. As EWOD actuation allows moving and merging droplets containing minutes amount of reagents, micro-chemistry can be performed in droplets, which are used as microreactors. Most of the applications reported so far are performed in aqueous solutions, which are often not suitable for chemical reactions. Indeed, a chemical processor requires other media, such volatile organic solvent (VOS). Usually, to prevent aqueous solutions from evaporation, droplets are handled between a matrix of electrodes and a top layer (closed system), and surrounding with an oil layer. However, the use of organic media in a closed system or under oil would introduce others problems, such as complication in the fluidic connections, cross-contamination and oil/solvent miscibility. We therefore decided to use ILs on our chemical chip, assuming that these particular solvents could be used in open microsystems (without top layer) without evaporating.

ILs have recently attracted considerable attention. Some of them, the so-called Room Temperature Ionic Liquids (RTILs) are free-flowing liquids at room temperature. RTILs are increasingly used in chemical synthesis as solvents, electrolyte materials and in biological applications due to their remarkable properties such as high ionic conductivity, very low evaporation rate, large potentials range in electrochemistry [4]. Moreover, a unique subclass of ILs, task-specific ionic liquids (TSILs) have greatly widened the utilisation scope of ILs [5, 6]. Their functional groups, covalently linked to the cation and/or the anion, confer additional properties to these salts, therefore expanding their potential applications far beyond those of conventional ionic liquids. Moreover, TSILs and RTILs can be combined in solutions. Hence, supported synthesis in homogeneous solution can be achieved, which is a major advantage. Thus, the simplicity and flexibility of EWOD actuation combined with ILs properties, offer the opportunity to design powerful tools for high throughput combinatorial synthesis of organic compounds, protocol optimization or multi-step synthesis [7].

As only few data existed on ILs under EWOD actuation, we investigated their behaviour. Firstly, we observed that ILs are sensitive to the EWOD phenomena and their spreading on an single electrode was quantified and compared to the one of aqueous solution. Eleven ILs were moved on our chip and their dynamic behaviour were also investigated and compared qualitatively to the one of aqueous solution. ILs show lower speeds (1-10 mm/s) than DI water (120 mm/s) due to their high viscosities (50 to 300 times more than water), but a wider range of actuation is available for ILs motion. Indeed, the minimum potential Vmin (minimum potential required to move a droplet) is lower for ILs than for water, due to their smaller hysteresis and lower surface tension, whereas the maximum potential Vmax (no more gain in the EWOD force above this potential) is similar to DI water. Moreover, we demonstrated that the splitting of ILs droplets could be done on an open system under air, which was never made with aqueous solution. We also performed dispensing [8] of ILs in closed chip from a reservoir, but under an oil layer.

Next to the validation of these basic fluidic functions with ILs, as a feasibility study, a Grieco's reaction [9] was performed on our chip. Grieco's reaction is an easy, fast and total reaction leading to tetrahydroquinolines and involving four components: an aniline, an aldehyde, an olefin and an acid as a catalyst. We used a TSIL, with an aniline or a aldehyde grafted on it. On the one hand, by doing so, we benefit of outstanding chemical reaction in the same way as classic homogeneous organic chemistry, and on the other hand simplified purification as in supported chemistry. As it is a multi-component reaction, we easily obtained 8 different components. The validation of the droplet reaction was firstly performed by ESI/MS. Then, we developed a HPLC method with an internal reference, in order to investigate the matrix influence of different IL on the Grieco reaction at the microscopic scale. Main conclusion was that in ILs with an NTf2- as anion, a 100% conversion rate is reached in 40 minutes, whereas 10 minutes are enough at the macroscopic scale. The lower rate enhancement in droplet compared to batch is explained by the lack of agitation in droplets. Indeed, at room temperature, the viscosities of ILs are so high, that only the diffusion of molecules permitting the reaction's progress. However, by heating the droplet to 80°C, the reaction is fully performed in 10 minutes, due to the significant decrease of the viscosities of ILs and to the apparition of convection flow inside the droplet.

The ESI/MS and HPLC used previously require taking off the droplet from the chip. In order to get an on-line detection (directly on the chip), we used a two catena microchip to perform on line electrochemistry detection. The first catenary plays the role of the working electrode and the second is both the counter electrode and the reference electrode. To get an easy detection, we used nitrobenzaldehyde for Grieco's reaction, and have targeted the reduction of the nitro group in the desired final product.

Concerning micro-chemical application, we are currently working on new on-line detection, such Raman spectroscopy, and on the design of new chips with large number of reservoirs for handling a large number of chemical reagents, in order to create high-throughput parallel synthesis reactor.

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