We report an approach for the online coupling of digital microfluidics (DMF) with mass spectrometry (MS) using a chip-integrated microspray hole (μSH). The technique uses an adapted electrostatic spray ionization (ESTASI) method to spray a portion of a sample droplet through a microhole in the cover plate, allowing its chemical content to be analyzed by MS. This eliminates the need for chip disassembly or the introduction of capillary emitters for MS analysis, as required by state-of-the-art. For the first time, this allows the essential advantage of a DMF devicefree droplet movementto be retained during MS analysis. The broad applicability of the developed seamless coupling of DMF and mass spectrometry was successfully applied to the study of various on-chip organic syntheses as well as protein and peptide analysis. In the case of a Hantzsch synthesis, we were able to show that the method is very well suited for monitoring even rapid chemical reactions that are completed in a few seconds. In addition, the strength of the low resource consumption in such on-chip microsyntheses was demonstrated by the example of enzymatic brominations, for which only a minute amount of a special haloperoxidase is required in the droplet. The unique selling point of this approach is that the analyzed droplet remains completely movable after the MS measurement and is available for subsequent on-DMF chip processes. This is illustrated here for the example of MS analysis of the starting materials in the corresponding droplets before they are combined to investigate the reaction progress by DMF-MS further. This technology enables the ongoing and almost unlimited tracking of multistep chemical processes in a DMF chip and offers exciting prospects for transforming digital microfluidics into automated synthesis platforms.

In this work, we introduce an approach to merge droplet microfluidics with a HPLC/MS functionality on a single chip to analyze the contents of individual droplets. This is achieved by...

Improving the performance of chemical transformations catalysed by microbial biocatalysts requires a deep understanding of cellular processes. While the cellular heterogeneity of cellular characteristics, such as the concentration of high abundant cellular content, is well studied, little is known about the reactivity of individual cells and its impact on the chemical identity, quantity, and purity of excreted products. Biocatalytic transformations were monitored chemically specific and quantifiable at the single-cell level by integrating droplet microfluidics, cell imaging, and mass spectrometry. Product formation rates for individual Saccharomyces cerevisiae cells were obtained by i) incubating nanolitresized droplets for product accumulation in microfluidic devices, ii) an imaging setup to determine the number of cells in the droplets, and iii) electrospray ionisation mass spectrometry for reading the chemical contents of individual droplets. These findings now enable the study of whole-cell biocatalysis at single-cell resolution.

Microfluidic double-emulsion droplets allow the realization and study of biphasic chemical processes such as chemical reactions or extractions on the nanoliter scale. Double emulsions of the rare type (o1/w/o2) are used here to realize a lipase-catalyzed reaction in the non-polar phase. The surrounding aqueous phase induces the transfer of the hydrophilic product from the core oil phase, allowing on-the-fly MS analysis in single double droplets. A microfluidic two-step emulsification process is developed to generate the (o1/w/o2) double-emulsion droplets. In this first example of microfluidic double-emulsion MS coupling, we show in proof-of-concept experiments that the chemical composition of the water layer can be read online using ESI–MS. Double-emulsion droplets were further employed as two-phase micro-reactors for the hydrolysis of the lipophilic ester p-nitrophenyl palmitate catalyzed by the Candida antarctica lipase B (CalB). Finally, the formation of the hydrophilic reaction product p-nitrophenol within the double-emulsion droplet micro-reactors is verified by subjecting the double-emulsion droplets to online ESI–MS analysis. Graphical abstract

We report a novel approach for surface-enhanced Raman spectroscopy (SERS) detection in digital microfluidics (DMF). This is made possible by a microspray hole (μSH) that uses an electrostatic spray (ESTAS) for sample transfer from inside the chip to an external SERS substrate. To realize this, a new ESTAS-compatible stationary SERS substrate was developed and characterized for sensitive and reproducible SERS measurements. In a proof-of-concept study, we successfully applied the approach to detect various analyte molecules using the DMF chip and achieved micro-molar detection limits. Moreover, this technique was exemplarily employed to study an organic reaction occurring in the DMF device, providing vibrational spectroscopic data.