Investigating and Quantifying Molecular Complexity Using Assembly Theory and Spectroscopy.

Current approaches to evaluate molecular complexity use algorithmic complexity, rooted in computer science, and thus are not experimentally measurable. Directly evaluating molecular complexity could be used to study directed vs undirected processes in the creation of molecules, with potential applications in drug discovery, the origin of life, and artificial life. Assembly theory has been developed to quantify the complexity of a molecule by finding the shortest path to construct the molecule from building blocks, revealing its molecular assembly index (MA). In this study, we present an approach to rapidly infer the MA of molecules from spectroscopic measurements. We demonstrate that the MA can be experimentally measured by using three independent techniques: nuclear magnetic resonance (NMR), tandem mass spectrometry (MS/MS), and infrared spectroscopy (IR). By identifying and analyzing the number of absorbances in IR spectra, carbon resonances in NMR, or molecular fragments in tandem MS, the MA of an unknown molecule can be reliably estimated. This represents the first experimentally quantifiable approach to determining molecular assembly. This paves the way to use experimental techniques to explore the evolution of complex molecules as well as a unique marker of where an evolutionary process has been operating.

A robotic prebiotic chemist probes long term reactions of complexifying mixtures.

To experimentally test hypotheses about the emergence of living systems from abiotic chemistry, researchers need to be able to run intelligent, automated, and long-term experiments to explore chemical space. Here we report a robotic prebiotic chemist equipped with an automatic sensor system designed for long-term chemical experiments exploring unconstrained multicomponent reactions, which can run autonomously over long periods. The system collects mass spectrometry data from over 10 experiments, with 60 to 150 algorithmically controlled cycles per experiment, running continuously for over 4 weeks. We show that the robot can discover the production of high complexity molecules from simple precursors, as well as deal with the vast amount of data produced by a recursive and unconstrained experiment. This approach represents what we believe to be a necessary step towards the design of new types of Origin of Life experiments that allow testable hypotheses for the emergence of life from prebiotic chemistry.

Identifying molecules as biosignatures with assembly theory and mass spectrometry.

The search for alien life is hard because we do not know what signatures are unique to life. We show why complex molecules found in high abundance are universal biosignatures and demonstrate the first intrinsic experimentally tractable measure of molecular complexity, called the molecular assembly index (MA). To do this we calculate the complexity of several million molecules and validate that their complexity can be experimentally determined by mass spectrometry. This approach allows us to identify molecular biosignatures from a set of diverse samples from around the world, outer space, and the laboratory, demonstrating it is possible to build a life detection experiment based on MA that could be deployed to extraterrestrial locations, and used as a complexity scale to quantify constraints needed to direct prebiotically plausible processes in the laboratory. Such an approach is vital for finding life elsewhere in the universe or creating de-novo life in the lab.

A programmable chemical computer with memory and pattern recognition.

Current computers are limited by the von Neumann bottleneck, which constrains the throughput between the processing unit and the memory. Chemical processes have the potential to scale beyond current computing architectures as the processing unit and memory reside in the same space, performing computations through chemical reactions, yet their lack of programmability limits them. Herein, we present a programmable chemical processor comprising of a 5 by 5 array of cells filled with a switchable oscillating chemical (Belousov-Zhabotinsky) reaction. Each cell can be individually addressed in the 'on' or 'off' state, yielding more than 2.9 × 1017 chemical states which arise from the ability to detect distinct amplitudes of oscillations via image processing. By programming the array of interconnected BZ reactions we demonstrate chemically encoded and addressable memory, and we create a chemical Autoencoder for pattern recognition able to perform the equivalent of one million operations per second.

Environmental control programs the emergence of distinct functional ensembles from unconstrained chemical reactions.

Many approaches to the origin of life focus on how the molecules found in biology might be made in the absence of biological processes, from the simplest plausible starting materials. Another approach could be to view the emergence of the chemistry of biology as process whereby the environment effectively directs "primordial soups" toward structure, function, and genetic systems over time. This does not require the molecules found in biology today to be made initially, and leads to the hypothesis that environment can direct chemical soups toward order, and eventually living systems. Herein, we show how unconstrained condensation reactions can be steered by changes in the reaction environment, such as order of reactant addition, and addition of salts or minerals. Using omics techniques to survey the resulting chemical ensembles we demonstrate there are distinct, significant, and reproducible differences between the product mixtures. Furthermore, we observe that these differences in composition have consequences, manifested in clearly different structural and functional properties. We demonstrate that simple variations in environmental parameters lead to differentiation of distinct chemical ensembles from both amino acid mixtures and a primordial soup model. We show that the synthetic complexity emerging from such unconstrained reactions is not as intractable as often suggested, when viewed through a chemically agnostic lens. An open approach to complexity can generate compositional, structural, and functional diversity from fixed sets of simple starting materials, suggesting that differentiation of chemical ensembles can occur in the wider environment without the need for biological machinery.

Development of a Minimal Photosystem for Hydrogen Production in Inorganic Chemical Cells.

Inorganic chemical cells (iCHELLs) are compartment structures consisting of polyoxometalates (POMs) and cations, offering structured and confined reaction spaces bounded by membranes. We have constructed a system capable of efficient anisotropic and hierarchical photo-induced electron transfer across the iCHELL membrane. Mimicking photosynthesis, our system uses proton gradients between the compartment and the bulk to drive efficient conversion of light into chemical energy, producing hydrogen upon irradiation. This illustrates the power of the iCHELL approach for catalysis, where the structure, compartmentalisation and variation in possible components could be utilised to approach a wide range of reactions.

A recursive microfluidic platform to explore the emergence of chemical evolution.

We propose that a chemically agnostic approach to explore the origin of life, using an automated recursive platform based on droplet microfluidics, could be used to induce artificial chemical evolution by iterations of growth, speciation, selection, and propagation. To explore this, we set about designing an open source prototype of a fully automated evolution machine, comprising seven modules. These modules are a droplet generator, droplet transfer, passive and active size sorting, splitter, incubation chamber, reservoir, and injectors, all run together via a LabVIEWTM program integration system. Together we aim for the system to be used to drive cycles of droplet birth, selection, fusion, and propagation. As a proof of principle, in addition to the working individual modules, we present data showing the osmotic exchange of glycylglycine containing and pure aqueous droplets, showing that the fittest droplets exhibit higher osomolarity relative to their neighbours, and increase in size compared to their neighbours. This demonstrates the ability of our platform to explore some different physicochemical conditions, combining the efficiency and unbiased nature of automation with our ability to select droplets as functional units based on simple criteria.

Miller-Urey Spark-Discharge Experiments in the Deuterium World.

We designed and conducted a series of primordial-soup Miller-Urey style experiments with deuterated gases and reagents to compare the spark-discharge products of a "deuterated world" with the standard reaction in the "hydrogenated world". While the deuteration of the system has little effect on the distribution of amino acid products, significant differences are seen in other regions of the product-space. Not only do we observe about 120 new species, we also see significant differences in their distribution if the two hydrogen isotope worlds are compared. Several isotopologue matches can be identified in both, but a large proportion of products have no equivalent in the corresponding isotope world with ca. 43 new species in the D world and ca. 39 new species in the H world. This shows that isotopic exchange (the addition of only one neutron) may lead to significant additional complexity in chemical space under otherwise identical reaction conditions.

An all-inorganic polyoxometalate-polyoxocation chemical garden.

Herein, we show it is possible to produce wholly inorganic chemical gardens from a cationic polyoxometalate (POM) seed in an anionic POM solution, demonstrating a wholly POM-based chemical garden system that produces architectures over a wide concentration range. Six concentration dependent growth regimes have been discovered and characterized: clouds, membranes, slugs, tubes, jetting and budding.

Formation of oligopeptides in high yield under simple programmable conditions.

Many high-yielding reactions for forming peptide bonds have been developed but these are complex, requiring activated amino-acid precursors and heterogeneous supports. Herein we demonstrate the programmable one-pot dehydration-hydration condensation of amino acids forming oligopeptide chains in around 50% yield. A digital recursive reactor system was developed to investigate this process, performing these reactions with control over parameters such as temperature, number of cycles, cycle duration, initial monomer concentration and initial pH. Glycine oligopeptides up to 20 amino acids long were formed with very high monomer-to-oligomer conversion, and the majority of these products comprised three amino acid residues or more. Having established the formation of glycine homo-oligopeptides, we then demonstrated the co-condensation of glycine with eight other amino acids (Ala, Asp, Glu, His, Lys, Pro, Thr and Val), incorporating a range of side-chain functionality.

Development of a 3D printer using scanning projection stereolithography.

We have developed a system for the rapid fabrication of low cost 3D devices and systems in the laboratory with micro-scale features yet cm-scale objects. Our system is inspired by maskless lithography, where a digital micromirror device (DMD) is used to project patterns with resolution up to 10 µm onto a layer of photoresist. Large area objects can be fabricated by stitching projected images over a 5 cm(2) area. The addition of a z-stage allows multiple layers to be stacked to create 3D objects, removing the need for any developing or etching steps but at the same time leading to true 3D devices which are robust, configurable and scalable. We demonstrate the applications of the system by printing a range of micro-scale objects as well as a fully functioning microfluidic droplet device and test its integrity by pumping dye through the channels.

Integrated 3D-printed reactionware for chemical synthesis and analysis.

Three-dimensional (3D) printing has the potential to transform science and technology by creating bespoke, low-cost appliances that previously required dedicated facilities to make. An attractive, but unexplored, application is to use a 3D printer to initiate chemical reactions by printing the reagents directly into a 3D reactionware matrix, and so put reactionware design, construction and operation under digital control. Here, using a low-cost 3D printer and open-source design software we produced reactionware for organic and inorganic synthesis, which included printed-in catalysts and other architectures with printed-in components for electrochemical and spectroscopic analysis. This enabled reactions to be monitored in situ so that different reactionware architectures could be screened for their efficacy for a given process, with a digital feedback mechanism for device optimization. Furthermore, solely by modifying reactionware architecture, reaction outcomes can be altered. Taken together, this approach constitutes a relatively cheap, automated and reconfigurable chemical discovery platform that makes techniques from chemical engineering accessible to typical synthetic laboratories.

Morphogenesis of polyoxometalate cluster-based materials to microtubular network architectures.

Pressed pellets of polyoxometalate (POM)-based materials are shown to undergo morphogenesis to produce microtubular network architectures without the need for a large single crystal precursor. The compression of the POM material into a pellet lowers the solubility sufficiently to allow tube initiation and growth from POMs that would otherwise be too soluble in their native crystalline state, thus yielding hollow, highly controllable, tubes of diameter 10-100 µm.

Real-time ion-flux imaging in the growth of micrometer-scale structures and membranes.

Real-time ion flux imaging: an ion-sensitive field-effect transistor (ISFET) array is coupled with optical microscopy to image the growth of, and ion flux through, micrometer-scale tubes and membranes built from polyoxometalate clusters. The correlation between the optical and ionic imaging data is excellent, showcasing the use of ISFET arrays for high-resolution spatial and temporal mapping of ionic movements.

Osmotically driven crystal morphogenesis: a general approach to the fabrication of micrometer-scale tubular architectures based on polyoxometalates.

The process of osmotically driven crystal morphogenesis of polyoxometalate (POM)-based crystals is investigated, whereby the transformation results in the growth of micrometer-scale tubes 10-100 µm in diameter and many thousands of micrometers long. This process initiates when the crystals are immersed in aqueous solutions containing large cations and is governed by the solubility of the parent POM crystal. Evidence is presented that indicates the process is general to all types of POMs, with solubility of the parent crystal being the deciding parameter. A modular approach is adopted since different POM precursor crystals can form tubular architectures with a range of large cationic species, producing an ion-exchanged material that combines the large added cations and the large POM-based anions. It is also shown that the process of morphogenesis is electrostatically driven by the aggregation of anionic metal oxides with the dissolved cations. This leads to the formation of a semi-permeable membrane around the crystal. The osmotically driven ingress of water leads to an increase in pressure, and ultimately rupture of the membrane occurs, allowing a saturated solution of the POM to escape and leading to the formation of a "self-growing" microtube in the presence of the cation. It is demonstrated that the growth process is sustained by the osmotic pressure within the membrane surrounding the parent crystal, as tube growth ceases whenever this pressure is relieved. Not only is the potential of the modular approach revealed by the fact that the microtubes retain the properties of their component parts, but it is also possible to control the direction of growth and tube diameter. In addition, the solubility limits of tube growth are explored and translated into a predictive methodology for the fabrication of tubular architectures with predefined physical properties, opening the way for real applications.

Unveiling the transient template in the self-assembly of a molecular oxide nanowheel.

Self-assembly has proven a powerful means of preparing structurally intricate nanomaterials, but the mechanism is often masked by the common one-pot mixing procedure. We employed a flow system to study the steps underlying assembly of a previously characterized molybdenum oxide wheel 3.6 nanometers in diameter. We observed crystallization of an intermediate structure in which a central {Mo36} cluster appears to template the assembly of the surrounding {Mo150} wheel. The transient nature of the template is demonstrated by its ejection after the wheel is reduced to its final electronic state. The template's role in the self-assembly mechanism is further confirmed by the deliberate addition of the template to the reaction mixture, which greatly accelerates the assembly time of the {Mo150} wheel and increases the yield.