Soft sensor based on Raman spectroscopy for the in-line monitoring of metabolites and polymer quality in the biomanufacturing of polyhydroxyalkanoates.

Polyhydroxyalkanoates (PHA) are among the most promising bio-based alternatives to conventional petroleum-based plastics. These biodegradable polyesters can in fact be produced by fermentation from bacteria like Cupriavidus necator, thus reducing the environmental footprint of the manufacturing process. However, ensuring consistent product quality attributes is a major challenge of biomanufacturing. To address this issue, the implementation of real-time monitoring tools is essential to increase process understanding, enable a prompt response to possible process deviations and realize on-line process optimization. In this work, a soft sensor based on in situ Raman spectroscopy was developed and applied to the in-line monitoring of PHA biomanufacturing. This strategy allows the collection of quantitative information directly from the culture broth, without the need for sampling, and at high frequency. In fact, through an optimized multivariate data analysis pipeline, this soft sensor allows monitoring cell dry weight, as well as carbon and nitrogen source concentrations with root mean squared errors (RMSE) equal to 3.71, 7 and 0.03 g/L, respectively. In addition, this tool allows the in-line monitoring of intracellular PHA accumulation, with an RMSE of 14 gPHA/gCells. For the first time, also the number and weight average molecular weights of the polymer produced could be monitored, with RMSE of 8.7E4 and 11.6E4 g/mol, respectively. Overall, this work demonstrates the potential of Raman spectroscopy in the in-line monitoring of biotechnology processes, leading to the simultaneous measurement of several process variables in real time without the need of sampling and labor-intensive sample preparations.

Current trends in the production of biodegradable bioplastics: The case of polyhydroxyalkanoates.

The global pollution caused by plastics and microplastics is stimulating intense research towards more environmentally friendly materials, preserving the remarkable application characteristics of the currently available polymers. Among these, polyhydroxyalkanoates (PHAs) have been hailed as the solution to replace conventional, oil-based plastics. Given their biodegradable nature and mechanical properties, their use can be envisioned in a wide range of applications reducing the environmental footprint. Several types of processes have been proposed for their production, which can be grouped in three main classes: (i) microbiological, (ii) enzymatic and (iii) chemical processes. Given the significant amount of literature available on this topic, this review aims to critically analyse what has been proposed so far in each of these classes, with specific reference to their potential to provide bioplastics that can actually replace the currently available materials. A comparison is made, based on the following aspects: achievable molecular structures (such as molecular weight and composition distributions), raw-material and production costs and availability of large-scale production technologies. Finally, some considerations and ideas on what should be further investigated and implemented to realize the economically sustainable production of PHA are brought forward.

Oligonucleotides: Current Trends and Innovative Applications in the Synthesis, Characterization, and Purification.

Oligonucleotides (ONs) are gaining increasing importance as a promising novel class of biopharmaceuticals. Thanks to their fundamental role in gene regulation, they can be used to develop custom-made drugs (also called N-to-1) able to act on the gene expression at pre-translational level. With recent approvals of ON-based therapeutics by the Food and Drug Administration (FDA), a growing demand for high-quality chemically modified ONs is emerging and their market is expected to impressively prosper in the near future. To satisfy this growing market demand, a scalable and economically sustainable ON production is needed. In this paper, the state of the art of the whole ON production process is illustrated with the aim of highlighting the most promising routes toward the auspicated market-size production. In particular, the most recent advancements in both the upstream stage, mainly based on solid-phase synthesis and recombinant technology, and the downstream one, focusing on chromatographic techniques, are reviewed. Since ON production is projected to expand to the large scale, automatized multicolumn countercurrent technologies will reasonably be required soon to replace the current ones based on batch single-column operations. This consideration is supported by a recent cutting-edge application of continuous chromatography for the ON purification.