Medeiros Garcia Alcântara João, Iannacci Francesco, Morbidelli Massimo, Sponchioni Mattia
Dept. of Chemistry, Materials and Chemical Engineering "Giulio Natta", Politecnico di Milano, via Mancinelli 7, Milano 20131, Italy.
Dept. of Chemistry, Materials and Chemical Engineering "Giulio Natta", Politecnico di Milano, via Mancinelli 7, Milano 20131, Italy.
J Biotechnol. 2023 Nov 20;377:23-33. doi: 10.1016/j.jbiotec.2023.10.005. Epub 2023 Oct 23.
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 g/g 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.
聚羟基脂肪酸酯（PHA）是传统石油基塑料最有前景的生物基替代品之一。事实上，这些可生物降解的聚酯可以通过像食酸戴尔福特菌这样的细菌发酵生产，从而减少制造过程的环境足迹。然而，确保产品质量属性的一致性是生物制造的一项重大挑战。为了解决这个问题，实施实时监测工具对于增进对过程的理解、对可能的过程偏差做出迅速反应以及实现在线过程优化至关重要。在这项工作中，开发了一种基于原位拉曼光谱的软传感器，并将其应用于PHA生物制造的在线监测。这种策略允许直接从培养液中收集定量信息，无需取样，且频率很高。事实上，通过优化的多变量数据分析流程，这种软传感器能够监测细胞干重以及碳源和氮源浓度，其均方根误差（RMSE）分别为3.71、7和0.03 g/L。此外，该工具还能在线监测细胞内PHA的积累，RMSE为14 g/g。首次还能够监测所生产聚合物的数均分子量和重均分子量，RMSE分别为8.7×10⁴和11.6×10⁴ g/mol。总体而言，这项工作展示了拉曼光谱在生物技术过程在线监测中的潜力，能够在无需取样和繁琐样品制备的情况下实时同时测量多个过程变量。
