Laboratório de Engenharia de Bioprocessos. Escola de Artes, Ciências e Humanidades (EACH), Universidade de São Paulo, Rua Arlindo Béttio, 1000, CEP 03828-000 São Paulo, SP, Brazil.
Laboratório de Biotecnologia Viral, Instituto Butantan, Av Vital Brasil 1500, CEP 05503-900 São Paulo, SP, Brazil.
Spectrochim Acta A Mol Biomol Spectrosc. 2025 Feb 5;326:125217. doi: 10.1016/j.saa.2024.125217. Epub 2024 Sep 24.
The Zika disease caused by the Zika virus was declared a Public Health Emergency by the World Health Union (WHO), with microcephaly as the most critical consequence. Aiming to reduce the spread of the virus, biopharmaceutical organizations invest in vaccine research and production, based on multiple platforms. A crescent vaccine production approach is based on virus-like particles (VLP), for not having genetic material in its composition, hypoallergenic and non-mutant character. For bioprocess, it is essential to have means of real-time monitoring, which can be assessed using process analysis techniques such as Near-infrared (NIR) spectroscopy, that can be combined with chemometric methods, like Partial-Least Squares (PLS) and Artificial Neural Networks (ANN) for prediction of biochemical variables. This work proposes a biochemical Zika VLP upstream production at-line monitoring model using NIR spectroscopy comparing sampling conditions (with or without cells), analytical blank (air, ultrapure water), and spectra pre-processing approaches. Seven experiments in a benchtop bioreactor using recombinant baculovirus/Sf9 insect cell platform in serum-free medium were performed to obtain biochemical and spectral data for chemometrics modeling (PLS and ANN), composed by a random data split (80 % calibration, 20 % validation) for cross-validation of the PLS models and 70 % training, 15 % testing, 15 % validation for ANN. The best models generated in the present work presented an average absolute error of 1.59 × 10 cell/mL for density of viable cells, 2.37 % for cell viability, 0.25 g/L for glucose, 0.007 g/L for lactate, 0.138 g/L for glutamine, 0.18 g/L for glutamate, 0,003 g/L for ammonium, and 0.014 g/L for potassium.
由 Zika 病毒引起的 Zika 病被世界卫生组织（WHO）宣布为公共卫生紧急事件，其最严重的后果是小头畸形。为了减少病毒的传播，生物制药组织基于多种平台投资于疫苗的研究和生产。一种新兴的疫苗生产方法是基于病毒样颗粒（VLP），因为其组成中没有遗传物质，具有低致敏性和非突变性。对于生物工艺，必须有实时监测的手段，可以使用过程分析技术（如近红外（NIR）光谱）进行评估，该技术可以与化学计量学方法（如偏最小二乘（PLS）和人工神经网络（ANN））相结合，用于预测生化变量。这项工作提出了一种使用 NIR 光谱对 Zika VLP 上游生物工艺进行在线监测的模型，比较了采样条件（有细胞和无细胞）、分析空白（空气、超纯水）和光谱预处理方法。在无血清培养基中使用重组杆状病毒/Sf9 昆虫细胞平台的台式生物反应器中进行了七项实验，以获得生化和光谱数据用于化学计量学建模（PLS 和 ANN），由随机数据分割（80%的校准，20%的验证）组成，用于 PLS 模型的交叉验证和 70%的训练、15%的测试、15%的验证用于 ANN。本工作生成的最佳模型对活细胞密度的平均绝对误差为 1.59×10 个细胞/mL，细胞活力的平均绝对误差为 2.37%，葡萄糖的平均绝对误差为 0.25g/L，乳酸的平均绝对误差为 0.007g/L，谷氨酰胺的平均绝对误差为 0.138g/L，谷氨酸的平均绝对误差为 0.18g/L，铵的平均绝对误差为 0.003g/L，钾的平均绝对误差为 0.014g/L。
