Non-destructive monitoring tools for biomass and astaxanthinproduction in Haematococcus pluvialis rotating biofilms: aspectroscopy approach

Description

Biofilm-based systems have gained increasing attention for microalgae cultivation dueto their potential to improve productivity, reduce water and energy demands andeventually operating costs. However, effective operation of these systems requiresprocess control, which relies on the development of on-line monitoring of processvariables such as biomass, pigments and lipids contents. While there are variousmethods and technologies for monitoring these biological parameters in microalgaesuspensions, at present, there is a lack of tools for non-destructively monitoring biofilmbased systems.In this study, we developed a biofilm-based rotating system for cultivatingHaematococcus pluvialis biofilms for astaxanthin production (high-value carotenoid).The dynamics of biomass and astaxanthin production were afterwards determinedunder several light and nutrient conditions. The data collected were then used todevelop regression models based on Fourier-transform infrared (FTIR) and reflectancespectroscopy (Vis-NIR; 380-1000 nm) to non-destructively monitor biofilm growth andastaxanthin content.FTIR spectroscopy is an effective technique for high-throughput screening in biologyas it can distinguish and quantify various components such as proteins, lipids, nucleicacids, and carbohydrates. In H. pluvialis, the biosynthesis of astaxanthin matches theaccumulation of triacylglycerols (TAGs) leading to a complex macromolecularreorganization (Figure 1), suggesting that FTIR spectra could be used to quantify theastaxanthin content. Indeed, cell populations rich in astaxanthin could be easilyidentified by the ratio between macromolecules, and a linear regression betweenastaxanthin content and the lipids-to-proteins ratio was obtained (Figure 2). Thismethod is non-invasive, does not require a chemical extraction step, and can be usedto monitor and predict astaxanthin production in microalgal biofilm systems. However,the strong absorption of water between 1700 and 900 cm-1 may limit its application forreal-time monitoring. To address this, we propose the use of reflectance spectroscopy,which stands on the light reflected at specific wavelengths to calculate indexes that arewidely applied to remotely characterize plants and microphytobenthos communities.These indexes strongly reflect specific physiological mechanisms and biochemicalcomposition, making them effective in estimating biomass, pigments, and identifyingstress changes. To demonstrate this, we applied the same principle to our biofilmbased system and develop indexes for estimating biomass and astaxanthin content.Reflectance measurements were performed at the biofilm surface over time (Figure 3).The most influential wavelengths were afterwards determined for these variables asλ525, λ637, λ563, λ678 nm and near-infrared (NIR λ750-900 nm). A strong correlationbetween biomass and an index based on NIR and λ525 nm was found (R2 = 0.938);another was established between an index using λ563 and λ637 and astaxanthincontent (R2= 0.944) (Figure 4).In summary, we developed, for the first time, non-destructive and in situ monitoringtools for microalgae biofilm characterization. Our findings address one of the limitationsin process control, and have the potential to improve the operation of biofilm-basedsystems, making them a promising technology for microalgal cultivation and theproduction of high-value compounds

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