Investigating the Impact of Fermentation on Polyphenolic Content in Cultivated Sugar Kelp (Saccharina latissima): A Comprehensive Study
Master thesis
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Date
2024-06-03Metadata
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- Department of Chemistry [464]
Abstract
Seaweeds, classified as macroalgae, are a marine bioresource garnering global attention, primarily due to the increasing demand for new sustainable bioresources. Seaweeds are rich in a variety of bioactive compounds that hold promising health benefits. One such group of compounds is polyphenols. These diverse compounds, which are biosynthesized in both plants and algae, are of special interest due to their numerous potential health advantages. The brown seaweed Saccharina latissima (Laminariaceae), commonly called sugar kelp, has been the pioneer species in Norwegian seaweed cultivation – and is still the most widely cultivated seaweed in Norway. To ensure the year-round availability of harvested sugar kelp, researchers are currently exploring the potential of lactic acid bacteria fermentation as a preservation technique. The present project investigates how fermentation affects the polyphenolic content in samples of cultivated sugar kelp, with a special focus on the liquid released during the fermentation process.
Lerøy Ocean Forest provided 13 fermented sugar kelp samples (F1-F13), stored at a controlled temperature at the Department of Chemistry, UiB. Fermentation duration, which was halted by freezing, ranged from one day (F1) to 36 weeks (F13), with intervals in between. This allowed a thorough study of the impact of fermentation duration on the polyphenolic contents in sugar kelp. The samples were categorized into biomass (stipe and leaves) and fermentation liquid, the latter being fluid released during fermentation. The fermentation liquid samples were initially dried and subjected to a total phenolic content (TPC) assay using Folin-Ciocalteu’s (FC) reagent. Despite several different approaches, the analysis was unable to identify a clear pattern among the liquid samples fermented with varying durations. This was likely due to the complex physiochemical properties of these samples, and most likely affected by high sugar content interfering with the analysis.
An alternative sample preparation technique was explored, including a work-up step with chromatography (Amberlite XAD-7) to semi-purify the samples prior to analysis, aiming to remove sugars affecting the analysis. This method was subsequently applied to a selected set of samples. For the new series of samples, liquid samples from both the initial and final stages of the fermentation process were selected. Additionally, the biomass originating from the same samples were chosen for comparison. These biomass samples were extracted using a 60:40 methanol:water solution. Following extraction, both the biomass extracts and the liquid samples were semi-purified using the prepared column filled with Amberlite XAD-7. The samples were first rinsed with 2-3 L of distilled water, then eluted with 100% methanol. The methanol extracts from this purification process were subsequently dried and subjected to further analysis.
The polyphenol contents in the semi-purified samples were quantified using two methods: the FC-TPC assay and a selective quantitative nuclear magnetic resonance spectroscopy (s-qNMR) method. The latter has been specifically optimized for seaweed polyphenol quantification inhouse by the Jordheim Group (Wekre et al., 2022). Both methods revealed a significant increase in the polyphenol content of the liquid samples as the fermentation duration increased. For instance, the liquid sample fermented for 36 weeks (L13) exhibited polyphenolic concentrations of 91.3±4.7 mg GAE/g DW with the TPC method and 44.7±1.3 mg GAE/g DW with the s-qNMR method. In contrast, the liquid sample that was only fermented for one day (L1) had considerably lower polyphenolic concentrations, measuring 10.9±0.6 mg GAE/g DW with the TPC method and 17.0±0.4 mg GAE/g DW with the s-qNMR method.
The HPLC-DAD analysis of these samples showed a higher quantity of compounds in sample L13 that absorb UV-Vis waves around 330±20 nm, compared to what was seen in sample L1. This could possibly be attributed to a higher amount of low-molecular-weight phenolic compounds like phenolic acids in sample L13, but further separation and isolation would be required to draw more definite conclusions from this analysis.The antioxidant abilities in the semi-purified samples were measured with DPPH assay analysis. The IC50 values, which represent the concentration of the sample required to neutralize 50% of the free radical DPPH, were 9.95±0.63 mg/mL for sample L1 and 0.82±0.07 mg/mL for sample L13. Given that the IC50 value is inversely proportional to antioxidant activity, and polyphenols are known to have antioxidant abilities, these findings support the polyphenol contents being higher in sample L13 (fermented 36 weeks).
In summary, the findings in this study suggest an increase in polyphenolic content and antioxidant capacity in liquid samples with extended fermentation duration. The underlying cause of this increase is yet to be determined. It may be linked to chemical changes in the composition of the fermentation liquid, resulting in higher antioxidant capacity in later stages of fermentation, or to an increased phenolic extraction capacity of the liquid with time. Despite challenges in pinpointing the exact origin of increased polyphenol concentrations, the fermentation liquid shows potential commercial value in food, feed, and health sectors, warranting further investigation.
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Postponed access: the file will be accessible after 2029-06-03