Superior triacylglycerol (TAG) accumulation in starchless mutants of Scenedesmus obliquus: (II) evaluation of TAG yield and productivity in controlled photobioreactors

Abstract

Background: Many microalgae accumulate carbohydrates simultaneously with triacylglycerol (TAG) upon nitrogen starvation, and these products compete for photosynthetic products and metabolites from the central carbon metabolism. As shown for starchless mutants of the non-oleaginous model alga Chlamydomonas reinhardtii, reduced carbohydrate synthesis can enhance TAG production. However, these mutants still have a lower TAG productivity than wild-type oleaginous microalgae. Recently, several starchless mutants of the oleaginous microalga Scenedesmus obliquus were obtained which showed improved TAG content and productivity.

Results: The most promising mutant, slm1, is compared in detail to wild-type S. obliquus in controlled photobioreactors. In the slm1 mutant, the maximum TAG content increased to 57 ± 0.2% of dry weight versus 45 ± 1% in the wild type. In the wild type, TAG and starch were accumulated simultaneously during initial nitrogen starvation, and starch was subsequently degraded and likely converted into TAG. The starchless mutant did not produce starch and the liberated photosynthetic capacity was directed towards TAG synthesis. This increased the maximum yield of TAG on light by 51%, from 0.144 ± 0.004 in the wild type to 0.217 ± 0.011 g TAG/mol photon in the slm1 mutant. No differences in photosynthetic efficiency between the slm1 mutant and the wild type were observed, indicating that the mutation specifically altered carbon partitioning while leaving the photosynthetic capacity unaffected.

Conclusions: The yield of TAG on light can be improved by 51% by using the slm1 starchless mutant of S. obliquus, and a similar improvement seems realistic for the areal productivity in outdoor cultivation. The photosynthetic performance is not negatively affected in the slm1 and the main difference with the wild type is an improved carbon partitioning towards TAG.

Keywords: Acutodesmus obliquus; Scenedesmus obliquus; mutant; starch; starchless; triacylglycerol (TAG).

Figure 1

Duplicate batch nitrogen run-out cultivations of the wt and the slm1 S. obliquus. Left (A, C): wild-type. Right (B, D): slm1. Top (A, B): biomass concentration (g DW/l) and dissolved NO₃-N concentration. Bottom (C, D): total fatty acid (TFA), TAG, and starch content. As indicated in the legend boxes in the figure, in the top figure, the black symbols represent the dry weight concentration and the open symbols represent the NO₃-N concentration. In the bottom figures, the open symbols represent the total fatty acid content (TFA), the gray symbols represent the TAG content, and the black symbols represent the starch content. The results indicated with #1 and #2 in the figure legend represent the replicate cultivations.

Figure 2

TAG concentration (A), time-averaged yield of TAG on photons (B), and time-averaged volumetric TAG productivity (C). Open symbols: wt. Gray symbols: slm1. The time-averaged yield and volumetric productivity are calculated over the period between inoculation and each time point. The results indicated with #1 and #2 in the figure legend represent the replicate cultivations.

Figure 3

Theoretical minimum amount of photons required to produce the observed biomass after nitrogen is depleted. Open symbols: wt. Gray symbols: slm1. The results indicated with #1 and #2 in the figure legend represent the replicate cultivations.

Figure 4

Impact of nitrogen starvation on photosynthesis. (A) Fv/Fm ratio. (B) Absorbance cross section. (C) Absorbance spectrum of the slm1 and wt under nitrogen replete and nitrogen depleted conditions, normalized to the absorbance at 680 nm. (D) Ratio of absorbance at 483 nm and 680 nm. In figure C, the time indicated in the figure legend represents the time after inoculation. Nitrogen starvation commenced 70 to 100 h after inoculation. The spectrum that is indicated with N+ in the figure legend represents the spectrum before nitrogen was depleted. Open symbols indicate the wt and gray symbols indicate the slm1. The results indicated with #1 and #2 in the figure legend represent the replicate cultivations.