Rapid detection and quantification of haptophyte alkenones by Fourier transform infrared spectroscopy (FTIR)

Abstract

Several haptophyte algae produce unique neutral long-chain (C-C 40) ketones (alkenones) with two to four trans-carbon-carbon double (C=C) bonds and a keto-group. These molecules are of great biological interest and may have substantial commercial relevance as biofuel sources. Unfortunately, their detection and quantification,involving extraction from cells with organic solvents and fractionation of lipids through columns,are rather cumbersome. Hence, wedeveloped a method for the rapid and reliable quantification of alkenones in intact cells using Fourier transform infrared spectroscopy (FTIR). The method is based on theabsorption band at 962.5 cm-1 ,whichcorresponds to atrans-C=C bond that isunequivocally attributable to alkenones. This peak was exclusively observed in alkenone-producing haptophytessuch as Emiliania huxleyi NIES-837, Emiliania huxleyi CCMP 2090,Tisochrysis lutea(formerIsochrysis galbanaT-iso), Tisochrysis luteaCCMP 463, and Chrysotila lamellosa CCMP 1307,as well as in appropriate standards. We compared our FTIR method with a typical GC analysis method. The alkenone quantitydetermined by these two methods showed similar values. Nevertheless, the FTIRmethod developed in this studydoes not require complexextraction procedures and is therefore a much easier and rapid quantification method. This FTIR method can also be used for the screening of strains and the optimization of culture conditions for alkenone production.

Phosphorus and nitrogen starvation reveal life‐cycle specific responses in the metabolome of Emiliania huxleyi (Haptophyta)

“…Recently, alkenones in E. huxleyi have been semi‐quantified using the peak at 963 cm −1 , attributed to the trans ‐double bonds (Pelusi et al ). Together with the peak at 1706 cm −1 of the keto groups, it is specific for alkenones (Pelusi et al ). Here, the signal at 1706 cm −1 could not be isolated, but the contribution of polyunsaturated hydrocarbons, such as alkenones and alkenes, was identified (3013 cm −1 and 963 cm −1 ), being ∼10% larger in the diploid (absorbance A = 0.0046) (Fig.…”

“…The contribution of alkenes and calcium carbonate to the bands was validated by generating reference spectra of hexatriacontane (98%, Sigma‐Aldrich), tetradecane (99%, Acros organics), trans‐7‐tetradecene (97%, Sigma‐Aldrich), and calcium carbonate (>99%, Sigma‐Aldrich). Bands of proteins, phospholipids, carbohydrates, and alkenones were identified based on comparison with literature (Giordano et al ; Naumann ; Stehfest et al ; Pelusi et al ).…”

“…1b,c). Recently, alkenones in E. huxleyi have been semi-quantified using the peak at 963 cm 21 , attributed to the trans-double bonds (Pelusi et al 2016). Together with the peak at 1706 cm 21 of the keto groups, it is specific for alkenones (Pelusi et al 2016).…”

Overexpression of Tisochrysis lutea Akd1 identifies a key cold-induced alkenone desaturase enzyme

“…The alkenone-producing haptophyte alga, Tisochrysis lutea (T-iso) (formerly Isochrysis galbana T-iso strain) 19 , 25 , kindly provided by Prof. T. Matsunaga of Tokyo University of Agriculture and Technology (Tokyo, Japan), was used in this study as describes in our previous report 34 . For this study, to select a high-alkenone-producing cell line, cells were separated on agar plate, and a single colony was isolated and established as a strain for further use.…”

“…In all experiments, the cultures were continuously bubbled and illuminated by 18 or 36 W fluorescent lamps at an intensity of 100 μmol photons m −2 s −1 . After sampling, TOC and alkenone contents were determined using a TOC-L analyser (Shimadzu Corporation) and GC-FID as describes in our previous report 34 . The data were obtained from four independent experiments.…”