Implementing stepwise solvent elution in sequential injection chromatography for fluorimetric determination of intracellular free amino acids in the microalgae Tetraselmis gracilis

The concept of sequential injection chromatography (SIC) was exploited to automate the fluorimetric determination of amino acids after pre-column derivatization with o-phthaldialdehyde (OPA) in presence of 2-mercaptoethanol (2MCE) using a reverse phase monolithic C₁₈ stationary phase. The method is low-priced and based on five steps of isocratic elutions. The first step employs the mixture methanol: tetrahydrofuran: 10 mmol L⁻¹ phosphate buffer (pH 7.2) at the volumetric ratio of 8:1:91; the other steps use methanol: 10 mmol L⁻¹ phosphate buffer (pH 7.2) at volumetric ratios of 20:80, 35:65, 50:50 and 65:35. At a flow rate of 10 μL s⁻¹ a 25 mm long-column was able to separate aspartic acid (Asp), glutamic acid (Glu), asparagine (Asn), serine (Ser), glutamine (Gln), glycine (Gly), threonine (Thr), citruline (Ctr), arginine (Arg), alanine (Ala), tyrosine (Tyr), phenylalanine (Phe), ornithine (Orn) and lysine (Lys) with resolution >1.2 as well as methionine (Met) and valine (Val) with resolution of 0.6. Under these conditions isoleucine (Ile) and leucine (Leu) co-eluted. The entire cycle of amino acids derivatization, chromatographic separation and column conditioning at the end of separation lasted 25 min. At a flow rate of 40 μL s⁻¹ such time was reduced to 10 min at the cost of resolution worsening for the pairs Ctr/Arg and Orn/Lys. The detection limits varied from 0.092 μmol L⁻¹ for Tyr to 0.51 μmol L⁻¹ for Orn. The method was successfully applied to the determination of intracellular free amino acids in the green alga Tetraselmis gracilis during a period of seven days of cultivation. Samples spiked with known amounts of amino acids resulted in recoveries between 94 and 112%.     

Evaluation of sample extraction methods for proteomics analysis of green algae Chlorella vulgaris

Many protein extraction methods have been developed for plant proteome analysis but information is limited on the optimal protein extraction method from algae species. This study evaluated four protein extraction methods, i.e. direct lysis buffer method, TCA‐acetone method, phenol method, and phenol/TCA‐acetone method, using green algae Chlorella vulgaris for proteome analysis. The data presented showed that phenol/TCA‐acetone method was superior to the other three tested methods with regards to shotgun proteomics. Proteins identified using shotgun proteomics were validated using sequential window acquisition of all theoretical fragment‐ion spectra (SWATH) technique. Additionally, SWATH provides protein quantitation information from different methods and protein abundance using different protein extraction methods was evaluated. These results highlight the importance of green algae protein extraction method for subsequent MS analysis and identification.     

Halogenated Sesquiterpenes from the Marine Red Alga Laurencia saitoi (Rhodomelaceae)

Four new halogenated sesquiterpenes, 10‐bromo‐3‐chloro‐2,7‐epoxychamigr‐9‐en‐8α‐ol ( 1 ), 2,10β‐dibromochamigra‐2,7‐dien‐9α‐ol ( 2 ), (9S)‐2‐bromo‐3‐chloro‐6,9‐epoxybisabola‐7(14),10‐diene ( 3 ), and (9R)‐2‐bromo‐3‐chloro‐6,9‐epoxybisabola‐7(14),10‐diene ( 4 ), were characterized from the marine red alga Laurencia saitoi. In addition, two known halosesquiterpenes, 2,10‐dibromo‐3‐chlorochamigr‐7‐en‐9α‐ol ( 5 ) and isolaurenisol ( 6 ), were also isolated and identified. Their structures were established on the basis of extensive analysis of spectroscopic data.     

Simultaneous determination of toxins in algae and water samples by high-performance liquid chromatography with triple quadrupole mass spectrometry

A facile method based on liquid chromatography coupled with electrospray ionization tandem triple quadrupole mass spectrometry working in selected reaction monitoring mode has been established to analyze toxins in the algae and water samples. Twelve types of toxins (anatoxin, cylindrospermopsin, dinophysistoxin-1, nodularin, okadaic acid, microcystins) were efficiently separated under optimized liquid chromatography coupled with mass spectrometry conditions in the selected reaction monitoring mode. Correlation coefficients of the calibration curves, all felt in the range of 0.9958-0.9998, indicated good linearity. The detection limits of toxins in this method were all lower than 0.20 ng/mL and the quantification limits were in the range from 0.04 to 0.60 ng/mL. Except for anatoxin, cylindrospermopsin, and nodularin, the other toxins' recoveries varied from 55.45 to 140.85%. And the relative standard deviations of interday and intraday precision were at 8.61% (n = 5). The high-performance liquid chromatography (HPLC)/electrospray ionization (ESI)-mass spectrometery (MS) method was also successfully applied to analyze the algae and water samples. Owing to its exclusive selectivity and excellent sensitivity, the developed method is a tool for comprehensive analyses of the 12 types of toxins at nanogram levels.     

CE–MS fingerprinting of Laurencia complex algae (Rhodophyta)

The use of CE–ESI‐MS has been considered as a new chemical strategy for the possible discernment of genera and species of the Laurencia complex. After the selection of the CE–MS and the extraction conditions, a total of 28 specimens of the complex, including different species of four genera (Laurencia, Laurenciella, Palisada, and Osmundea) collected from five intertidal locations on the Island of Tenerife (Canary Islands, Spain) were analyzed. CE–MS fingerprints revealed that CE–MS can be used as a useful tool for these studies in order to assess similarities and differences between them and that it constitutes an important starting point for further studies in the field.     

太湖藻及模型化合物的分级热解 (cited: 1)

以太湖藻为原料,通过分级热解来制备生物油,分别以棕榈酸、琼脂糖和蛋清粉模拟藻中的脂质、糖类和蛋白质,研究了藻分级热解生物油中主要组分的来源及热解过程中三大组分之间的相互作 用．结果表明,藻生物油中的烃类、十六酸及酯类均源自藻中脂质的热解,藻中蛋白质热解主要生成酚类、吲哚类、吡咯类和小分子酸;生物油中大部分的酰胺和腈类是脂质和蛋白质热解中间物相互作用的产物．藻组分中脂质和糖类的热解主要发生在473～573 K,而蛋白质的热解主要发生在523 K以上．藻生物油中仅有少量的糖类直接热解产物,可能是糖类热解产物在蛋白质热解产生的氨气等的作用下发生了聚合,使473和523 K所得生物油中有相当量的寡聚物;脂质与蛋白质的热解中间物相互作用生成了酰胺,酰胺再脱水又生成了腈类;而脂质与糖类之间相互作用不明显．     

Significance of transparent exopolymer particles derived from aquatic algae in membrane fouling

In recent years, Transparent exopolymer particles(TEPs) have been identified as significant contributors to membrane surface biofouling. Reported research on the effect of TEPs on membrane fouling has mainly focused on algae-derived TEPs in the ocean, and very limited investigations have been conducted on those in freshwater systems. In this study, we investigated the characteristics of TEPs derived from Microcystis aeruginosa and their influence on membrane fouling in an ultrafiltration (UF) system. The results indicated that bound TEPs could lead to more serious membrane fouling while free TEPs caused more serious irreversible membrane fouling. Further studies showed that in free TEP solutions, small-sized colloidal TEPs (c-TEPs) rather than large-sized particle TEPs (p-TEPs) showed a significantly positive correlation with irreversible membrane fouling. The presence of Ca²⁺ ions in influent water can reduce membrane fouling to some extent since a low concentration of Ca²⁺ ions (1 mM) can lead to the transformation of most free TEPs from the colloidal to particulate state. Both acidic and alkaline environments of free TEP solutions result in more serious membrane fouling compared to a neutral environment of free TEP solution. The negative impact of the acidic environment on membrane fouling was more significant than that of the alkaline environment. The abovementioned results show that when using a UF system to filter water with high algal content, greater attention should be paid to free TEPs, especially those in the colloidal state, because they can cause serious, irreversible membrane fouling.     

Evaluation of ionic liquid stationary phases for one dimensional gas chromatography-mass spectrometry and comprehensive two dimensional gas chromatographic analyses of fatty acids in marine biota

Ionic liquid stationary phases were tested for one dimensional gas chromatography-mass spectrometry (GC-MS) and comprehensive two dimensional gas chromatography (GC×GC) of fatty acid methyl esters from algae. In comparison with polyethylene glycol and cyanopropyl substituted polar stationary phases, ionic liquid stationary phases SLB-IL 82 and SLB-IL 100 showed comparable resolution, but lower column bleeding with MS detection, resulting in better sensitivity. The selectivity and polarity of the ionic liquid phases are similar to a highly polar biscyanopropyl-silicone phase (e.g. HP-88). In GC×GC, using an apolar polydimethyl siloxane×polar ionic liquid column combination, an excellent group-type separation of fatty acids with different carbon numbers and number of unsaturations was obtained, providing information that is complementary to GC-MS identification.     

Fractional Pyrolysis of Algae and Model Compounds (cited: 1)

Pyrolysis of algae from Taihu Lake water blooms for bio-oil production was conducted from 473 K to 773 K by a fractional way in six steps. Palmitic acid, agarose and egg white were used as model compounds to study the origin of bio-oil ingredients and interaction of the intermediates from the algae components. In the first step at 473 K, the bio-oil obtained was composed of n-heptadecane and some small molecule acids. Quantities of carboxylic acids (mainly palmitic acid) and some amides, hydrocarbons, esters etc. were evolved in the second step at 523 K. For the third step at 573 K, except the carboxylic acids (still mainly palmitic acid), amides, nitriles, and phenols also accounted for a large proportion whereas respectable amount of indoles and alcohol ketones were attained. The main products in the later three steps were nitriles and phenols at 623 K, hydrocarbons and phenols at 673 K, and only phenols at 773 K, respectively. A higher heating value (HHV) of 36.0 MJ/kg of the bio-oil was obtained at 673 K. The hydrocarbons, palmitic acid and esters in the bio-oil were derived from lipids. The phenols, indoles, pyrroles, small molecular acids, amides like acetamide and some nitriles like phenyl-acetonitrile were generated from proteins. Amides and nitriles were also dated from the interaction of pyrolytic intermediates of lipids and proteins. Fewer products directly from the direct pyrolysis of saccharides were detected in the algae bio-oil due to the interaction of pyrolytic intermediates of saccharides and proteins in algae, and those interactions resulted in the formation of oligomers in the bio-oil at 473 and 523 K. Whereas very weak interaction was observed between lipids and saccharides. The process of fractional pyrolysis by varying temperature provided an advisable way for improving the selectivity of bio-oil from direct pyrolysis, and made the bio-oil much more applicable in down streaming utilization.