Distribution of betaine lipids in 14 species of microalgae
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摘要: 采用超高效液相色谱−四极杆−飞行时间质谱联用分析系统(UPLC -Q- TOF MS)，对4个微藻门中的14种微藻的甜菜碱脂的分子组成及其相应脂肪酸的组成及分布进行分析。结果表明：在14种微藻中共鉴定出133种甜菜碱脂，包括53种DGCC、41种DGTS和39种DGTA。其中甲藻和定鞭藻中主要的甜菜碱脂种类为DGCC，绿藻中主要的甜菜碱脂种类为DGTS；而在硅藻中的甜菜碱脂主要包括两种：中心硅藻纲中为DGCC，羽纹硅藻纲中为DGTA。此外，不同微藻中甜菜碱脂脂肪酸组成差异仅限制在门或纲的水平上，在较低的分类水平上差异不明显。硅藻门、甲藻门和定鞭藻门中均含有DGCC，其脂肪酸链的组成均含有C14−C18的脂肪酸以及C20和C22的多不饱和脂肪酸，但是甲藻的C14−C18脂肪酸链为饱和的，而C19奇数碳原子的脂肪酸链仅在硅藻中发现。认为海洋微藻甜菜碱脂的研究可以为海洋微藻化学分类学以及缺磷极性脂类的生理、生态作用和功能特性研究提供重要参考依据。
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关键词:
- 海洋微藻 /
- 甜菜碱脂 /
- 超高效液相色谱−四极杆−飞行时间质谱
Abstract: The qualitative and quantitative analyses on betaine lipids of microalgae were conducted by means of the quadrupole flight time ultra-high performance liquid chromatography tandem mass spectrometry (UPLC-Q-TOF-MS) using electron spraying ionization. As a result, 133 betaine lipids including 53 DGCC, 41 DGTS and 39 DGTA were identified from 14 species of microalgae in four major phyla. DGCC was the dominant betaine lipid in dinoflagellates and haptophytes, while DGTS was the main betaine lipid in chlorophytes. There were two main types of betaine lipid in diatoms: DGCC in centric diatom and DGTA in pennate diatom. The differences of fatty acid of betaine lipid in marine microalgae were only observed at the phyla and class levels, but at lower taxonomic levels the differences were less stable. DGCC existed in diatoms, dinoflagellates and chlorophytes, which contained C14−C18 saturated and unsaturated fatty acids and C20 and C22 polyunsaturated fatty acids. The difference in DGCC is that saturated C14−C18 fatty acids existed in dinoflagellates, while the fatty acids of odd carbon numbers C19 occurred only in diatoms. The results of description and molecular characterization of betaine lipid in microalgae can be served in future investigation in chemotaxonomy, physiology, ecological role of microalgae and functional properties of these phosphorous-lacking polar lipids.-
Key words:
- microalgae /
- betaine lipids /
- UPLC-Q-TOF-MS
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图 1 正离子MSE模式，假微型海链藻中16:0/22:6 DGCC ([M+H]+，m/z=800.61)低能量下提取离子流色谱图 （a）；正离子模式下假微型海链藻保留时间为12.98 min16:0/22:6 DGCC [M+H]+（m/z=800.61）二级质谱图（b）
Fig. 1 EIC of 16:0/22:6 DGCC ([M+H]+, m/z= 800.61) in T. pseudonana at the low collision energy of 6 V in the positive mode with the MSE technique (a); the MS2 spectrum of the [M+H]+ ion at m/z=800.61 of 16:0/22:6 DGCC at retention time 12.98 min extracted from T. pseudonana (b)
图 2 正离子MSE模式，颗石藻总脂混合物的低能量总离子流色谱图（a）；高能量MSE色谱图中提取DGCC特征碎片离子m/z=104.11的离子流色谱图（b）
Fig. 2 TIC of the lipid mixture extracted from P. carterae at the low collision energy of 6 V in the positive mode with the MSE technique (a); EIC of m/z=104.11 for characteristic product ion of DGCC in P. carterae from high collision energy scans (ramp of 30–60 V) in the positive mode with the MSE technique (b)
表 1 14种海洋微藻的种名和种质编号
Tab. 1 The strain number and species name of 14 species of microalgae
序号 分类 种质编号 中文名称 拉丁名称 1 甲藻门 NMBjah045 东海原甲藻 Prorocentrum donghaiense 2 NMBjah047-1 卡罗藻 Karlodinum veneficum 3 NMBjah043 微小亚历山大藻 Alexandrium minutum 4 绿藻门 NMBluh015-1 小球藻 Chlorella sp. 5 NMBluh014 微绿球藻 Nannochloropsis oceanica 6 NMBluh014-1 云微藻 Nannochloris sp. 7 定鞭藻门 NMBjih026-1 颗石藻 Pleurochrysis carterae 8 NMBjih022-2 湛江等鞭金藻 Isochrysis zhanjiangensis 9 NMBjih021-2 球等鞭金藻 Isochrysis galbana 10 硅藻门 NMBguh003-4 角毛藻 Chaetoceros sp. 11 NMBguh005 假微型海链藻 Thalassiosira pseudonana 12 NMBguh021 威氏海链藻 Conticribra weissflogii 13 NMBguh001 三角褐指藻 Phaeodactylum tricornutum 14 NMBguh002 小新月菱形藻 Nitzschia closterium f. minutissima 
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表 2 9种海洋微藻中DGCC的组成及含量（nmol/mg干藻粉）
Tab. 2 The composition and content of DGCC in 9 microalgae（nmol/mg dry microalgae）
组成 [M+H]+ sn-1/sn-2 硅藻门 甲藻门 定鞭藻门 假微型海链藻 威氏海链藻 角毛藻 东海原甲藻 卡罗藻 微小亚历山大藻 颗石藻 湛江等鞭金藻 球等鞭金藻 DGCC-1 462.34 14:0 0.026±0.002 0.057±0.008 0.017±0.001 0.013±0.003 0.007±0.002 0.010±0.002 0.059±0.010 DGCC-2 486.35 16:2 0.004±0.001 DGCC-3 488.36 16:1 0.017±0.002 0.474±0.031 DGCC-4 490.37 16:0 0.055±0.004 0.219±0.027 0.139±0.014 0.125±0.031 0.075±0.015 1.053±0.080 0.025±0.003 DGCC-5 504.39 17:0 0.016±0.004 DGCC-6 510.34 18:4 0.077±0.005 DGCC-7 514.37 18:2 0.191±0.019 0.059±0.003 DGCC-8 516.39 18:1 0.009±0.001 0.153±0.016 0.096±0.006 0.133±0.011 DGCC-9 518.4 18:0 0.122±0.006 DGCC-10 536.36 20:5 0.046±0.004 0.628±0.091 0.423±0.028 0.418±0.034 DGCC-11 538.37 20:4 0.218±0.009 DGCC-12 562.37 22:6 0.053±0.004 0.739±0.086 0.272±0.018 0.046±0.008 1.419±0.054 0.135±0.036 0.691±0.065 DGCC-13 672.54 14:0/14:0 0.016±0.002 0.020±0.002 0.118±0.007 DGCC-14 698.56 14:0/16:1 0.144±0.005 0.326±0.020 DGCC-15 700.53 14:0/16:0 0.027±0.007 0.025±0.003 0.281±0.020 0.047±0.005 DGCC-16 720.53 16:0/16:4 0.178±0.009 DGCC-17 720.54 14:0/18:4 0.298±0.049 0.112±0.034 0.139±0.012 DGCC-18 724.57 16:1/16:1 0.091±0.005 0.430±0.024 DGCC-19 726.59 16:0/16:1 0.175±0.009 0.205±0.020 0.421±0.016 0.030±0.005 DGCC-20 728.6 16:0/16:0 0.092±0.031 0.012±0.003 DGCC-21 746.56 16:0/18:5 0.255±0.012 DGCC-22 746.55 14:0/20:5 0.404±0.013 1.762±0.170 0.010±0.003 DGCC-23 748.57 16:0/18:4 0.103±0.004 2.990±0.059 DGCC-24 752.59 16:0/18:2 0.095±0.007 0.244±0.034 DGCC-25 754.63 16:0/18:1 0.055±0.002 0.164±0.022 DGCC-26 756.64 16:0/18:0 0.031±0.007 DGCC-27 768.54 16:3/20:5 0.072±0.005 DGCC-28 770.56 16:2/20:5 0.153±0.010 DGCC-29 772.58 16:1/20:5 0.728±0.045 0.855±0.011 1.002±0.065 DGCC-30 772.57 14:0/22:6 0.902±0.060 0.574±0.028 0.097±0.018 1.542±0.260 9.156±0.935 DGCC-31 774.59 14:0/22:5 0.017±0.002 DGCC-32 774.58 16:0/20:5 0.124±0.005 0.993±0.173 0.354±0.018 0.023±0.004 8.025±0.171 DGCC-33 776.61 16:0/20:4 0.131±0.012 0.478±0.033 DGCC-34 780.63 18:1/18:1 0.056±0.003 DGCC-35 794.55 18:4/20:5 0.142±0.009 0.224±0.005 DGCC-36 794.55 16:3/22:6 0.077±0.038 DGCC-37 796.58 18:3/20:5 0.355±0.063 DGCC-38 798.6 16:1/22:6 0.179±0.010 0.145±0.016 DGCC-39 798.57 16:0/22:5 0.073±0.014
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表 4 3种海洋微藻中甜菜碱脂的组成及含量（nmol/mg干藻粉）
Tab. 4 The composition and content of betaine lipid in 3 microalgae（nmol/mg dry microalgae）
组成 [M+H]+ sn-1/sn-2 硅藻门 甲藻门 小新月菱形藻 三角褐指藻 卡罗藻 DGTA-1 472.36 16:1 0.173±0.001 DGTA-2 496.36 18:3 0.598±0.016 DGTA-3 498.38 18:2 0.554±0.050 0.523±0.033 DGTA-4 500.39 18:1 1.744±0.129 0.686±0.040 DGTA-5 520.35 20:5 0.227±0.017 1.115±0.102 DGTA-6 522.38 20:4 0.172±0.008 1.593±0.095 DGTA-7 546.37 22:6 0.383±0.034 0.336±0.019 DGTA-8 654.53 14:0/14:1 0.146±0.004 DGTA-9 656.54 14:0/14:0 0.579±0.035 DGTA-10 682.57 14:0/16:1 0.188±0.028 0.556±0.033 DGTA-11 704.55 14:0/18:4 0.060±0.006 DGTA-12 706.57 14:0/18:3 0.408±0.023 DGTA-13 708.58 16:1/16:1 0.081±0.005 DGTA-14 708.57 14:0/18:2 0.066±0.007 0.621±0.039 DGTA-15 710.59 14:0/18:1 1.024±0.152 0.863±0.049 DGTA-16 730.57 14:0/20:5 0.268±0.031 1.260±0.106 DGTA-17 732.57 14:0/20:4 0.072±0.021 1.303±0.045 DGTA-18 734.59 14:0/20:3 0.301±0.033 DGTA-19 734.59 16:1/18:4 0.047±0.002 DGTA-20 736.62 16:1/18:1 0.361±0.011 DGTA-21 736.6 16:0/18:2 0.193±0.012 DGTA-22 738.62 16:0/18:1 0.125±0.015 0.126±0.008 DGTA-23 752.55 16:3/20:5 0.021±0.004 DGTA-24 754.56 16:2/20:5 0.026±0.003 0.014±0.002 DGTA-25 756.57 16:1/20:5 0.094±0.007 0.209±0.030 DGTA-26 756.57 14:0/22:6 0.025±0.002 0.507±0.026 DGTA-27 758.59 16:1/20:4 0.350±0.016 DGTA-28 758.6 16:0/20:5 0.187±0.014 DGTA-29 760.61 16:0/20:4 0.026±0.003 0.209±0.015 DGTA-30 764.64 18:1/18:1 0.112±0.021 DGTA-31 778.56 18:4/20:5 0.035±0.003 DGTA-32 780.58 18:3/20:5 0.030±0.001 0.431±0.032 DGTA-33 782.59 16:1/22:6 0.070±0.005 DGTA-34 784.61 18:1/20:5 0.517±0.026 0.603±0.042 DGTA-35 804.57 20:5/20:5 0.776±0.144 0.588±0.029 DGTA-36 806.59 20:4/20:5 0.078±0.013 0.358±0.039 DGTA-37 806.59 20:5/20:4 1.189±0.098 DGTA-38 808.6 20:4/20:4 0.226±0.012 DGTA-39 830.59 20:5/22:6 0.065±0.003 0.247±0.015
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表 3 8种海洋微藻中DGTS的组成及含量（nmol/mg干藻粉）
Tab. 3 The composition and content of DGTS in 8 microalgae（nmol/mg dry microalgae）
组成 [M+H]+ sn-1/sn-2 硅藻门 甲藻门 绿藻门 定鞭藻门 威氏海链藻 卡罗藻 小球藻 微绿球藻 云微藻 颗石藻 湛江等鞭金藻 球等鞭金藻 DGTS-1 472.36 16:1 0.214±0.045 0.332±0.060 DGTS-2 474.38 16:0 0.156±0.027 0.175±0.032 0.045±0.002 DGTS-3 496.36 18:3 0.191±0.034 0.215±0.005 DGTS-4 498.38 18:2 0.039±0.006 0.053±0.010 0.063±0.014 DGTS-5 520.36 20:5 1.305±0.322 2.247±0.303 DGTS-6 522.37 20:4 0.136±0.028 0.487±0.099 DGTS-7 682.57 14:0/16:1 0.434±0.082 0.105±0.007 DGTS-8 706.56 16:0/16:3 0.168±0.039 DGTS-9 708.57 14:0/18:2 0.080±0.008 DGTS-10 708.57 16:1/16:1 0.258±0.046 0.099±0.004 DGTS-11 708.59 16:0/16:2 0.077±0.013 DGTS-12 710.59 14:0/18:1 0.568±0.082 0.482±0.051 DGTS-13 710.57 16:0/16:1 0.138±0.013 1.509±0.183 0.748±0.031 DGTS-14 724.59 16:0/17:1 0.071±0.007 DGTS-15 728.55 16:3/18:3 0.035±0.006 DGTS-16 730.57 16:3/18:2 0.037±0.005 DGTS-17 730.55 14:0/20:5 0.546±0.045 0.263±0.006 DGTS-18 732.57 14:0/20:4 0.131±0.023 DGTS-19 732.58 16:3/18:1 0.045±0.010 DGTS-20 734.58 16:0/18:3 0.045±0.006 0.130±0.016 DGTS-21 734.59 18:0/16:3 0.286±0.049 DGTS-22 736.61 16:0/18:2 0.119±0.023 0.480±0.064 DGTS-23 736.6 18:0/16:2 0.091±0.013 DGTS-24 738.63 16:0/18:1 0.033±0.004 0.069±0.006 0.037±0.001 DGTS-25 754.55 16:2/20:5 0.108±0.017 DGTS-26 756.57 16:1/20:5 1.186±0.076 0.537±0.007 DGTS-27 758.59 16:0/20:5 2.622±0.278 1.683±0.050 DGTS-28 758.6 18:2/18:3 0.066±0.015 DGTS-29 760.61 18:2/18:2 0.055±0.003 DGTS-30 760.61 16:0/20:4 0.477±0.077 0.417±0.011 DGTS-31 762.62 20:0/16:3 0.402±0.055 DGTS-32 762.63 18:1/18:2 0.011±0.00 DGTS-33 764.65 18:1/18:1 0.017±0.006 0.020±0.002 DGTS-34 764.64 20:0/16:2 0.152±0.026 DGTS-35 782.59 18:2/20:5 0.340±0.056 DGTS-36 784.6 18:1/20:5 0.286±0.038 DGTS-37 784.59 16:0/22:6 0.352±0.017 DGTS-38 786.59 18:1/20:4 0.062±0.007 DGTS-39 792.68 20:0/18:2 0.041±0.003 DGTS-40 804.56 20:5/20:5 3.023±0.304 0.855±0.057 DGTS-41 806.58 20:4/20:5 1.015±0.083 0.565±0.046
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表 5 4个门的微藻中甜菜碱脂脂肪酸的组成特点
Tab. 5 Characteristics of fatty acids of betaine lipids in marine microalgae
硅藻门 甲藻门 绿藻门 定鞭藻门 DGTA 14:0, 16:0, 16:1, 16:2, 16:3, 18:1, 18:2, 18:3, 18:4, 20:3, 20:4, 20:5, 22:6 14:0, 14:1 14:0, 16:1, 18:3, 18:4, 20:5, 22:6 DGTS 14:0, 15:0, 16:0,16:1, 17:1, 18:1, 18:2 18:1, 18:2 14:0, 16:0, 16:1, 16:2, 16:3, 17:1, 18:0, 18:1, 18:2, 18:3, 20:0, 20:4, 20:5 14:0, 16:0, 16:1, 18:0, 18:1, 18:2, 18:3, 18:4, 20:5, 22:6 DGCC 14:0, 16:0, 16:1, 16:2, 16:3, 16:4, 18:1, 18:2, 18:3, 18:4, 19:0, 20:4, 20:5, 22:6 14:0, 16:0, 17:0, 18:0, 20:5, 22:6 14:0, 16:0, 16:1, 18:1, 18:3, 18:4, 18:5, 20:4, 20:5, 22:5, 22:6
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