193
Australia. Additional peak identification was carried out by accurate mass (<10 ppm)
matching using METLIN database, by manually interpreting the mass spectra, considering the data obtained from literature and fragmentation pattern of the compounds. Samples were analysed in triplicates and only those molecules identified in
all the three samples are reported.
9.3 Results and Discussion
In this study, the metabolite profile of Z. muelleri was analysed. A total of 63 components were identified in the GC-MS analysis accounting for >90% of the total
peak areas. This could be classified into four metabolite categories: organic acids
(40%), sugar and sugar alcohols (25%), amino acids (23%), phytosterols and others
(10%). A list of annotated molecules using GC-MS is listed in Table 9.1.
Forty-four metabolites were annotated using LC-MS analysis. Identified metabolites belonged to various classes including phenolic acids, amino acids, sugar alcohols, flavonoid glycosides/methyl ether derivatives and fatty acids. Based on
metabolite identification, LC-MS chromatogram of Z. muelleri indicated three main
regions, 0.6–1.5 min with eluted peaks linked to amino acids and sugar, 3.5–7.2 min
for organic acid and flavonoids and 11–13 min for fatty acid and derivatives. The
compounds annotated from Z. muelleri using LC-MS is summarized in Table 9.2.
9.3.1 Identification of Rosmarinic Acid and Zosteric Acid
Earlier studies have reported rosmarinic acid and zosteric acid as the marker compounds of Zostera species (Quackenbush et al. 1986). Rosmarinic acid, a dimer of
caffeic acid, was characterized by the presence of molecular ion at m/z 361.0922
and fragments at m/z 343.0815 (M + H-18, loss of water molecule), 181.0494
(M + H-180, loss of caffeic acid and m/z 163.088 (M + H-198, loss of R(+)-β-(3,4dihydroxyphenyl) lactic acid) (Barros et al. 2013; Taamalli et al. 2015). The molecular ion peak at m/z 245.0110 (Rt 3.48 min) was assigned to zosteric acid. Targeted
QTOF analysis in negative ion mode showed product ions at m/z 198.991 and
163.038 corresponding to loss of carboxylic acid (−COOH) and sulphonic acid (−
SO 3 H), respectively, thus indicating the peak as zosteric acid.
9.3.2 Profile of Z. muelleri
In the last few decades, considerable progress has made in identifying secondary
metabolites in seagrass using targeted and/or non-targeted metabolomic approaches.
Several studies have demonstrated the high existence of natural secondary
9 Gas and Liquid Chromatography-Mass Spectrometry-Based Metabolic Profiling…
Australia. Additional peak identification was carried out by accurate mass (<10 ppm)
matching using METLIN database, by manually interpreting the mass spectra, considering the data obtained from literature and fragmentation pattern of the compounds. Samples were analysed in triplicates and only those molecules identified in
all the three samples are reported.
9.3 Results and Discussion
In this study, the metabolite profile of Z. muelleri was analysed. A total of 63 components were identified in the GC-MS analysis accounting for >90% of the total
peak areas. This could be classified into four metabolite categories: organic acids
(40%), sugar and sugar alcohols (25%), amino acids (23%), phytosterols and others
(10%). A list of annotated molecules using GC-MS is listed in Table 9.1.
Forty-four metabolites were annotated using LC-MS analysis. Identified metabolites belonged to various classes including phenolic acids, amino acids, sugar alcohols, flavonoid glycosides/methyl ether derivatives and fatty acids. Based on
metabolite identification, LC-MS chromatogram of Z. muelleri indicated three main
regions, 0.6–1.5 min with eluted peaks linked to amino acids and sugar, 3.5–7.2 min
for organic acid and flavonoids and 11–13 min for fatty acid and derivatives. The
compounds annotated from Z. muelleri using LC-MS is summarized in Table 9.2.
9.3.1 Identification of Rosmarinic Acid and Zosteric Acid
Earlier studies have reported rosmarinic acid and zosteric acid as the marker compounds of Zostera species (Quackenbush et al. 1986). Rosmarinic acid, a dimer of
caffeic acid, was characterized by the presence of molecular ion at m/z 361.0922
and fragments at m/z 343.0815 (M + H-18, loss of water molecule), 181.0494
(M + H-180, loss of caffeic acid and m/z 163.088 (M + H-198, loss of R(+)-β-(3,4dihydroxyphenyl) lactic acid) (Barros et al. 2013; Taamalli et al. 2015). The molecular ion peak at m/z 245.0110 (Rt 3.48 min) was assigned to zosteric acid. Targeted
QTOF analysis in negative ion mode showed product ions at m/z 198.991 and
163.038 corresponding to loss of carboxylic acid (−COOH) and sulphonic acid (−
SO 3 H), respectively, thus indicating the peak as zosteric acid.
9.3.2 Profile of Z. muelleri
In the last few decades, considerable progress has made in identifying secondary
metabolites in seagrass using targeted and/or non-targeted metabolomic approaches.
Several studies have demonstrated the high existence of natural secondary
9 Gas and Liquid Chromatography-Mass Spectrometry-Based Metabolic Profiling…