Limonium Tetragonum Research Articles

Development and characterization of expressed sequence tag-simple sequence repeat markers for the near-threatened halophyte, Limonium tetragonum (Thunb.) A. A. Bullock (Plumbaginaceae).

Here, next-generation RNA sequencing analysis was performed to develop 13 novel expressed sequence tag-simple sequence repeat markers to evaluate the genetic variation in the near-threatened halophyte, Limonium tetragonum (Thunb.) A. A. Bullock. In the four populations, the total number of alleles at each locus ranged from two to seven, with an average of 3.1 and average observed and expected heterozygosities ranged from 0.00 to 0.13 and 0.28 to 0.78, respectively. Three of thirteen loci had the same homo alleles within populations, however, different alleles among different populations. Compared to other halophytes, relatively low genetic diversity was observed in this species. Further studies are necessary to determine the population demography of L. tetragonum and clarify the cause of its low genetic diversity.

Effect of NaCl concentration on the growth and phytochemicals of hydroponically grown Limonium tetragonum in a vertical farm

Effect of NaCl concentration on the growth and phytochemicals of hydroponically grown <i>Limonium tetragonum</i> in a vertical farm

Physiological and biochemical responses of Limonium tetragonum to NaCl concentrations in hydroponic solution.

Limonium (L.) tetragonum (Thunb.) A. A. Bullock, a halophyte that grows all over the southwest coast of Korea, is a medicinal plant with various pharmacological effects. The salt defense mechanism stimulates the biosynthesis of various secondary metabolites and improves functional substances. In this study, we investigated the optimal NaCl concentration for the growth and enhancement of secondary metabolites in hydroponically grown L. tetragonum. The seedlings grown for 3 weeks in a hydroponic cultivation system were treated with 0-, 25-, 50-, 75-, and 100-mM NaCl in Hoagland's nutrient solution for 8 weeks. No significant effect on the growth and chlorophyll fluorescence was observed for the NaCl concentrations below 100-mM. The increase in the NaCl concentration resulted in the decrease in the water potential of the L. tetragonum leaves. The Na+ content accumulated in the aerial part increased rapidly and the content of K+, which acts as an antagonist, decreased with the increase in NaCl concentrations in hydroponics. The total amino acid content of L. tetragonum decreased compared to the 0-mM NaCl, and most of the amino acid content decreased as the NaCl concentration increased. In contrast, the content of urea, proline (Pro), β-alanine, ornithine, and arginine was increased with an increase in NaCl concentration. The Pro content at 100-mM NaCl accounted for 60% of the total amino acids and was found to be a major osmoregulator as an important component of the salt defense mechanisms. The top five compounds identified in the L. tetragonum were classified as flavonoids while the flavanone compound was detected only in the NaCl treatments. A total of four myricetin glycosides were increased in comparison to the 0-mM NaCl. Among the differentially expressed genes, a significantly large change in Gene ontology was seen in the circadian rhythm. NaCl treatment enhanced the flavonoid-based substances of L. tetragonum. The optimum NaCl concentration for the enhancement of secondary metabolites of the L. tetragonum in the vertical farm-hydroponic cultivation system was 75-mM NaCl.

Chemical Constituents, Anticancer and Anti-Proliferative Potential of Limonium Species: A Systematic Review.

Limonium species represent a source of bioactive compounds that have been widely used in folk medicine. This study aimed to synthesize the anticancer and anti-proliferative potential of Limonium species through a systematic review. Searches were performed in the electronic databases PubMed/MEDLINE, Scopus, and Scielo and via a manual search. In vivo or in vitro studies that evaluated the anticancer or anti-proliferative effect of at least one Limonium species were included. In total, 942 studies were identified, with 33 articles read in full and 17 studies included for qualitative synthesis. Of these, 14 (82.35%) refer to in vitro assays, one (5.88%) was in vivo, and two (11.76%) were designed as in vitro and in vivo assays. Different extracts and isolated compounds from Limonium species were evaluated through cytotoxic analysis against various cancer cells lines (especially hepatocellular carcinoma-HepG2; n = 7, 41.18%). Limonium tetragonum was the most evaluated species. The possible cellular mechanism involved in the anticancer activity of some Limonium species included the inhibition of enzymatic activities and expression of matrix metalloproteinases (MMPs), which suggested anti-metastatic effects, anti-melanogenic activity, cell proliferation inhibition pathways, and antioxidant and immunomodulatory effects. The results reinforce the potential of Limonium species as a source for the discovery and development of new potential cytotoxic and anticancer agents. However, further studies and improvements in experimental designs are needed to better demonstrate the mechanism of action of all of these compounds.

Limonium tetragonum Promotes Running Endurance in Mice through Mitochondrial Biogenesis and Oxidative Fiber Formation.

The purpose of this study was to examine whether Limonium tetragonum, cultivated in a smart-farming system with LED lamps, could increase exercise capacity in mice. C57BL/6 male mice were orally administered vehicle or Limonium tetragonum water extract (LTE), either 30 or 100 mg/kg, and were subjected to moderate intensity treadmill exercise for 4 weeks. Running distance markedly increased in the LTE group (100 mg/kg) by 80 ± 4% compared to the vehicle group, which was accompanied by a higher proportion of oxidative fibers (6 ± 6% vs. 10 ± 4%). Mitochondrial DNA content and gene expressions related to mitochondrial biogenesis were significantly increased in LTE-supplemented gastrocnemius muscles. At the molecular level, the expression of PGC-1α, a master regulator of fast-to-slow fiber-type transition, was increased downstream of the PKA/CREB signaling pathway. LTE induction of the PKA/CREB signaling pathway was also observed in C2C12 cells, which was effectively suppressed by PKA inhibitors H89 and Rp-cAMP. Altogether, these findings indicate that LTE treatment enhanced endurance exercise capacity via an improvement in mitochondrial biosynthesis and the increases in the formation of oxidative slow-twitch fibers. Future study is warranted to validate the exercise-enhancing effect of LTE in the human.

Endophytic Fungi of Salt-Tolerant Plants: Diversity and Ability to Promote Plant Growth.

Suaeda australis, Phragmites australis, Suaeda maritima, Suaeda glauca Bunge, and Limonium tetragonum in the Seocheon salt marsh on the west coast of the Korean Penincula were sampled in order to identify the endophytes inhabiting the roots. A total of 128 endophytic fungal isolates belonging to 31 different genera were identified using the fungal internal transcribed spacer (ITS) regions and the 5.8S ribosomal RNA gene. Fusarium, Paraconiothyrium and Alternaria were the most commonly isolated genera in the plant root samples. Various diversity indicators were used to assess the diversity of the isolated fungi. Pure cultures containing each of the 128 endophytic fungi, respectively, were tested for the plant growth-promoting abilities of the fungus on Waito-C rice germinals. The culture filtrate of the isolate Lt-1-3-3 significantly increased the growth of shoots compared to the shoots treated with the control. Lt-1-3-3 culture filtrate was analyzed and showed the presence of gibberellins (GA1 2.487 ng/ml, GA3 2.592 ng/ml, GA9 3.998, and GA24 6.191 ng/ml). The culture filtrate from the Lt-1-3-3 fungal isolate produced greater amounts of GA9 and GA24 than the wild-type Gibberella fujikuroi, a fungus known to produce large amounts of gibberellins. By the molecular analysis, fungal isolate Lt-1-3-3 was identified as Gibberella intermedia, with 100% similarity.

The complete chloroplast genome of Limonium tetragonum (Plumbaginaceae) isolated in Korea

The chloroplast genome of Limonium tetragonum (Thunb.) Bullock, a halophytic species, was sequenced to understand genetic differences based on its geographical distribution. The cp genome of L. tetragonum was 154,689 bp long (GC ratio is 37.0%) and has four subregions: 84,572 bp of large single-copy (35.3%) and 12,813 bp of small singlecopy (31.5%) regions were separated by 28,562 bp of inverted repeat (40.9%) regions. It contained 128 genes (83 proteincoding genes, eight rRNAs, and 37 tRNAs). Thirty-five single-nucleotide polymorphisms and 33 INDEL regions (88 bp in length) were identified. Maximum-likelihood and Bayesian inference phylogenetic trees showed that L. tetragonum formed a sister group with L. aureum, which is incongruent with certain previous studies, including a phylogenetic analysis.

Two new records of powdery mildews (Erysiphaceae) from Japan: Erysiphe actinidiicola sp. nov. and Erysiphe sp. on Limonium tetragonum.

Erysiphe actinidiicola on Actinidia polygama is described based on morphological and molecular data. Erysiphe actinidiicola is distinguished from E. actinidiae var. actinidiae by having irregularly to dichotomously branched chasmothecial appendages, larger chasmothecia sizes and numbers of asci per chasmothecium. Molecular analyses indicated that this species forms a clade separated from E. actinidiae var. actinidiae. An epitype is proposed for E. actinidiae var. actinidiae with ex-epitype sequences. A powdery mildew found on Limonium tetragonum is tentatively described as Erysiphe sp. This species is distinguished from E. limonii, a powdery mildew on Limonium spp., based on the DNA sequence differences in the 28S rDNA and internal transcribed spacer region as well as the morphological differences in the length of the conidiophores. This is the first record of powdery mildew on L. tetragonum in the world.

Protective effects of halophyte complex extract against UVB-induced damage in human keratinocytes and the skin of hairless mice.

Limonium tetragonum, Triglochin maritimum, Artemisia scoparia and red ginseng have been used as folk remedies for treating a variety of diseases. In the current study, the protective effects of halophyte and red ginseng against ultraviolet (UV)-induced skin damage were investigated. Halophyte red ginseng complex extract (HRCE) was prepared and its effects on UV-B irradiated human keratinocytes and mouse skin were studied through ELISA, Western blotting immunofluorescence and histological staining. HRCE inhibited peroxide-induced damage in human keratinocytes. HRCE also inhibited UVB-induced collagen and elastin degradation in human keratinocytes and mouse skin. In addition, HRCE inhibited mast cell infiltration in the skin of mice irradiated with UVB light. This effect was likely due to HRCE inhibiting the activation of MAPK and NF-κB. By protecting the skin from UVB-induced skin damage, HRCE has the potential to be used in the treatment and prevention of UV-induced skin damage and photoaging.

Diversity and Plant Growth Promotion of Fungal Endophytes in Five Halophytes from the Buan Salt Marsh.

The diversity and plant growth-promoting ability of fungal endophytes that are associated with five halophytic plant species (Phragmites australis, Suaeda australis, Limonium tetragonum, Suaeda glauca Bunge, and Suaeda maritima) growing in the Buan salt marsh on the west coast of South Korea have been explored. About 188 fungal strains were isolated from these plant samples' roots and were then studied with the use of the internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2). The endophytic fungal strains belonged to 33 genera. Alternaria (18%) and Fusarium (12.8%), of the classes Dothideomycetes and Sordariomycetes, were most rampant in the coastal salt marsh plants. There was a higher diversity in fungal endophytes that are isolated from S. glauca Bunge than in isolates from other coastal salt marsh plants. Plant growth-promoting experiments with the use of Waito-C rice seedlings show that some of the fungal strains could encourage a more efficient growth than others. Furthermore, gibberellins (GAs) GA1, GA3, and GA9 were seen in the Sa-1-4-3 isolate (Acrostalagmus luteoalbus) culture filtrate with a gas chromatography/mass spectrometry.

Importance of Biotic and Soil Factors in Determining the Distribution Strategies of Coastal Salt Marsh Plants

The distribution of plant communities in the salt marshes of the southwestern coasts of South Korea was studied, along with environmental or plant factors, by canonical correspondence analysis (CCA) and the competitor (C), stress tolerator (S), and ruderal (R) (CSR) ecological strategies. The coastal salt-marsh plants were classified into three plant-factor groups in the CCA biplot diagram. Group 1 was correlated with LS and FP. Group 2 was correlated with CH and SLA, and Group 3 was correlated with LA, LDMC and LDW. The salt-marsh plants were classified into four soil-factor groups in the CCA biplot diagram. First, the group factor was correlated with TN, TOC, and Ca2+. Second, the group factor was distributed according to Mg2+, soil texture as Clay and Silt. Third, the group factor was distributed according to Salinity and Na+ content. Fourth, the group factor was distributed according to Sand content. To clarify the relative significance of competition, stress, and disturbance in the distribution process of plant communities, the CSR distribution model was adopted. The nine species showed CR (competitor-ruderal) strategies: Artemisia fukudo, Artemisia scoparis, Aster tripolium, Atriplex gmelinii, Imperata cylindrica var. koenigii, Salicornia europaea, Suaeda japonica, and Suaeda maritima. The four species with C (competitor) strategies were Artemisia capillaris, Limonium tetragonum, Triglochin maritimum, and Zoysia sinica. Carex scabrifolia and Phragmites communis displayed SC (stress-tolerant-competitor). Both distribution patterns of the CCA diagrams and CSR triangles may provide a useful scientific basis for protecting and restoring salt marshes and their valuable ecosystem services, considering the increasing disturbances.

Diversity and Plant Growth-Promoting Effects of Fungal Endophytes Isolated from Salt-Tolerant Plants.

Fungal endophytes are symbiotic microorganisms that are often found in asymptomatic plants. This study describes the genetic diversity of the fungal endophytes isolated from the roots of plants sampled from the west coast of Korea. Five halophytic plant species, Limonium tetragonum, Suaeda australis, Suaeda maritima, Suaeda glauca Bunge, and Phragmites australis, were collected from a salt marsh in Gochang and used to isolate and identify culturable, root-associated endophytic fungi. The fungal internal transcribed spacer (ITS) region ITS1-5.8S-ITS2 was used as the DNA barcode for the classification of these specimens. In total, 156 isolates of the fungal strains were identified and categorized into 23 genera and two phyla (Ascomycota and Basidiomycota), with Dothideomycetes and Sordariomycetes as the predominant classes. The genus Alternaria accounted for the largest number of strains, followed by Cladosporium and Fusarium. The highest diversity index was obtained from the endophytic fungal group associated with the plant P. australis. Waito-C rice seedlings were treated with the fungal culture filtrates to analyze their plant growth-promoting capacity. A bioassay of the Sm-3-7-5 fungal strain isolated from S. maritima confirmed that it had the highest plant growth-promoting capacity. Molecular identification of the Sm-3-7-5 strain revealed that it belongs to Alternaria alternata and is a producer of gibberellins. These findings provided a fundamental basis for understanding the symbiotic interactions between plants and fungi.

Talaromyces halophytorum sp. nov. Isolated from Roots of Limonium tetragonum in Korea

Talaromyces halophytorum sp. nov. was isolated from the roots of halophyte Limonium tetragonum collected from Seocheon-gun, Korea in November 2015. It showed a slow growth on yeast extract sucrose agar at 25 °C, no growth at 4 °C or 37 °C and produced smooth-walled and globose to sub-globose conidia. T. halophytorum is phylogenetically distinct from the other reported Talaromyces species of section Trachyspermi based on multi-locus sequence typing results using partial fragments of β-tubulin, calmodulin, ITS, and RNA polymerase II genes.

Canonical Correspondence Analysis Ordinations and Competitor, Stress Tolerator, and Ruderal Strategies of Coastal Dune Plants in South Korea

Lee, J.-S.; Son, D.-H.; Lee, S.-H.; Myeong, H.-H.; Cho, J.-S.; Lee, J.-C.; Lee, J.-Y.; Park, C.-S., and Kim, J.-W., 2020. Canonical correspondence analysis ordinations and competitor, stress tolerator, and ruderal strategies of coastal dune plants in South Korea. Journal of Coastal Research, 36(3), 528–535. Coconut Creek (Florida), ISSN 0749-0208.The distribution of plant communities in the sand dunes of the SW coasts of South Korea was studied, along with environmental factors and plant traits, by canonical correspondence analysis (CCA). The competitor, stress tolerator, and ruderal (CSR) ecological strategies were also evaluated. The coastal sand dune plants were classified into two plant trait groups in the CCA biplot diagram. First, vegetation was correlated with leaf dry weight, canopy height, specific leaf area, leaf dry matter content, and lateral spread. Second, it was correlated with the flowering period and flowering start. Coastal sand dune plants were classified into three soil factor groups in axes 1 and 2 of the CCA biplot diagram. First, the vegetation was correlated with total nitrogen (T-N), K+, Silt, Mg2+, Na+, and clay contents. Second, it was correlated with Ca2+. Third, it was correlated with the sand contents. To elucidate the relative significance of competition, stress, and disturbance in the distribution process of plant communities, the CSR distribution model was adopted. Many coastal plants (12 species) showed competitor-ruderal/competitor-stress-tolerant-ruderal strategies: Artemisia fukudo, Atriplex gmelinii, Carex kobomugi, Calystegia soldanella, Digitaria ciliaris, Linaria japonica, Messerschmidia sibirica, Oenothera bieenis, Salicornia europaea, Salsola komarovii, Suaeda glauca, and Suaeda maritima. The four species with stress-tolerant-competitor/competitor-stress-tolerant-ruderal strategies were Carex pumila, Imperata cylindrica var. koenigii, Limonium tetragonum, and Zoysia sinica. Conyza canadensis, Ischaemum anthephoroides, Phragmites communis, and Vitex rotundifolia displayed competitor/stress-tolerant-competitor, competitor-ruderal, stress-tolerant-competitor, and C/SC strategies, respectively. The differences in distribution and restoration patterns of the CCA diagrams and CSR triangles may be attributable to different adaptions of plant traits or soil factors.

The Standardized Extract of Limonium tetragonum Alleviates Chronic Alcoholic Liver Injury in C57Bl/6J Mice.

Background:In traditional folk medicine, Limonium tetragonum is used in the treatment of uterine hemorrhage, tinnitus, and oligomenorrhea.Objective:This study aimed to identify the therapeutic effect of L. tetragonum EtOAc extract (EALT) on liver of mice with chronic alcohol poisoning.Materials and Methods:C57BL/6J mice were administered 100 mg/kg of EALT with a single binge ethanol/Lieber-DeCarli liquid diet for 8 weeks.Results:The chronic-binge ethanol diet induced a significant increase in liver marker enzyme activities. Coadministration of EALT reversed the elevation of serum total cholesterol and triglyceride as well as aspartate aminotransferase and alanine aminotransferase due to chronic alcohol consumption. Histologic findings including markedly attenuated fat accumulation in hepatocytes were observed in EALT-treated mice. EALT supplementation prevented alcoholic liver injury through attenuation of inflammatory mediators such as toll-like receptor-4, cytochrome P4502E1, and cyclooxygenase-2, and inflammatory cytokine interleukin-6.Conclusion:Results provided direct experimental evidence for the hepatoprotective effect of EALT in the NIAAA mouse model. Therapeutic attempts with the L. tetragonum extract might be useful in the management of alcoholic liver disease.SUMMARY Halophyte Limonium tetragonum has recently been of interest in Korea for its nutritional value and salty taste which made it an ideal vegetablePhytochemical analysis of L. tetragonum EtOAc extract (EALT) resulted in nine compounds including catechins and myricetin glycosides as main componentsAdministration of EALT for 8 weeks showed hepatoprotective effect on Lieber-DeCarli diet-fed mouse modelA significant decrease in liver marker enzymes and inflammatory mediators was also detected. Abbreviations used: EALT: L. tetragonum EtOAc extract; TC: Total cholesterol; TG: Triglyceride; ROS: Reactive oxygen species; CYP2E1: Cytochrome P4502E1; TLR-4: Toll-like receptor-4; COX-2: Cyclooxygenase-2.

Comparison of Biological Activities of Korean Halophytes

Halophytes are expected to possess abundant secondary metabolites and various biological activities because of habitat in extreme environments. In this study, we collected 14 halophytes (Asparagus oligoclonos, Calystegia soldanella, Carex pumila, Chenopodium glaucum, Elymus mollis, Glehnia littoralis, Limonium tetragonum, Messerschmidia sibirica, Rosa rugosa, Salsola komarovii, Spergularia marina, Suaeda glauca, Suaeda maritima, and Vitex rotundifolia) native to Korea and compared their total polyphenol contents, antioxidant and anti-inflammatory activities. The total polyphenol contents of R. rugosa (27.28%) and L. tetragonum (13.17%) were significantly higher than those of the other 12 halophytes and L. tetragonum, R. rugosa, and M. sibirica showed significantly greater antioxidant activities than the other 11 halophytes, as determined by DPPH (2,2-diphenyl-1-picrylhydrazyl). A. oligoclonos, E. mollis, and C. pumila showed significantly greater anti-inflammatory activities than the other 11, as determined by NO (Nitric oxide) and PGE2 (Prostaglandin E2) levels. In contrast, these three extracts had normal and low total polyphenol contents among the 14 halophytes. Consequently, the total polyphenol content in the 14 studied halophytes appeared to be related to antioxidant, but not anti-inflammatory activity levels.

Antioxidant and Anti-inflammatory Activity of Six Halophytes in Korea

The aim of this study was to measure and compare polyphenol content, antioxidant, and antiinflammatory activity of six halophytes (Limonium tetragonum, Suaeda glauca, Suaeda japonica, Salicornia europaea, Triglochin maritimum, and Sonchus brachyotus). Depending on the total polyphenol content, the plants were categorized into two groups: (1) a high total polyphenol content group that included L. tetragonum, S. brachyotus, and S. europaea, and, (2) a low total polyphenol content group consisting of S. glauca, T. maritima, and S. japonica. Antioxidant activity was evaluated using DPPH and hydroxyl radical scavenging assays, and by measuring ROS. Anti-inflammatory activity was evaluated by measuring NO and PGE2. L. tetragonum and S. brachyotus, that have high polyphenol content, also showed strong antioxidant activity. In addition, L. tetragonum, S. brachyotus, and S. europaea showed good anti-inflammatory activity. Consequently, the total polyphenol content was thought to be related to antioxidant and anti-inflammatory activity. Therefore, S. brachyotus and L. tetragonum are good candidates for use in pharmaceuticals and functional foods.

염생식물 갯질경 추출물의 미백 활성

To identify the natural and novel cosmeceutical ingredients which have whitening function, we examined the effects of tyrosinase activity and melanin production on halophyte Limonium tetragonum (Thunb.) A. A. Bullock extracts. As a result, the extracts showed whitening effect with no cytotoxicity. Melanin contents of B16F10 melanoma cells were decreased in a dose-dependent manner at 50, 100, 200 μg/mL treatment of the extracts. In tyrosinase activity inhibition test, L. tetragonum (Thunb.) A. A. Bullock extracts showed decreased tyrosinase activity as the concentration of α-MSH was increased. Furthermore, it was observed that the tyrosinase and MITF expression was significantly downregulated by adding L. tetragonum (Thunb.) A. A. Bullock extracts in α-MSH, a melanogenesis inducing material, treated B16F10 melanoma cells. These results indicate that L. tetragonum (Thunb.) A. A. Bullock extract might be an effective whitening agent by inhibit melanin formation.

Anti-Melanogenic Effects of Flavonoid Glycosides from Limonium tetragonum (Thunb.) Bullock via Inhibition of Tyrosinase and Tyrosinase-Related Proteins.

Overproduction and stimulation of tyrosinase result in increased melanogenesis of which several skin disorders such as freckles, spots, and hyperpigmentation appear as complications. Limonium tetragonum is a halophyte well-known for its antioxidative properties. This study investigated the anti-melanogenic effects of solvent-partitioned L. tetragonum extracts (LTEs) and its bioactive constituents, two isolated flavonoid glycosides. Current study followed a set of experiments on B16-F10 mouse melanoma cell model with a focus on tyrosinase activity and production. The anti-melanogenic capacity of LTEs was confirmed by their tyrosinase inhibitory effects, prevention of DOPA oxidation, and suppression of melanin production. The inhibition of tyrosinase and DOPA oxidation by LTEs was suggested to be related with the downregulation of microphthalmia-associated transcription factor, tyrosinase, tyrosinase-related protein-1, and tyrosinase-related protein-2, verified with mRNA and protein expression levels. Among all tested LTEs, 85% aq. MeOH and n-BuOH were found to be the most active fractions which later yielded the two known compounds, myricetin 3-galactoside and quercetin 3-O-β-galactopyronaside. The anti-melanogenic potential of the compounds were confirmed by their tyrosinase inhibitory effects. These results suggested that L. tetragonum may serve as a potential source of bioactive substances with effective anti-melanogenesis properties.

Identification of Hepatoprotective Constituents in Limonium tetragonum and Development of Simultaneous Analysis Method using High-performance Liquid Chromatography.

Background:Limonium tetragonum, a naturally salt-tolerant halophyte, has been studied recently and is of much interest to researchers due to its potent antioxidant and hepatoprotective activities.Objective:In the present study, we attempted to elucidate bioactive compounds from ethyl acetate (EtOAc) soluble fraction of L. tetragonum extract. Furthermore, the simultaneous analysis method of bioactive EtOAc fraction of L. tetragonum has been developed using high-performance liquid chromatography (HPLC).Materials and Methods:Thirteen compounds have been successfully isolated from EtOAc fraction of L. tetragonum, and the structures of 1–13 were elucidated by extensive one-dimensional and two-dimensional spectroscopic methods including 1H-NMR, 13C-NMR, 1H-1H COSY, heteronuclear single quantum coherence, heteronuclear multiple bond correlation, and nuclear Overhauser effect spectroscopy. Hepatoprotection of the isolated compounds against liver fibrosis was evaluated by measuring inhibition on hepatic stellate cells (HSCs) undergoing proliferation.Results:Compounds 1–13 were identified as gallincin (1), apigenin-3-O-β-D-galactopyranoside (2), quercetin (3), quercetin-3-O-β-D-galactopyranoside (4), (−)-epigallocatechin (5), (−)-epigallocatechin-3-gallate (6), (−)-epigallocatechin-3-(3″-O-methyl) gallate (7), myricetin-3-O-β-D-galactopyranoside (8), myricetin-3-O-(6″-O-galloyl)-β-D-galactopyranoside (9), myricetin-3-O-α-L-rhamnopyranoside (10), myricetin-3-O-(2″-O-galloyl)-α-L-rhamnopyranoside (11), myricetin-3-O-(3″-O-galloyl)-α-L-rhamnopyranoside (12), and myricetin-3-O-α-L-arabinopyranoside (13), respectively. All compounds except for 4, 8, and 10 are reported for the first time from this plant.Conclusion:Myricetin glycosides which possess galloyl substituent (9, 11, and 12) showed most potent inhibitory effects on the proliferation of HSCs.SUMMARYIn the present study, we have successfully isolated 13 compounds from bioactive fraction of Limonium tetragonum. The structures of compounds isolated have been fully elucidated, and hepatoprotective activities of compounds against liver fibrosis were evaluated by measuring inhibition on hepatic stellate cells undergoing proliferation. Furthermore, the simultaneous analysis method of bioactive ethyl acetate fraction of L. tetragonum has been developed using HPLC. Ten compounds identified herein are reported for the first time from this plant.Abbreviations used: HSQC: Heteronuclear single quantum coherence; HMBC: Heteronuclear multiple bond correlation; NOESY: Nuclear Overhauser effect spectroscopy; EGCG: Epigallocatechin-3-gallate; EGC: Epigallocatechin; HSC: Hepatic stellate cell; MTT: 3-(4,5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide.