Sequencing and Analysis of the Complete Mitochondrial Genome of Lentipes ikeae.

We sequenced and analyzed the complete mitochondrial genome of Lentipes ikeae and explored the phylogenetic relationships among Sicydiinae based on mitochondrial genome sequences. The complete mitochondrial genome sequence of L. ikeae was determined using the Illumina HiSeq X Ten sequencing platform, and the gene structural characteristics and base composition were analyzed. Based on the mitochondrial genome sequences of 28 Sicydiinae species published in GenBank and mitochondrial protein-coding genes (PCGs), Acanthogobius flavimanus (Gobionellinae) was selected as an outgroup to construct phylogenetic trees of Sicydiinae using the maximum likelihood and Bayesian inference methods. The mitochondrial genome of L. ikeae (GenBank number: OP764680) has a total length of 16,498 bp and encodes 13 PCGs, 22 transfer RNA genes, two ribosomal RNA genes, and a D-loop (control) region. Gene rearrangement is not observed. The mitochondrial genome of L. ikeae exhibits an AT preference, with AT skew > 0 and GC skew < 0 across the entire genome. The phylogenetic relationships of Sicydiinae based on 13 mitochondrial PCG sequences are Sicydium + (Stiphodon + (Sicyopus + Lentipes)) + Sicyopterus, indicating that Sicydium, Sicyopterus, Lentipes, and Stiphodon are all monophyletic groups.

Comparative Analysis and Phylogenetic Study of Dawkinsia filamentosa and Pethia nigrofasciata Mitochondrial Genomes.

Smiliogastrinae are recognized for their high nutritional and ornamental value. In this study, we employed high-throughput sequencing technology to acquire the complete mitochondrial genome sequences of Dawkinsia filamentosa and Pethia nigrofasciata. The gene composition and arrangement order in these species were similar to those of typical vertebrates, comprising 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 1 non-coding region. The mitochondrial genomes of D. filamentosa and P. nigrofasciata measure 16,598 and 16,948 bp, respectively. Both D. filamentosa and P. nigrofasciata exhibit a significant preference for AT bases and an anti-G bias. Notably, the AT and GC skew values of the ND6 gene fluctuated markedly, suggesting that the selection and mutation pressures on this gene may differ from those affecting other genes. Phylogenetic analysis, based on the complete mitochondrial genomes of 23 Cyprinidae fishes, revealed that D. filamentosa is closely related to the sister group comprising Dawkinsia denisonii and Sahyadria chalakkudiensis. Similarly, P. nigrofasciata forms a sister group with Pethia ticto and Pethia stoliczkana.

Mitogenome of a monotypic genus, Oliotius Kottelat, 2013 (Cypriniformes: Cyprinidae): Genomic characterization and phylogenetic position.

We examined the evolutionary status of the genus Oliotius Kottelat, 2013, clarified the mitogenome structure of Smiliogastrinae, and analyzed its phylogenetic information to provide a reference for the taxonomy, evolutionary genetics, and germplasm identification of Smiliogastrinae. Using next-generation sequencing, the complete mitogenome sequence of the monotypic species Oliotius oligolepis was obtained, annotated, and characterized. The mitogenomic sequences of Smiliogastrinae fish were compared and analyzed, and a phylogenetic tree was constructed. The mitogenome of O. oligolepis was 16,636 base pairs long and contained 22 tRNAs, 13 protein-coding genes, 2 rRNAs, and 1 D-loop. The Smiliogastrinae species exhibited gene arrangement consistency, with slight differences in their genome lengths and compositions. Among the 13 protein-coding genes, ND3 showed the highest polymorphism rate and largest genetic distance. Our phylogenetic analysis showed that the target species O. oligolepis is closely related to Barbodes binotatus. The genus Oliotius is monotypic, and the genus Barbodes is not supported as a monophyletic group. This molecular information provides a foundation for further research on the phylogeny of the genera Oliotius and Barbodes and provides insight into the phylogenetic features of fishes belonging to Smiliogastrinae.

Genetic population structure and demographic history of the endemic fish Paralichthys olivaceus of the Northwest Pacific Ocean.

The Northwest Pacific marginal waters comprising the South China Sea, East China Sea, Yellow Sea, and the Sea of Japan have unique geomorphic features. The Japanese flounder Paralichthys olivaceus, which is endemic to the Northwest Pacific, has high nutritional, economic, and ecological value. To allow the examination of the demographic history and population structure of the most common P. olivaceus species range over the five marginal seas (East China Sea, Yellow Sea, Bohai Sea, Northwest Pacific Ocean, and the Sea of Japan), the mitochondrial DNA control region of 91 individuals from six populations in China was sequenced. These sequences were combined with 233 sequences from four populations distributed in the Northwestern Pacific Ocean for analysis. Higher levels of nucleotide diversity (0.032 ± 0.016) and haplotype diversity (0.996 ± 0.001) were observed. The peripheral Fuqing population in the East China Sea had the relatively lowest genetic diversity and highest differentiation. Furthermore, when the results of the isolation by distance test, spatial analysis of molecular variation and geographic barrier analysis are also considered, there is a clear need to prioritize resource conservation and enhancement measures in this area. The phylogenetic trees, structure assignment test, and haplotypes network revealed no significant differences in the genealogical structure among ten populations. Mismatch distribution analysis, Bayesian skyline plots, and neutrality tests suggested that P. olivaceus experienced population expansion during the Pleistocene. Ocean currents and climate change play important roles in shaping the geographical distribution and genetic population structure of P. olivaceus.

Comparative analysis of the mitogenomes of two Corydoras (Siluriformes, Loricarioidei) with nine known Corydoras, and a phylogenetic analysis of Loricarioidei.

Corydoras is a speciose catfish genus from South America with widely investigated phylogenetic and evolutionary relationships. The complete mitogenomes of C.aeneus and C.paleatus were sequenced, assembled, and annotated using next-generation sequencing. The genome arrangements, gene contents, genome structures, base compositions, evolutionary features, codon usage, and tRNA structures of the two mitogenomes were compared and analyzed with nine published mitogenomes of Corydoras. Phylogenetic analysis was performed using concatenated nucleotide sequences with 13 protein-coding genes and two rRNAs with 44 mitogenomes of Siluriformes. These results provide information on the mitogenomes of eleven Corydoras species and evolutionary relationships within the suborder Loricarioidei, which may be applicable for further phylogenetic and taxonomic studies on Siluriformes and Loricarioidei.

Complete mitochondrial genome of Ovalona pulchella (Branchiopoda, Cladocera) as the first representative in the family Chydoridae: Gene rearrangements and phylogenetic analysis of Cladocera.

Chydoridae are phytophilic-benthic microcrustaceans that make up a significant proportion of species diversity and play an important role in the littoral zone of freshwater ecosystems worldwide. Here, we provide the complete mitochondrial genome of Ovalona pulchella (King, 1853), determined by next-generation sequencing. The entire mitochondrial genome is 15,362 bp in length; this is the first sequenced mitochondrial genome in the family Chydoridae. The base composition and codon usage were typical of Cladocera species. The mitochondrial gene arrangement (37 genes) was not consistent with that of other Branchiopoda. Both maximum likelihood and Bayesian analyses supported each suborder and family of Branchiopoda as monophyletic groups. The relationships among the families were as follows: [(Leptestheriidae + Limnadiidae) + (Sididae + (Bosminidae + (Chydoridae + Daphniidae)))] + Triopsidae. The newly sequenced O. pulchella was most closely related to the family Daphniidae. The complete mitochondrial genome of O. pulchella also provides valuable molecular information for further analysis of the phylogeny of the Chydoridae and the taxonomic status of the Branchiopoda.

Mitogenome of Knodus borki (Cypriniformes: Characidae): genomic characterization and phylogenetic analysis.

BACKGROUND: The taxonomic status of Knodu in the family Characidae is not yet clear. This study aimed to address this by sequencing and annotating Knodu borki Zarske, 2008. MATERIALS AND RESULTS: K. borki Zarske, 2008 was sequenced using a Hiseq platform and the complete mitogenome was assembled in SPAdes v3.15.2 and SOAPdenovo2 v.2.01. The mitogenome of K. borki from Guangzhou, the first sequenced species of the genus Knodu, is 16,837 bp in length and contains 13 protein-coding genes (PCGs), two ribosomal (r) RNAs, 22 transfer (t) RNAs, and one D-loop. Among these 37 genes, 28 are encoded by the heavy strand, while nine are encoded by the light strand. Twenty-one of the tRNAs can form typical cloverleaf secondary structures, except tRNA-Ser1, which lacks dihydrouridine arms. All PCGs have the same start codon (ATG), with the exception of COI (GTG). Four PCGs (ND1, ATP8, ND4L, and ND5) have TAA as the stop codon, ND6 has TAG as the stop codon, COI has AGG as the stop codon, and the remaining seven genes have incomplete stop codons of TA-/T-(ND2, COII, COIII, ND3, ND4, and Cyt b as T-, ATP6 as TA-). Phylogenetic analysis showed that K. borki belongs to the family Characidae. CONCLUSIONS: Our findings demonstrate that K. borki belongs to the family Characidae, due to consistency with the morphological identification. This study provides molecular information for further research on the phylogeny of the genus Knodus and for analyses of the taxonomic status of Characidae.

DNA barcoding of Scomberomorus (Scombridae, Actinopterygii) reveals cryptic diversity and misidentifications.

The genus Scomberomorus is economically important; however, the taxonomic status and phylogenetic relationships in this genus are not clearly resolved, making it difficult to effectively protect and exploit fish resources. To clarify the taxonomic status of Scomberomorus species, mitochondrial cytochrome c oxidase I (COI) gene sequences of 150 samples were analyzed. The average genetic distance among 14 species was approximately 11 times greater than the distances within species, in accordance with the '10× rule' of species identification. Five of the 14 species did not form monophyletic clades based on a Bayesian inference gene tree. The application of four DNA-based species delimitation methods (automatic barcode gap discovery, barcode index numbers, Poisson tree process, and the K/θ method) yielded several key results. (1) Cryptic species were detected within Scomberomoruscommerson. (2) A Scomberomorusqueenslandicus sample from Australia was misidentified as S.commerson in the Barcode of Life Data System (BOLD). (3) Specimens originally identified as Scomberomorusguttatus was differentiated into four OTUs or species, two in the Yellow, South China, and Java seas, and two in geographically distant areas, one each in the Arabian Sea and the Bay of Bengal. (4) Six specimens from South Africa originally identified as S.plurilineatus most likely do not belong to the species. (5) Specimens identified as S.maculatus and S.regalis were conspecific; however, introgression cannot be ruled out. Our findings revealed cryptic diversity and difficulties in morphological identification of species in the genus Scomberomorus. This study provides scientifically based support for the conservation of germplasm resources of the genus Scomberomorus.

Complete mitogenome of Treron sphenurus (Aves, Columbiformes): the first representative from the genus Treron, genomic comparisons and phylogenetic analysis of Columbidae.

The wedge-tailed green pigeon (Treron sphenurus) has a protective value in the evolution of the family Columbidae. In this study, the complete mitogenome of T. sphenurus from Baise City, China, which represents the first sequenced species of the genus Treron in Tribe Treronini, is reported. This was accomplished using PCR-based methods and a primer-walking sequencing strategy with genus-specific primers. The mitogenome was found to be 18,919 bp in length comprising 37 genes, including 13 protein-coding genes, two rRNA genes, 22 tRNA genes, and one control region. In terms of structure and composition, many similarities were found between the T. sphenurus and Hemiphaga novaeseelandiae (New Zealand pigeon) mitogenomes. This was further supported by phylogenetic analysis showing that T. sphenurus has a close evolutionary relationship with H. novaeseelandiae. The complete mitogenome of T. sphenurus reported here is expected to provide valuable molecular information for further studies on the phylogeny of the genus Treron and for analyses of the taxonomic status of the family Columbidae.

Mitochondrial Genome Structures and Phylogenetic Analyses of Two Tropical Characidae Fishes.

The Characidae family contains the largest number of tropical fish species. Morphological similarities make species identification difficult within this family. Here, the complete mitogenomes of two Characidae fish were determined and comparatively analyzed with those of nine other Characidae fish species. The two newly sequenced complete mitogenomes are circular DNA molecules with sizes of 16,701 bp (Hyphessobrycon amandae; MT484069) and 16,710 bp (Hemigrammus erythrozonus; MT484070); both have a highly conserved structure typical of Characidae, with the start codon ATN (ATG/ATT) and stop codon TAR (TAA/TAG) or an incomplete T--/TA-. Most protein-coding genes of the 11 Characidae mitogenomes showed significant codon usage bias, and the protein-coding gene cox1 was found to be a comparatively slow-evolving gene. Phylogenetic analyses via the maximum likelihood and Bayesian inference methods confirmed that H. amandae and H. erythrozonus belong to the family Characidae. In all Characidae species studied, one genus was well supported; whereas other two genera showed marked differentiation. These findings provide a phylogenetic basis for improved classification of the family Characidae. Determining the mitogenomes of H. erythrozonus and H. amandae improves our understanding of the phylogeny and evolution of fish species.

Genetic diversity of wild wintering red-crowned crane (Grus japonensis) by microsatellite markers and mitochondrial Cyt B gene sequence in the Yancheng reserve.

The red-crowned crane (Grus japonensis) is one of the most endangered cranes in the world, and its wild population is still declining. To characterize the genetic resources of East Asian migratory populations, we studied the genetic variation in wild red-crowned cranes at the Yancheng reserve. Based on a partial Cyt b gene sequence, 32 wild red-crowned crane samples were screened from 100 feathers with unknown sample information. Twelve haplotypes were detected using 32 wild red-crowned crane samples. Six pairs of published microsatellite primers were selected for genotyping. A total of 47 alleles were obtained, with an average of 7.8 alleles per locus. All microsatellite loci were highly polymorphic; the average polymorphic information content and expected heterozygosity were 0.721 ± 0.080 and 0.768 ± 0.071, respectively. These results show that the East Asian migratory population of wild red-crowned cranes exhibits high polymorphism. These data are useful for informing reintroduction efforts. The study results provide a basis for understanding the population genetic properties of an endangered crane.

Complete mitogenomic and phylogenetic characteristics of the speckled wood-pigeon (Columba hodgsonii).

The speckled wood-pigeon, Columba hodgsonii, is mainly distributed in Bhutan, China, India, Laos, Myanmar, Nepal, Pakistan, and Thailand. Although there are several studies on birds in the family Columbidae, no study has focused on C. hodgsonii, a member of this family. Therefore, this study aimed to clarify the phylogenetic status of C. hodgsonii. The complete mitochondrial genome (mitogenome) of C. hodgsonii was sequenced and characterized and compared with those of other Columba species. The C. hodgsonii mitogenome was found to be 17,477 bp in size and contained 13 PCGs, two rRNAs, 22 tRNAs, and one CR. Of the 37 genes encoded by the C. hodgsonii mitogenome, 28 were on the heavy strand and nine were on the light strand. Twelve PCGs were initiated by ATN codons and one PCG harbored an incomplete termination codon (T-). The base composition of C. hodgsonii PCGs was A = 29.44%, T = 24.37%, G = 12.43%, and C = 33.76%. For the whole mitogenome, including PCGs, rRNAs, tRNAs, and the control region, the AT-skew was positive, and the GC-skew was negative. Phylogenetic analysis based on the base sequences of 13 PCGs from 28 Columbidae species and one outgroup using maximum likelihood and Bayesian inference indicated that C. hodgsonii belongs to the genus Columba and that the family Columbidae is monophyletic.

Mitogenome of the little owl Athene noctua and phylogenetic analysis of Strigidae.

New advances in molecular approaches for DNA analysis have enhanced our understanding of the phylogenetic relationship of birds. The Little Owl (Athene noctua) is of great significance for the integrated management of forest diseases and control of regional pests. Here, we sequenced and annotated the 17,772 bp complete mitogenome of A. noctua. The mitogenome encoded 37 typical mitochondrial genes: 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA, and one non-coding control region (D-loop). The organization and location of genes in the A. noctua mitogenome were consistent with those reported for other Strigidae birds. Phylogenetic relationships based on Bayesian inference and Maximum likelihood methods showed that A. noctua has close relationships with Athene brama and Glaucidium cuculoides, confirming that A. noctua belongs to the Strigidae family. The phylogenetic relationships among seven genera of the Strigidae family used in this study were: Ninox and the other six genera were far apart, Otus and the clade ((Bubo + Strix) + Asio) were clustered into one branch, and Athene and Glaucidium were clustered into one branch. This phylogenetic classification is consistent with prior taxonomic studies on the Strigidae family. Our results provide new mitogenomic data to support further phylogenetic and taxonomic studies of Strigidae.

Five new mitogenomes of Phylloscopus (Passeriformes, Phylloscopidae): Sequence, structure, and phylogenetic analyses.

The genus Phylloscopus belongs to the order Passeriformes and subfamily Phylloscopinae within the family Sylviidae. Phylloscopus, a small insectivore widely distributed in the old world, includes 66 species and 112 subspecies. We identified five new Phylloscopus mitogenomes: P. tenellipes (16,904 base pairs (bp), MK390475), P. coronatus (16,905 bp, MK533705), P. borealis (16,881 bp, MK390476), P. schwarzi (16,920 bp, MK411584), and P. borealoides (16,904 bp, MN125373). All contained 37 genes, including 13 protein-coding genes, two rRNA genes, 22 tRNA genes, and two control regions. Seven Phylloscopus species were compared to the five new mitogenomes and the published complete P. inornatus and P. proregulus mitochondrial sequences. The genetic distance of 13 single protein-coding genes exhibited low variation within all seven Phylloscopus mitogenomes. Based on the Ka/Ks ratios, the molecular evolution patterns of single protein-coding genes were relatively consistent among Phylloscopus bird species. Phylogenetic analysis verified that subspecies of the Pale-legged Leaf Warbler P. tenellipes borealoides could be promoted to the Sakhalin Leaf Warbler P. borealoides. Gene order and genome organization information is useful to understand evolutionary relationships among Phylloscopus species. The complete mitogenomes of these five Phylloscopus species provide genetic markers for species identification, population genetics, and phylogeographic studies of birds.

Analysis of the Gut Microbiome of Wild and Captive Père David's Deer.

Père David's deer (Elaphurus davidianus or milu) is a highly endangered species originating from China, and many deer are currently being raised in captivity for gradual re-introduction to the wild. Wild and captive deer currently live in the same region but have vastly different diets. In this study, we used 16S rRNA high-throughput sequencing to identify the healthy core microbiome in the gut of wild and captive Père David's deer and investigate how dietary factors influence the gut microbiome by comparing their differences. A core shared gut microbiome was identified in healthy Père David's deer, which was similar to that of other ruminants, mainly comprising the phyla Firmicutes and Bacteroidetes. There were no differences in the richness or diversity of the gut microbiome between the wild and captive deer. However, PCA and ANOSIM demonstrated clear differences in the microbial community structure between the captive and wild deer, which mainly manifested as changes in the relative abundance of 39 bacterial genera. As the majority of these genera were not dominant in the deer gut, no significant difference was detected in functional modules related to the microbiome between the two groups. Therefore, the difference in dietary factors does not appear to affect the healthy core gut microbiome between captive and wild Père David's deer, suggesting strong co-evolution and the possibility of re-establishment in the wild. These data could guide future applications of population management in Père David's deer conservation.

[Photosynthetic characteristics and active ingredients differences of Asarum heterotropoides var. mandshuricum under different light irradiance].

Chlorophyll content,leaf mass to per area,net photosynthetic rate and bioactive ingredients of Asarum heterotropoides var. mandshuricum,a skiophyte grown in four levels of solar irradiance were measured and analyzed in order to investigate the response of photosynthetic capability to light irradiance and other environmental factors. It suggested that the leaf mass to per area of plant was greatest value of four kinds of light irradiance and decreasing intensity of solar irradiance resulted in the decrease of leaf mass to per area at every phenological stage. At expanding leaf stage,the rate of Chla and Chlb was 3. 11 when A. heterotropoides var. mandshuricum grew in full light irradiance which is similar to the rate of heliophytes,however,the rate of Chla and Chlb was below to 3. 0 when they grew in shading environment. The content of Chla,Chlb and Chl( a+b) was the greatest value of four kinds of light irradiance and decreasing intensity of solar irradiance resulted in its decreasing remarkably( P<0. 05). The rate of Chla and Chlb decreased but the content of Chla,Chlb and Chl( a+b) increased gradually with continued shading. The maximum value of photosynthetically active radiation appeared at 10: 00-12: 00 am in a day. The maximum value of net photosynthetic rate appeared at 8: 30-9: 00 am and the minimum value appeared at 14: 00-14: 30 pm at each phenological stage if plants grew in full sunlight. However,when plants grew in shading,the maximum value of net photosynthetic rate appeared at about 10: 30 am and the minimum value appeared at 12: 20-12: 50 pm at each phenological stage. At expanding leaf stage and flowering stage,the average of net photosynthetic rate of leaves in full sunlight was remarkably higher than those in shading and it decreased greatly with decreasing of irradiance gradually( P < 0. 05). However,at fruiting stage,the average of net photosynthetic rate of leaves in full sunlight was lower than those in 50% and 28% full sunlight but higher than those in 12% full sunlight. All photosynthetic diurnal variation parameters of plants measured in four kinds of different irradiance at three stages were used in correlation analysis. The results suggested that no significant correlation was observed between net photosynthetic rate and photosynthetically active radiation,and significant negative correlation was observed between net photosynthetic rate and environmental temperature as well as vapor pressure deficit expect for 12% full sunlight. Positive correlation was observed between net photosynthestic rate and relative humidity expect for 12% full sunlight. Significant positive correlation was observed between net photosynthetic rate and stomatal conductance in the four light treatments. Only,in 12% full sunlight,the net photosynthetic rate was significantly related to photosynthetically active radiation rather than related to environmental temperature,vapor pressure deficit and relative humidity. In each light treatment,a significant positive correlation was observed between environmental temperature and vapor pressure deficit,relative humidity as well as stomatal conductance. Volatile oil content was 1. 46%,2. 16%,1. 56%,1. 30% respectively. ethanol extracts was 23. 44%,22. 45%,22. 18%,21. 12% respectively. Asarinin content was 0. 281%,0. 291%,0. 279% and 0. 252% respectively. The characteristic components of Asarum volatile oil of plant in different light treatments did not change significantly among different groups.

Comparative analysis of the gut microbiota of hornbill and toucan in captivity.

Gut microbiota plays an important role in animals and are considered microbial organs. Hornbill and toucan are birds of the same ecotypes with high appreciative value. In this study, we characterized and compared the gut microbiota of toco toucan (Ramphastos toco), great hornbill (Buceros bicornis) and wreathed hornbill (Rhyticeros undulatus) using 16S rRNA high-throughput sequencing technology, and further discussed the influence of host bird genetics on its gut microbiota. We identified 10,847 operational taxonomic units (OTUs) from the hyper-variable V4-V5 region, representing 14 phyla that were dominated by the Firmicutes, Proteobacteria, Cyanobacteria, Fusobacteria, Actinobacteria, and Bacteroidetes. Alpha diversity indices showed no significant difference among the three species (p > 0.1). However, great hornbill and toco toucan shared a high number of OTUs. Principal component analysis also revealed highly similar gut microbiotas between the two distant species. Therefore, environmental factors might dominate over host genetics in shaping the gut microbiotas of hornbill and toucan. Our study would contribute in elucidating adaptation of the hornbill and toucan to environmental change.

Genetic diversity analysis of Peking gecko (Gekko swinhonis) in mid-Eastern China based on mitochondrial COI and Cyt b gene sequences.

To understand the genetic diversity of Peking gecko (Gekko swinhonis) populations in its endemic region, 60 individuals were sampled from Lushan, Qi, and Linying counties in Henan Province, China. Through PCR amplification and Sanger sequencing, 120 sequences with lengths of 652 bp (COI) and 739 bp (Cyt b) were obtained, and nine haplotypes were detected for each gene. Overall, results indicated that Peking gecko populations in China have high genetic diversity and significant genetic differentiation. This study provides necessary scientific basis for the protection of Peking gecko germplasm resources.

The complete mitochondrial genome of dark-sided flycatcher Muscicapa sibirica (Passeriformes: Muscicapidae).

We report the mitochondrial genome of Muscicapa sibirica. The overall base composition of the dark-sided flycatcher mitogenome is 24% T, 31.8% C, 29.4% A, and 14.8% G, with an A + T content of 53.4%. The total length of the sequence is 17,879 bp (13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 2 control regions). Phylogenetic analysis was performed based on the concatenated nucleotide sequences of cytochrome c oxidase subunit I and cytochrome b using the neighbor-joining method and the Kimura 2-parameter model in MEGA 7.0 with 1000 bootstrap replicates.

Complete (1)H-NMR and (13)C-NMR spectral assignment of five malonyl ginsenosides from the fresh flower buds of Panax ginseng.

BACKGROUND: Ginsenosides are the major effective ingredients responsible for the pharmacological effects of ginseng. Malonyl ginsenosides are natural ginsenosides that contain a malonyl group attached to a glucose unit of the corresponding neutral ginsenosides. METHODS: Medium-pressure liquid chromatography and semipreparative high-performance liquid chromatography were used to isolate purified compounds and their structures determined by extensive one-dimensional- and two-dimensional nuclear magnetic resonance (NMR) experiments. RESULTS: A new saponin, namely malonyl-ginsenoside Re, was isolated from the fresh flower buds of Panax ginseng, along with malonyl-ginsenosides Rb1, Rb2, Rc, Rd. Some assignments for previously published (1)H- and (13)C-NMR spectra were found to be inaccurate. CONCLUSION: This study reports the complete NMR assignment of malonyl-ginsenoside Re, Rb1, Rb2, Rc, and Rd for the first time.