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Lupinus mutabilis is an under-domesticated legume species from the Andean region of South America. It belongs to the New World lupins clade, which groups several lupin species displaying large genetic variation and adaptability to highly different environments. L. mutabilis is attracting interest as a potential multipurpose crop to diversify the European supply of plant proteins, increase agricultural biodiversity, and fulfill bio-based applications. This study reports the first high-quality L. mutabilis genome assembly, which is also the first sequenced assembly of a New World lupin species. Through comparative genomics and phylogenetics, the evolution of L. mutabilis within legumes and lupins is described, highlighting both genomic similarities and patterns specific to L. mutabilis, potentially linked to environmental adaptations. Furthermore, the assembly was used to study the genetics underlying important traits for the establishment of L. mutabilis as a novel crop, including protein and quinolizidine alkaloids contents in seeds, genomic patterns of classic resistance genes, and genomic properties of L. mutabilis mycorrhiza-related genes. These analyses pointed out copy number variation, differential genomic gene contexts, and gene family expansion through tandem duplications as likely important drivers of the genomic diversity observed for these traits between L. mutabilis and other lupins and legumes. Overall, the L. mutabilis genome assembly will be a valuable resource to conduct genetic research and enable genomic-based breeding approaches to turn L. mutabilis into a multipurpose legume crop.

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Intrahepatic cholangiocarcinoma (ICC) is a universally lethal malignancy with increasing incidence. However, ICC patients receive limited benefits from current drugs; therefore, we must urgently explore new drugs for treating ICC. Quinolizidine alkaloids, as essential active ingredients extracted from Sophora alopecuroides Linn, can suppress cancer cell growth via numerous mechanisms and have therapeutic effects on liver-related diseases. However, the impact of quinolizidine alkaloids on intrahepatic cholangiocarcinoma has not been fully studied. In this article, the in vitro anti-ICC activities of six natural quinolizidine alkaloids were explored. Aloperine was the most potent antitumor compound among the tested quinolizidine alkaloids, and it preferentially inhibited RBE cells rather than HCCC-9810 cells. Mechanistically, aloperine can potentially decrease glutamate content by inhibiting the hydrolysis of glutamine, reducing D-2-hydroxyglutarate levels and, consequently, leading to preferential growth inhibition in isocitrate dehydrogenase (IDH)-mutant ICC cells. In addition, aloperine preferentially resensitizes RBE cells to 5-fluorouracil, AGI-5198 and olaparib. This article demonstrates that aloperine shows preferential antitumor effects in intrahepatic cholangiocarcinoma cells harboring the mutant IDH1 by decreasing D-2-hydroxyglutarate, suggesting that aloperine could be used as a lead compound or adjuvant chemotherapy drug to treat ICC harboring the mutant IDH.

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Lupines (Lupinus spp.) are a common plant species on western U.S. rangelands with several lupine species containing alkaloids that can be toxic and/or teratogenic to livestock. In North America, more than 150 lupine species are recognized with some ranches or grazing allotments containing multiple species. One or more of these lupine species may contain alkaloids that are teratogenic to cattle. Previous work has shown that lupine alkaloids can be detected in earwax of cattle grazing lupine infested rangelands. Our hypothesis is that earwax can be used to determine if cattle have been exposed to teratogenic alkaloids from multiple lupine species. Two lupine species, L. sericeus and L. polyphyllus, were present on a rangeland in east-central Idaho. The teratogen, anagyrine, was detected in L. sericeus and the teratogen, ammodendrine, was detected in L. polyphyllus plants collected on this rangeland. In this study, earwax was collected from 69 pregnant cows that had previously grazed a rangeland containing two different lupine species containing alkaloids that cause crooked calf syndrome (CCS). Anagyrine was detected in the earwax of all 69 cows sampled. Ammodendrine, was detected in the earwax of 28 of the 69 cows sampled. Earwax is a good non-invasive sample to aid in the diagnosis of cattle that have consumed lupine and does appear, in this case, to be a good diagnostic tool to differentiate between more than one lupine species that may be the cause of CCS. Concentrations of anagyrine or ammodendrine did not correlate with the incidence of CCS.

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Lupins are used in animal feed because of their excellent nutritional composition. Australian and European Lupinus angustifolius seeds are incorporated in compound feed of calves for veal production in Belgium. To investigate the co-occurrence of quinolizidine alkaloids (QAs) and phomopsin A (PHO A) in lupin seeds and lupin-containing feed, and the potential transfer to animal-derived foods, representative samples were obtained from various actors in the chain. A UHPLC-MS/MS method was validated for the simultaneous quantification of seven QAs and PHO A in relevant matrices. Results indicate highly consistent total QA (TQA) levels in Australian lupins (173 ± 24 mg/kg) (n = 25), while European samples showed a high variability (1442 ± 1497 mg/kg) (n = 15). PHO A was detected in 7 of 40 samples. Lupin-containing feed had a mean TQA content of 42 ± 28 mg/kg (n = 20). An in vivo feeding trial demonstrated the transfer of QAs to muscle and liver of calves that were fed the lupin-containing feed. Highest concentrations were found for lupanine in liver tissue samples (67 ± 46 µg/kg). PHO A concentrations were below the LOD in all feed and tissue samples. These results indicate that animal-derived foods (veal meat/liver) are a potential route for QAs to enter the food chain.

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Alkaloids play an essential role in protecting plants against herbivores. Humans can also benefit from the pharmacological effects of these compounds. Plants produce an immense variety of structurally different alkaloids, including quinolizidine alkaloids, a group of bi-, tri-, and tetracyclic compounds produced by Lupinus species. Various lupin species produce different alkaloid profiles. To study the composition of quinolizidine alkaloids in lupin seeds, we collected 31 populations of two wild species native to Israel, L. pilosus and L. palaestinus, and analyzed their quinolizidine alkaloid contents. Our goal was to study the alkaloid profiles of these two wild species to better understand the challenges and prospective uses of wild lupins. We compared their profiles with those of other commercial and wild lupin species. To this end, a straightforward method for extracting alkaloids from seeds and determining the quinolizidine alkaloid profile by LC-MS/MS was developed and validated in-house. For the quantification of quinolizidine alkaloids, 15 analytical reference standards were used. We used GC-MS to verify and cross-reference the identity of certain alkaloids for which no analytical standards were available. The results enabled further exploration of quinolizidine alkaloid biosynthesis. We reviewed and re-analyzed the suggested quinolizidine alkaloid biosynthesis pathway, including the relationship between the amino acid precursor l-lysine and the different quinolizidine alkaloids occurring in seeds of lupin species. Revealing alkaloid compositions and highlighting some aspects of their formation pathway are important steps in evaluating the use of wild lupins as a novel legume crop.

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Quinolizidine alkaloids (QAs) are toxic secondary metabolites of the Lupinus species, the presence of which limits the expansion of lupin beans consumption, despite their high protein content. Evaluation of the level of alkaloids in edible Lupinus species is crucial from a food safety point of view. However, quantitation of QAs is complicated by the fact that not all important alkaloids used for quantitation are commercially available. In this context, we developed a method for the simultaneous quantitation of eight major lupin alkaloids using quantitative NMR spectroscopy (qNMR). Quantitation and analysis were performed in 15 different seed extracts of 11 Lupinus spp. some of which belonged to the same species, with different geographical origins and time of harvest, as well as in all aerial parts of L. pilosus. The mature seeds of L. pilosus were found to be a uniquely rich source of multiflorine. Additionally, we developed a protocol using adsorption or ionic resins for easy, fast, and efficient debittering of the lupine seeds. The protocol was applied to L. albus, leading to a decrease of the time required for alkaloids removal as well as water consumption and to a method for QA isolation from the debittering wastewater.

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As part of our ongoing investigation of natural bioactive substances from the genus Thermopsis of the tribe Fabaceae for agricultural protection, the chemical constituents of the herb Thermopsis lupinoides were systematically investigated, which led to the isolation of 39 quinolizidine alkaloids (QAs) (1-39), including 14 new QAs (1-14) and 14 isoflavones (40-53). Their structures were elucidated through comprehensive spectroscopic data analysis (IR, UV, NMR, HRESIMS), ECD calculations, and X-ray crystallography. The antitomato spotted wilt virus (TSWV) and antifungal (against Botrytis cinerea, Gibberella zeae, Phytophythora capsica, and Alternaria alternata) and insecticidal (against Aphis fabae and Tetranychus urticae) activities of the isolated compounds were screened using the lesion counting method, mycelial inhibition assay, and spray method, respectively. The bioassay results showed that 34 exhibited excellent protective activity against TSWV, with an EC50 value of 36.04 µg/mL, which was better than that of the positive control, ningnanmycin (86.03 µg/mL). The preliminary mechanistic exploration illustrated that 34 induced systemic acquired resistance in the host plant by acting on the salicylic acid signaling pathway. Moreover, 1 showed significant antifungal activity against B. cinerea (EC50 value of 20.83 µg/mL), while 2 exhibited good insecticidal activity against A. fabae (LC50 value of 24.97 µg/mL). This research is promising for the invention of novel pesticides from QAs with high efficiency and satisfactory ecological compatibility.

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Endophytic fungi of medicinal plants are symbiotic with the host and play an important role in determining metabolites. To understand the relationship between the accumulation of Sophora alopecuroides' medicinal bioactive compounds and the ecological succession of endophytic fungi, here we collected samples from S. alopecuroides at four developmental stages (adult, flowering, podding, and mature) and different organs (roots, stems, leaves, and seeds) at the mature stage. We then used high-performance liquid chromatography-mass spectrometry and high-throughput sequencing on the internal transcribed spacer region to identify the medicinal compounds and endophytic fungal communities in each sample. The endophytic fungal community characteristics and accumulation of medicinally bioactive compounds of S. alopecuroides varied with the host's developmental stages and organs, with the highest total alkaloids content of 111.9 mg/g at the mature stage. Membership analysis and network connection analysis showed a total of 15 core endophytic fungi in different developmental stages and 16 core endophytic fungi in different organs at the mature stage. The unclassified Ascomycota, Aspergillus, and Alternaria were significantly and positively correlated with the medicinal compounds of S. alopecuroides at the mature stage (r > 0.6 or r < -0.6; P < 0.05). In this study, we identified key endophytic fungal resources that affect the content of medicinally bioactive compounds in S. alopecuroides. This discovery could lay the foundation for enhancing the yield of medicinally bioactive compounds in S. alopecuroides and the development and application of functional endophytic fungi.IMPORTANCESophora alopecuroides is a traditional Chinese herbal medicine. The major medicinal chemicals are considered to be quinolizidine alkaloids. Quinolizidine alkaloids have been widely used for the treatment of tumors, dysentery, and enteritis. Previous studies have found that endophytic fungi in S. alopecuroides can promote the accumulation of host quinolizidine alkaloids. However, the relationship between the accumulation of S. alopecuroides' medicinal bioactive compounds and the ecological succession of endophytic fungi remains unclear. In this study, we screened the key endophytic fungal resources affecting the content of medicinally bioactive compounds and laid the foundation for subsequent research on the mechanism by which endophytic fungi promote the accumulation of medicinally bioactive compounds in S. alopecuroides.

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Three new quinolizidine alkaloids (1 - 3), including one new naturally isoflavone and cytisine polymer (3), along with 6 known ones were isolated from the ethanol extract of Sophora tonkinensis Gagnep. Their structures were elucidated by comprehensive spectroscopic data analysis (IR, UV, HRESIMS, 1D and 2D NMR), combined with ECD calculations. The antifungal activity against Phytophythora capsica, Botrytis cinerea, Gibberella zeae, and Alternaria alternata of the compounds was evaluated in a mycelial inhibition assay. Biological tests indicated that compound 3 exhibited strong antifungal activity against P. capsica with EC50 values of 17.7 µg/ml.

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Cotton aphids, Aphis gosspyii and cotton jassids, Amrasca biguttula are destructive piercing sucking pests to many strategic crops, especially cotton, not only in Egypt but also all over the world. Using synthetic pesticides to control these pests led to several deleterious impacts. Natural pesticides can be used as a harmless alternative. Nine compounds were isolated from different fractions of Retama raetam using chromatographic techniques and identified by spectroscopic methods as eugenol (1), alpinumisoflavone (2), licoflavone C (3), ephedroidin (4), anagyrine (5), spartiene (6), genistein-8ß-C-glucoside (7), isoprunetin (8) and isoprunetin 7-O-ß-D-glucopyranoside (9). The methanol crude extract and its fractions (hexane, chloroform, ethyl acetate and butanol), as well as the isolated compounds were examined against A. gosspyii and A. biguttula as insecticides. The results showed that chloroform fraction was the most potent fraction against A. gosspyii and A. biguttula, with LC50 values of 65.66 and 64.43 ppm, respectively. As well, compounds 1, 5 and 6 were found to be more active, with LC50 values of 69.84, 25.49 and 27.22 ppm for A. gosspyii and 65.17, 24.07 and 24.78 ppm for A. biguttula, respectively. The most potent compounds (1, 5 and 6) exhibited AChE inhibition toward A. gosspyii compared with the control. So, it can be concluded that the isolated compounds eugenol 1, anagyrine 5 and spartiene 6 are the active principles due to their capability to inhibit AchE activity.

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In the current era, it is important to consider economic and ecological sustainability issues while optimally meeting the nutrient needs of poultry. The use and research of alternative feedstuffs have gained importance due to these factors. The aim of this study is to reveal the raw lupin seeds' nutrient ingredients as an alternative feedstuff and the effects of debittering methods. In the present study, two different treatments (germination for 2 days; heat treatment in an autoclave at 130 °C for 20 min) were applied to white and blue lupin seeds, and the differences in nutrient compositions between them and raw seeds were determined. When fatty acid compositions were analyzed, oleic, γ-linolenic, arachidic, behenic, erucic, and lignoceric acid values were found to be the highest in the raw, autoclaved, and germinated forms of white lupin (p < 0.01). The highest values of palmitic, stearic, and linoleic acids were observed in blue lupin (p < 0.01). While the value of total quinolizidine alkaloids (QA) in raw white lupin grains was higher than 1.943 mg/g, it was higher than 1.800 mg/g in autoclaved and germination-treated grains. Similarly, the total QA value of raw blue lupin grains was 0.894 mg/g, 0.609 ± 0.244 mg/g in germination-treated seeds, and 0.705 ± 0.282 mg/g in autoclave-treated seeds. As a result of these findings, it can be said that the methods applied for the removal of bitterness gave promising results. Furthermore, it would be rewarding to use these lupin varieties in in vitro and in vivo experiments to reveal the impacts and mechanisms of debittering methods on poultry.

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From the leaves and stem bark of the Kenyan medicinal plant Calpurnia aurea subsp. aurea, four previously undescribed quinolizidine alkaloids namely, 2ß-methoxy-13α-O-(2'-pyrrolylcarbonyl) virgiline, 2α-methoxy-13ß-O-(2'-pyrrolylcarbonyl) virgiline, 3α-O-angelate-2ß-hydroxy-13α-O-(2'-pyrrolylcarbonyl) virgiline, 2,3-dehydro-virgiline were isolated together with four known ones. Structural elucidation of the compounds was based on 1D and 2D NMR spectroscopy and mass spectrometry. Their relative configurations were determined by NOESY correlations and literature. The quinolizidine alkaloids were tested against Trichophyton rubrum, Trichophyton interdigitale, Trichophyton benhamiae, Microsporum canis and Nannizzia gypsea, common causative agents of most of the tinea infections in human. All the isolated quinolizidine alkaloids exhibited antidermatophytic activity with MIC ranging from 37.5 µg/ml to 300 µg/ml.

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Fourteen quinolizidine derivatives, structurally related to the alkaloids lupinine and cytisine and previously studied for other pharmacological purposes, were presently tested for antiarrhythmic, and other cardiovascular effects on isolated guinea pig heart tissues in comparison to well-established reference drugs. According to their structures, the tested compounds are assembled into three subsets: (a) N-(quinolizidinyl-alkyl)-benzamides; (b) 2-(benzotriazol-2-yl)methyl-1-(quinolizidinyl)alkyl-benzimidazoles; (c) N-substituted cytisines. All compounds but two displayed antiarrhythmic activity that was potent for compounds 4, 1, 6, and 5 (in ascending order). The last compound (N-(3,4,5-trimethoxybenzoyl)aminohomolupinane) was outstanding, exhibiting a nanomolar potency (EC50 = 0.017 µM) for the increase in the threshold of ac-arrhythmia. The tested compounds shared strong negative inotropic activity; however, this does not compromise the value of their antiarrhythmic action. On the other hand, only moderate or modest negative chronotropic and vasorelaxant activities were commonly observed. Compound 5, which has high antiarrhythmic potency, a favorable cardiovascular profile, and is devoid of antihypertensive activity in spontaneously hypertensive rats, represents a lead worthy of further investigation.

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Lupin, a protein-rich grain legume, and products thereof, are becoming increasingly important in our diets. However, variable and high concentrations of quinolizidine alkaloids (QAs) may hamper this evolution. This study assessed the fate of QAs when processing Lupinus albus seeds and lupin-based foods, to give a first indication of the food industry's ability to sufficiently reduce the QA concentration. Typical unit processes, including toasting, dehulling, sterilization (sterilized jarred lupins), oven baking (cookies), frying (chips) and boiling in water (pasta), were simulated on lab-scale. A quantitative determination of five QAs and qualitative screening of other relevant QAs, in the derived fractions and lupin-based foods, was performed with a validated UHPLC-MS/MS and -HRMS method, respectively. Results revealed that the reduction in quinolizidine alkaloid content is highly dependent on the applied unit process, that QAs appear to be heat stabile, and that the depletion can be attributed to the leaching in cooking water.

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This study describes the analysis of five quinolizidine alkaloids (QA), i.e. 13-OH-lupanine, lupanine, lupinine, angustifoline and sparteine in 30 samples of lupine flours, lupine seeds and derived products collected 2019-2021 on the German retail market. QA occur as secondary metabolites in plants of lupine species. Certain QA are of toxicological relevance. The analytical determination performed by LC-MS/MS revealed some samples with high QA concentrations (up to 21,000 mg/kg), particularly in bitter lupine seeds. As those concentrations would result in substantial exceedances of the maximum tolerable intake values proposed by health authorities, they must be considered as a health concern.

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Quinolizidine alkaloids isolated from various marine and terrestrial animals and plants are primarily composed of lupinine-, matrine-, and sparteine-type alkaloids. Matrine, phenanthroquinolizidines, bis-quinolizidines, and small molecules from amphibian skins are representative compounds of such alkaloids. Quinolizidine alkaloids harbor anticancer, antibacterial, antiinflammatory, antifibrosis, antiviral, and anti-arrhythmia. In this chapter, we comprehensively outline the biological activity and pharmacological action of quinolizidine alkaloids and discuss new avenues toward the discovery of novel and more efficient drugs based on these naturally occurring compounds. It is urgent for basic research and clinical practice to conduct more targeted comprehensive research based on the lead drugs of quinolizidine alkaloids with significant pharmacological activity.

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Total syntheses of geissoschizine-type monoterpenoid indole alkaloids (MTIAs) are reported. An intramolecular Pictet-Spengler cyclization was developed for the selective construction of the 3R stereocenter. The first total synthesis of (+)-villocarine A was then achieved. Furthermore, the first total synthesis of the highly strained (-)-apogeissoschizine was also accomplished in an aza-Michael cyclization/E1cB elimination/stereoselective olefin isomerization sequence. Finally, (+)-geissoschizine, a common biosynthetic intermediate of MTIAs, was obtained from apogeissoschizine through ring-opening along with a release of ring strain.

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Previous studies have individually shown the antidiabetic potential of gamma conglutin (Cγ) and lupanine from lupins. Until now, the influence of combining both compounds and the effective dose of the combination have not been assessed. Moreover, the resulting gene expression profile from this novel combination remains to be explored. Therefore, we aimed to evaluate different dose combinations of Cγ and lupanine by the oral glucose tolerance test (OGTT) to identify the higher antidiabetic effect on a T2D rat model. Later, we administered the selected dose combination during a week. Lastly, we evaluated biochemical parameters and liver gene expression profile using DNA microarrays and bioinformatic analysis. We found that the combination of 28 mg/kg BW Cγ + 20 mg/kg BW lupanine significantly reduced glycemia and lipid levels. Moreover, this treatment positively influenced the expression of Pdk4, G6pc, Foxo1, Foxo3, Ppargc1a, Serpine1, Myc, Slc37a4, Irs2, and Igfbp1 genes. The biological processes associated with these genes are oxidative stress, apoptosis regulation, and glucose and fatty-acid homeostasis. For the first time, we report the beneficial in vivo effect of the combination of two functional lupin compounds. Nevertheless, further studies are needed to investigate the pharmacokinetics and pharmacodynamics of the Cγ + lupanine combined treatment.

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Despite its worldwide relevance as an invasive plant, there are few studies on Ulex europaeus (gorse) and its allelopathic activity is almost unexplored. The allelochemical profile of gorse was analysed through methanolic extract of pods and roots, and its phytotoxic effects on Lactuca sativa germination. The methanolic extract of pods had no effect in germination, while extract of roots resulted in a U-shaped dose-response curve: reducing the germination at concentration 0.5 mg mL-1. GC-MS analysis detected compounds with proven antimicrobial and antioxidant activities in the pods and cytotoxic compounds in the roots, which could explain the bioassay results. The quinolizidine alkaloids (QAs) composition was evaluated to predict possible biological functions. It showed the presence of QAs in gorse that are absent in their native range, indicating broad defense strategies against bacteria, fungi, plants, and insects in the Chilean ecosystem. This could explain the superiority of gorse in the invaded areas.

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Sophora alopecuroides L. is a traditional Chinese medicine used for the treatment of several different disease states including bacillary dysentery and enteritis. But importantly, it also plays a role as an anti-tumor agent. That said, little is known about the role endophytes play regarding the clinically bioactive metabolites in S. alopecuroides. In order to explore the effects of endophytic fungi on the accumulation, quality, and correlation in the content of the medicinal compounds, the structural diversity of endophytic fungi in S. alopecuroides was analyzed. The relationship between endophytes and quinolizidine alkaloids (QAs), housed within the seeds of S. alopecuroides, which were interpreted based on established methods of high-throughput sequencing and high-performance liquid chromatography. A total of 1,034,418 effective sequence reads and 257 operational taxonomic units (OTUs) were obtained from 33 samples which were sourced from 11 different sampling sites and further classified into 9 phyla, 20 classes, 45 orders, 85 families, and 118 genera. Ascomycota was found to be the dominant phylum of endophytic fungi in S. alopecuroides, with a relative abundance ranging from 60.85 to 98.30%. Alternaria, Cladosporium, Filobasidium, and an unidentified Ascomycota were the core-shared endophytes, accounting for 49.96, 27.12, 14.83, and 7.88%, respectively. Correlation analysis showed that the Simpson's diversity index of endophytic fungal community in S. alopecuroides was significantly positively correlated with the Oxymatrine (OMA) content in different areas, while the Chao and Shannoneven indexes were significantly negatively correlated with OMA. The endophytic fungi of Alternaria were positively correlated with the content of OMA, Oxysophocarpine (OSC), and total QAs. This study has mastered the endophytic fungi resources of S. alopecuroides, explored potential functional endophytic fungi, and provided a scientific basis for using biological fertilization strategies to improve the quality of S. alopecuroides.