Comparative study on dynamic in vitro digestion characteristics of lotus seed starch-EGCG complex prepared by different processing methods.

To study the effect of starch-polyphenol interaction induced by different processing methods on digestion characteristics, a dynamic in vitro human gastrointestinal system was employed to investigate the digestive characteristics of lotus seed starch-epigallocatechin gallate (EGCG) complex (LS-EGCG) prepared by different processing methods. Digestion altered crystal structure, particle size, morphology, pH, starch hydrolysis, and EGCG content. Processing broke physical barriers, reducing particle size by enzyme erosion. Enzymatic hydrolysis gradually exposed EGCG, indicated by green fluorescence. Heat and high pressure treatments enhanced starch dissolution, increasing sugar accumulation and hydrolysis. However, ultrasonic-microwave and high pressure microfluidization treatments formed dense structures, decreasing hydrolysis rates. Overall, the complex formed by high pressure microfluidization showed better enzyme resistance. The results provide a scientific basis for the development of food with quality and functional properties.

High-performance peptide biosensor based on unified structure of lotus silk.

The sensitive materials of current gas sensors are fabricated on planar substrates, significantly limiting the quantity of sensitive material available on the sensor and the complete exposure of the sensitive material to the target gas. In this work, we harnessed the finest, resilient, naturally degradable, and low-cost lotus silk derived from plant fibers, to fabricate a high-performance bio-sensor for toxic and harmful gas detection, employing peptides with full surface connectivity. The proposed approach to fabricate gas sensors eliminated the need for substrates and electrodes. To ascertain the effectiveness and versatility of the sensors created via this method, sensors for three distinct representative gases (isoamyl alcohol, 4-vinylanisole, and benzene) were prepared and characterized. These sensors surpassed reported detection limits by at least one order of magnitude. The inherent pliancy of lotus silk imparts adaptability to the sensor architecture, facilitating the realization of 1D, 2D, or 3D configurations, all while upholding consistent performance characteristics. This innovative sensor paradigm, grounded in lotus silk, represents great potential toward the advancement of highly proficient bio gas sensors and associated applications.

Investigating the efficacy of purified tannin extracts from underutilized temperate forages in reducing enteric methane emissions in vitro.

The study investigated how the concentration and composition of purified tannin extracts, at various inclusion rates, affect the ruminal in vitro fermentation parameters. Tannin extracts were isolated from four different forage species: birdsfoot trefoil (Lotus corniculatus), sulla (Hedysarum coronarium), big trefoil (Lotus pedunculatus), and salad burnet (Sanguisorba minor). Plants extracts were purified by Sephadex LH-20 gel chromatography and analyzed by UPLC-ESI-MS/MS. The results showed a large variation among the extracts from different species in terms of tannin composition and structural features. The extracts from salad burnet were dominated by hydrolysable tannins, comprising mainly ellagitannins. The extracts derived from sulla and big trefoil contained predominantly proanthocyanidins (PA), primarily composed of prodelphinidins with high mean degree of polymerisation (mDP). Birdsfoot trefoil extracts comprised procyanidin-rich PAs with low mDP. To determine whether the combined presence of tannins and flavonoid together lead to synergistic or antagonistic effects, the tannin extracts were incubated both with or without rutin at concentrations of 10, 20, and 30 g/kg DM, using a base substrate of perennial ryegrass (Lolium perenne, control). In general, all the tannin extracts decreased methane (CH4) production compared to the control, while no significant effect of rutin was observed on both gas (GP) and CH4 production, neither pure, nor in the simultaneous presence of tannins. The highest CH4 reduction (15%, at 30 g/kg DM) was observed from sulla and big trefoil extracts compared to control, but this was also supplemented with a concomitant reduction in GP (11%) indicating a reduction in feed digestibility. The extracts from birdsfoot trefoil and salad burnet reduced CH4 by up to 12% without significantly reducing GP, indicating the importance of tannin composition on ruminal fermentation.

Ultrasound-assisted extraction of polyphenols from lotus rhizome epidermis by alcohol/salt-based aqueous two-phase system: Optimization, extraction mechanism and antioxidant activities.

In this study, ultrasonic-assisted (UA) alcohol/salt-based aqueous two-phase system (ATPS) method was constructed to extract lotus rhizome epidermis (LRE) polyphenols. The extraction conditions were optimized as salt concentration 26.75 %, ethanol concentration 25.45 %, ultrasonic power 487 W and liquid-solid ratio 35.33 mL/g by comparing response surface methodology (RSM) and artificial neural network (ANN) models. Then, l-dopa (2.35 ± 0.036 mg/g dw), gallocatechin (1.66 ± 0.0035 mg/g dw) and epigallocatechin (1.37 ± 0.0035 mg/g dw) were determined as major polyphenols in LRE by using UA-ATPS method. Moreover, study showed that ultrasound, van der Waals force, hydrogen bond and salting out could accelerate the mass transfer and extraction of polyphenols in LRE cells. The high-pressure cavity and collapse effect of ultrasound could also accelerate the extraction of polyphenols. In vitro antioxidant experiments showed that LRE polyphenols have good antioxidant ability. In sum, this study developed a green and efficient extraction method to enhance the profitability of LRE in food and medicine industries.

Structure, spectral properties and antioxidant activity of melanoidins extracted from high temperature sterilized lotus rhizome juice.

Melanoidins are complex macromolecular compounds closely associated with the browning phenomenon in high-temperature sterilized lotus rhizome juice (HTSL). This study aimed to preliminarily investigate the structural properties of melanoidins extracted from HTSL. Results showed that the average molecular weight of HTSL melanoidins ranged from 1.48 to 41.40 kDa. Medium and high molecular weight melanoidins were the main contributors to the brown color of HTSL. Sugars, proteins, and phenolics were present in HTSL, among which sugar was the most abundant, with glucose being the predominant monosaccharide in acid degradation products of melanoidins. Through fluorescence and ultraviolet spectral analysis, we found that the melanoidins contained carboxyl and carbonyl compounds, as well as furan and pyran heterocyclic compounds. The infrared spectra and nuclear magnetic resonance spectra revealed a prominent sugar absorption peak, indicating that sugar was the main component of the melanoidins of HTSL. Furthermore, in vitro antioxidant experiments showed that the antioxidant activity of melanoidins was significantly positively correlated with phenolic compounds. Our results indicated that there were differences in the structural properties of melanoidins fractions with different molecular weights. MW-H fraction significantly impacted the color and antioxidant activity of HTSL.

Comparison of anti-allergic activities of different types of lotus seed resistant starch in OVA-induced mouse model.

Currently, evidence from observational studies suggests dietary fiber intake may be associated with decreased risk of food allergy. As a type of dietary fiber, resistant starch was also widely reported to possess anti-allergic properties. However, there is a relative paucity of studies assessing the influence of resistant starch types on their anti-allergic activity and its possible underlying mechanisms. In the current study, the anti-allergic effects of RS3-type (retrograded starch), RS4-type (chemically modified starch, cross-bonded), and RS5-type (starch-palmitic acid complex) of lotus seed resistant starch were evaluated in the OVA (100 mg/kg)-induced food allergic mice model. The results showed that oral administration of RS3 or RS4 lotus seed resistant starch (0.3 g/100 g b.w.) for 25 days significantly improved adverse symptoms of food allergy such as weight loss, increases in allergy symptom score and diarrhea rate; with significant reduction of serum specific antibody IgE, TNF-α, IL-4 levels and improved Th1/Th2 balance being observed. The mechanism may involve the regulation of lotus seed resistant starch on intestinal flora and the metabolites short-chain fatty acids and bile acids. Taken together, the findings may enhance understanding towards ameliorative effects of resistant starch on food allergy, and offer valuable insights for the exploration of novel anti-allergic bioactive compounds.

Study on the formation mechanism of blackening in damaged lotus rhizome epidermis: Effects of polyphenols and iron.

Lotus rhizome is an important aquatic vegetable, but the blackening of lotus rhizome epidermis (LRE) seriously affects its appearance and quality, which makes lotus rhizome products unmarketable. In this study, the effects of polyphenols and iron on the LRE color were studied to explore the possible mechanism of LRE blackening. Results indicated that the measurable total phenols contents in the mud treatment (MT) group were significantly reduced, and the total iron contents were significantly increased compared with the bruised treatment group (p < 0.05). The high-performance liquid chromatography results showed that the main polyphenols in LRE were dopa, gallocatechin, and catechin, as well as a small amount of catechol, epicatechin, proanthocyanidin B2, and proanthocyanidin C1. Moreover, the results of color difference and ultraviolet adsorption spectroscopy showed that there were obviously black or brown-gray of dopa (525 nm), gallocatechin (504.5 nm), and catechin (550 and 504.5 nm) with FeCl2. The simulated system treatment of LRE further confirmed that the chromaticity effect of dopa and iron in bruised LRE was similar to that of the MT group, whereas 1% (w/w) ascorbic acid, 2% (w/w) EDTA-2Na, or 3% (w/w) citric acid could solely prohibit the blackening. This suggested that the dopa in LRE and FeCl2 in mud may mainly combine into [2(DOPA-2H+)+Fe3+]- through non-covalent interaction, which leads to the blackening of bruised LRE under neutral conditions. These results can guide the storage of lotus rhizomes and improve the development of the lotus rhizome industry.

Lotus Leaf Extract Alleviates Lipopolysaccharide-Induced Intestinal Injury in Mice by Regulating Oxidative Stress and Inflammation.

Inflammatory bowel disease, a disease featured by intestinal epithelial barrier destruction and dysfunction, has been a constant threat to animal health. The primary objective of this research was to assess the impact of the extract derived from lotus leaves (LLE) on lipopolysaccharide (LPS) induced damage to the intestines in mice, as well as to investigate the fundamental mechanism involved. The LLE was prepared using ultrasonic extraction in this experiment, and the LLE total flavonoid content was 117.02 ± 10.73 mg/g. The LLE had strong antioxidant activity in vitro, as assessed by 2, 2-diphenyl-1-picrylhydrazyl, ferric reducing antioxidant power, and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) methods. In the vivo experiment, different doses of LLE (50, 100, and 200 mg/kg) were administered for 2 weeks before LPS treatment in mice. The results revealed that LLE alleviates intestinal tissue damage in LPS-induced mice. In the jejunum tissue, LLE significantly upregulated mRNA and protein expression levels of tight junction proteins, such as ZO-1, occludin, and claudin-1, and decreased the contents of the inflammatory cytokines, interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α. Furthermore, the malondialdehyde and lactate dehydrogenase contents increased by LPS in the liver were significantly reduced after administration of LLE, and the total antioxidant capacity, superoxide dismutase, and reduced glutathione decreased by LPS were remarkably increased by LLE. It was found that LLE could relieve LPS-induced oxidative stress by upregulating mRNA and protein expression of Nrf2 and HO-1 in jejunum tissue. In conclusion, LLE alleviates LPS-induced intestinal damage through regulation of the Nrf2/HO-1 signal pathway to alleviate oxidative stress, reducing inflammatory factors and increasing the expression of tight junction proteins in mice.

Modifications of native lotus (Nelumbo nucifera G.) rhizome starch and its overall characterization: A review.

Lotus (Nelumbo nucifera G.) rhizomes are an under-utilized and sustainable starch source that constitutes up to 20 % starch. The review mainly focused on the extraction methods of starch, the chemical composition of LRS, and techno-functional characteristics such as swelling power, solubility, in vitro digestibility, pasting property, and gelatinization is highlighted in LRS review. Lotus rhizome starch (LRS) is also used as a water retention agent, thickening, gelling, stabilizing, and filling in food and non-food applications. Native starch has limited functional characteristics in food applications so by modifying the starch, functional characteristics are enhanced. Single and dual treatment processes are available to enhance microstructural properties, resistant starch, techno-functional, morphological, and, film-forming properties. Compared with other starch sources, there is a lack of systematic information on the LRS. Many industries are interested in developing food products based on starch such as nanoparticles, hydrogels, edible films, and many others. Additionally, there are several recommendations to improve the applications in the food industry. Finally, we provide an outlook on the future possibility of LRS.

Inhibitory mechanism of carboxymethyl chitosan-lotus seedpod oligomeric procyanidin nanoparticles on dietary advanced glycation end products released from glycated casein during digestion.

Lotus seedpod oligomeric procyanidins (LSOPC) are potent inhibitors of advanced glycation end products (AGEs), whose gastrointestinal susceptibility to degradation limits their use in vivo. In this study, carboxymethyl chitosan-lotus seedpod oligomeric procyanidin nanoparticles (CMC-LSOPC NPs) were constructed with a binding ratio of 1:6.51. CMC-LSOPC NPs significantly inhibited the release of AGEs from glycated casein (G-CS) during digestion, increasing the inhibition rate by 25.76% in the gastric phase and by 14.33% in the intestinal phase compared with LSOPC alone. To further investigate the inhibition mechanism, fluorescence microscopy, scanning electron microscopy and FTIR were used to find that CMC-LSOPC NPs could form cohesions to encapsulate G-CS in the gastric phase and hinder G-CS hydrolysis. In the intestinal phase, LSOPC was targeted for release and bound to trypsin through hydrophobic interactions and hydrogen bonding, resulting in protein peptide chain rearrangement, changes in secondary structure and significant reduction in trypsin activity. In addition, CMC-LSOPC NPs increased the antioxidant capacity of digestive fluid and could reduce the oxidative stress in the gastrointestinal tract caused by the release of AGEs. It's the first time that CMC-LSOPC NPs were constructed to enhance the stability of LSOPC during digestion and explain the mechanism by which CMC-LSOPC NPs inhibit the release of AGEs from G-CS in both stomach and intestine. This finding will present a novel approach for reducing AGEs during gastrointestinal digestion.

Molecular Mechanism of Lotus Seedpod Oligomeric Procyanidins Inhibiting the Absorption of Oligopeptide-Advanced Glycation End Products.

Research on advanced glycation end product (AGEs) inhibition has generally focused on food processing, but many protein-AGEs will still be taken. Oligopeptide (OLP)-AGEs, as the main form after digestion, will damage human health once absorbed. Here, we investigated the ability of lotus seedpod oligomeric procyanidins (LSOPC) to inhibit the absorption of the OLP-AGEs and elucidated the underlying mechanism. Our results showed that the inhibition rate of LSOPC on the absorption of OLP-AGEs was about 50 ± 5.38%. 0.1, 0.2, and 0.3 mg/mL could upregulate the expression of ZO-1 and downregulate the expression of PepT1 and clathrin. Molecular docking showed that LSOPC could compete with the binding of OLP-AGEs to PepT1 and AP-2, thus inhibiting the absorption of OLP-AGEs. Furthermore, the interaction of LSOPC with the OLP-AGEs reduced the surface hydrophobicity of OLP-AGEs. It altered the secondary structure of the OLP-AGEs, thus weakening the affinity of the OLP-AGEs to the transporter protein to inhibit the absorption of OLP-AGEs. Together, our data revealed potential mechanisms by which LSOPC inhibit the absorption of OLP-AGEs and opened up new perspectives on the application of LSOPC in reducing the increasing health risks posed by OLP-AGEs.

Structure relationship of non-covalent interactions between lotus seedpod oligomeric procyanidins and glycated casein hydrolysate during digestion.

Procyanidin-amino acid interactions during transmembrane transport cause changes in the structural and physical properties of peptides, which limits further absorption of oligopeptide-advanced glycation end products (AGEs). In this study, glycated casein hydrolysates (GCSHs) were employed to investigate the structure and interaction mechanism of GCSH with lotus seedpod oligomeric procyanidin (LSOPC) complexes in an intestinal environment. LSOPC can interact with GCSH under certain conditions to form hydrogen bonds and hydrophobic interactions to form GCSH-LSOPC complexes. Results showed that procyanidin further leads to the transformation of a GCSH secondary structure and the increase of surface hydrophobicity (H0). The strongest non-covalent interaction between GCSH and (-)-epigallocatechin gallate (EGCG) was due to the polyhydroxy structure of EGCG. Binding site analysis showed that EGCG binds to the internal cavity of P1 to maintain the relative stability of the binding conformation. The antioxidant capacity of GCSH was remarkably elevated by GCSH-LSOPC. This study will provide a new reference for the accurate control of oligopeptide-AGEs absorption by LSOPC in vivo.

Rhamnogalacturonan I-Enriched Pectin, Flavonoids, and Alkaloids from Lotus Leaf Infusion in Regulating Glycolipid Absorption and Metabolism: Isolation, In Vitro Bioactivity Verification, and Structural Characterization.

Lotus leaf is effective in regulating glycolipid absorption and metabolism, but the roles of small-molecule compounds and polysaccharides are unknown. In this study, the small-molecule compounds including flavonoids, alkaloids, and polysaccharides were gradually isolated from lotus leaf infusion by multi-column chromatography and applied to in vitro activity verification and structural characterization. Although flavonoids and alkaloids were effective in inhibiting pancrelipase and α-glucosidase, polysaccharides more effectively bounded bile acids, inhibited cholesterol micelle solubility, and stimulated the growth of Bifidobacterium than lotus leaf infusion. Polysaccharides, presented as spherical conformation in water, were identified as rhamnogalacturonan I-enriched (93%) low-ester pectin with multiple branches mainly composed of arabinan, arabinogalactan-type II, and galactan formed by →3)-Galp-(1→, →5)-Araf-(1→ and →4)-Galp-(1→ residues. Polysaccharides, which were a key constituent of lotus leaf infusion in regulating glycolipid absorption and metabolism, should be paid more attention and developed as a functional food ingredient.

Study on the structure, antioxidant activity and degradation pattern of polysaccharides isolated from lotus seedpod.

The lotus (Nelumbo nucifera Gaertn.) is the largest aquatic vegetable in Asia. The lotus seedpod (LS) is an inedible part of the mature flower receptacle of the lotus plant. However, the polysaccharide isolated from the receptacle has been less studied. The purification of LS resulted in two polysaccharides (LSP-1 and LSP-2). Both polysaccharides were found to be medium-sized HG pectin, with a Mw of 74 kDa. Their structures were elucidated via GC-MS and NMR spectrum and proposed as the repeating sugar units of GalA connected via α-1,4-glycosidic linkage, with LSP-1 having a higher degree of esterification. They have certain content of antioxidant and immunomodulatory activities. The esterification of HG pectin would have an adverse effect on these activities. Furthermore, the degradation pattern and kinetics of LSPs by pectinase conformed to the Michaelis-Menten model. There is a large amount of LS, resulting from the by-product of locus seed production, and thus a promising source for the isolation of the polysaccharide. The findings of the structure, bioactivities, and degradation property provide the chemical basis for their applications in the food and pharmaceutical industries.

Analysis of proanthocyanidins and flavonols in the seedpods of Chinese Antique Lotus: A rich source of antioxidants.

Antique Lotus (Nelumbo) is a perennial aquatic plant with unique historical significance and cultural value, whereas its potential economic value hasn't been fully explored. The present study showed that lotus seedpods had significantly higher antioxidant capacity than other parts by FRAP, ABTS, and ORAC assays and analyzed the proanthocyanidins and flavonols in the seedpods of Antique Lotus. Polyphenols contributed to great antioxidant activity and 51 polyphenols were identified by UPLC-TQ-MS analysis. In which, 27 compounds were identified from lotus seedpods for the first time, including 20 trimers, 5 dimers and 2 tetramers of proanthocyanidin. Total proanthocyanidins explained 70%-90% of the different antioxidant activities and the content of proanthocyanidin trimers showed the strongest correlations with the antioxidant activities. This study provided a fundamental reference for the research of polyphenols in lotus and found that Antique Lotus seedpod extracts have the promising prospects of additives used in feed and food processing.

Biosensor-based active ingredient recognition system for screening TNF-α inhibitors from lotus leaves.

Erhuangquzhi granules (EQG) have been clinically proven to be effective in nonalcoholic steatohepatitis (NASH) treatment. However, the active components and molecular mechanisms remain unknown. This study aimed to screen active components targeting tumor necrosis factor α (TNF-α) in EQG for the treatment of NASH by a surface plasmon resonance (SPR) biosensor-based active ingredient recognition system (SPR-AIRS). The amine-coupling method was used to immobilize recombinant TNF-α protein on an SPR chip, the specificity of the TNF-α-immobilized chip was validated, and nine medicinal herbs in EQG were prescreened. Nuciferine (NF), lirinidine (ID), and O-nornuciferine (NNF) from lotus leaves were found and identified as TNF-α ligands by UPLC‒MS/MS, and the affinity constants of NF, ID, and NNF to TNF-α were determined by SPR experiments (Kd = 61.19, 31.02, and 20.71 µM, respectively). NF, ID, and NNF inhibited TNF-α-induced apoptosis in L929 cells, the levels of secreted IL-6 and IL-1ß were reduced, and the phosphorylation of IKKß and IκB was inhibited in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. In conclusion, a class of new active small-molecule TNF-α inhibitors was discovered, which also provides a valuable reference for the material basis and mechanism of EQG action in NASH treatment.

Explainable machine learning-assisted origin identification: Chemical profiling of five lotus (Nelumbo nucifera Gaertn.) parts.

Five homologous lotus parts, namely, the leaf, stamen, plumule, flower and leaf base, are all ancient nutrient sources, but their chemical differences are poorly understood. Identification of these parts of origin could contribute to determining reasonable edible and/or medicinal applications without misuse/waste risk. The present work aimed to investigate the feasibility of using metabolic profiles coupled with explainable machine learning (ML) for tracing lotus parts of origin. Assisted with molecular networking, 151 compounds were systematically annotated through an untargeted metabolomics approach. Twenty-eight representative constituents were subsequently quantified for the construction of the ML algorithm. Because most ML algorithms are data-driven black boxes with opaque inner workings, the SHaply Additive exPlanation technique was innovatively used to understand model outputs. By offering an integral analytical platform for phytochemical characterization and information interpretation, these results could serve as a basis for an explainable tool for identification of the specific lotus part of origin.

Inhibition of Three Diabetes-Related Enzymes by Procyanidins from Lotus (Nelumbo nucifera Gaertn.) Seedpods.

The inhibitory effects of procyanidins from lotus (Nelumbo nucifera Gaertn.) seedpods on the activities of α-amylase, α-glucosidase and protein tyrosine phosphatase 1B (PTP1B), were studied and compared with those of (+)-catechin, (-)-epicatechin, epigallocatechin gallate (EGCG), procyanidin dimer B2 and trimer C1. The results showed that Lotus procyanidin extract (LPE) significantly inhibited α-amylase, α-glucosidase and PTP1B with IC50 values of 5.5, 1.0, and 0.33 µg/mL, respectively. The inhibition increased with the degree of polymerization and the existence of galloyl or gallocatechin units. Kinetic analysis showed that LPE inhibited α-glucosidase activity in a mixed competitive and noncompetitive mode. Fluorescence quenching revealed that α-glucosidase interacted with LPE or EGCG in an apparent static mode, or the model of "sphere of action". The apparent static (K) and bimolecular (kq) constants were 4375 M-1 and 4.375 × 1011 M-1 s-1, respectively, for LPE and 1195 M-1 and 1.195 × 1011 M-1 s-1, respectively, for EGCG. Molecular docking analysis provided further information on the interactions of (+)-catechin, (-)-epicatechin, EGCG, B2 and C1 with α-glucosidase. It is hypothesized that LPE may bind to multiple sites of the enzyme through hydrogen bonding and hydrophobic interactions, leading to conformational changes in the enzyme and thus inhibiting its activity. These findings first elucidate the inhibitory effect of LPE on diabetes-related enzymes and highlight the usefulness of LPE as a dietary supplement for the prophylaxis of diabetes.

Processing tactics for low-cost production of pure nuciferine from lotus leaf.

Nuciferine is an important drug candidate for the treatment of many diseases. However, there is no general method for its low - cost production. In this work, a feasible method for the production of nuciferine from lotus leaf, using ultrasonic-assisted extraction-solid phase extraction (UAE-SPE) as extraction and cleanup procedure, was developed. Petroleum ether and silica gel have been successfully used as extraction solvent and adsorbent to integrate UAE with SPE, respectively. Except for filtration, no treatment (e.g. concentration and redissolution, etc) was needed on UAE extract before SPE and the effluents obtained in the loading process of SPE could be used as UAE extraction solvent without purification. No obvious decline in the extraction efficiency of UAE and adsorption capacity of SPE was observed at least for 5 runs, which provides a feasible way for the continuous production of nuciferine in industry, i.e. Cyclic UAE-SPE. Moreover, SPE column could be conveniently regenerated and reused without significant decline in its adsorption capacity at least for 5 cycles, which can be used to reduce the cost of the whole system further. In comparison with other cleanup procedures, Cyclic UAE-SPE showed apparent advantages in energy conservation and emission reduction. LLE and crystallization were applied to separate nuciferine from other impurities further. Under optimum conditions, the total recovery rate of nuciferine with a purity over 90.0% from lotus leaf reached 50.1%. All in all, the developed method has advantages in convenient operation, low cost, and high efficiency, thus, is fitting for the production of high purity nuciferine.

Inhibitory Effect of Lotus Leaf-Enriched Flavonoid Extract on the Growth of HT-29 Colon Cancer Cells through the Expression of PI3K-Related Molecules.

Objectives: Lotus leaf is rich in flavonoids, and this study is aimed at examining the inhibitory effect of lotus leaf-enriched flavonoid extract (LLEFE) on HT-29 colon cancer cells through phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) expression regulation. Methods: Lotus leaves were extracted by ethanol and purified using FL-3 macroporous resin to create the LLEFE. HT-29 colon cancer cells were tested using various methods: their proliferation was observed by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, their survival status was observed by fluorescence staining, their oxidative stress level was observed by biochemical kits, and their mRNA expression was determined by quantitative polymerase chain reaction (qPCR) assay. Additionally, the composition of the flavonoids in lotus leaf was determined by HPLC. Results: The results showed that the proliferation of NCM460 normal human colon cells was not affected by 0-500 µg/mL LLEFE but the proliferation of HT-29 human colon cancer cells decreased. LLEFE increased the LDH level in an HT-29 colon cancer cell culture medium; also increased the superoxide dismutase (SOD), catalase (CAT) activities, and glutathione (GSH) level in HT-29 cells; and decreased the malondialdehyde (MDA) level. Further experimental results showed that LLEFE upregulated the expression of SOD1, CAT, and GSH mRNA and downregulated the expression of PI3K, Akt, and mammalian target of rapamycin (mTOR) in HT-29 cells. The high-performance liquid chromatography (HPLC) results showed that kaempferin, hyperoside, astragaloside, phloridzin, and quercetin were the main chemical constituents of lotus leaf. Conclusion: Lotus leaves contain functional flavonoids that inhibit the proliferation of HT-29 colon cancer cells and regulate the expression of PI3K/Akt through five important chemicals.