Upregulated synthesis and production of bioactive compounds in Lotus arabicus L. by in vitro feeding with dried powder of date palm seeds.

BACKGROUND: Plants are considered the primary source of many principal bioactive compounds that have been utilized in a wide range of applications including the pharmaceutical and biotechnological industries. Therefore, there is an imperative need to modulate the production of natural bioactive components. The present study aimed to determine the importance of dried and pulverized date palm seeds (DPS) as a natural elicitor for the synthesis of secondary metabolites in Lotus arabicus L. RESULTS: The presence of various antioxidant compounds, simple sugars, amino acids, fatty acids and reasonable mineral contents was distinct in the phytochemical characterization of DPS. The major components detected in DPS analysis were the 5-(hydroxymethyl) furfural and 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-pyranone. The induced callus of L. arabicus (seven weeks old) was supplemented with DPS at different concentrations (0, 2, 4, 8 and 10 g/l) in culture media. Treatment with 8 g/l DPS induced the highest antioxidant capacity, ascorbic acid content and secondary metabolites (total phenolics and flavonoids) in the produced callus. Stress biomarkers (hydrogen peroxide and malondialdehyde) were found in the control ranges except at 10 g/l DPS. The expression patterns of key genes involoved in secondary metabolism modulation, such as phenylalanine ammonia lyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), flavonol synthase (FLS) and deoxyxylulose phosphate reductoisomerase (DXR), were triggered after DPS treatments. Moreover, the quantitative profiling of phenolic and flavonoid compounds showed that supplementation with DPS, especially at 8 g/l, led to pronounced increases in most of the measured compounds. CONCLUSION: The marked upregulation of eliciting-responsive genes and overproduction of secondary metabolites provide molecular-based evidence for intensifying the principal pathways of phenylpropanoid, flavonoid and terpenoid biosynthesis. Overall, the present in vitro study highlights the stimulating capacity of DPS utilization to improve the bioactive components of L. arabicus at the physiological and molecular levels, enhancing its potential as a medicinal herb.

Cooperative interactions between nitrogen fixation and phosphorus nutrition in legumes.

Legumes such as soybean are considered important crops as they provide proteins and oils for humans and livestock around the world. Different from other crops, leguminous crops accumulate nitrogen (N) for plant growth through symbiotic nitrogen fixation (SNF) in coordination with rhizobia. A number of studies have shown that efficient SNF requires the cooperation of other nutrients, especially phosphorus (P), a nutrient deficient in most soils. During the last decades, great progress has been made in understanding the molecular mechanisms underlying the interactions between SNF and P nutrition, specifically through the identification of transporters involved in P transport to nodules and bacteroids, signal transduction, and regulation of P homeostasis in nodules. These studies revealed a distinct N-P interaction in leguminous crops, which is characterized by specific signaling cross talk between P and SNF. This review aimed to present an updated picture of the cross talk between N fixation and P nutrition in legumes, focusing on soybean as a model crop, and Medicago truncatula and Lotus japonicus as model plants. We also discuss the possibilities for enhancing SNF through improving P nutrition, which are important for high and sustainable production of leguminous crops.

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.

Protective effects of lotus plumule ethanol extracts on bleomycin-induced pulmonary fibrosis in mice.

Pulmonary fibrosis (PF) is a progressive fibrosing disease, characterized by excessive accumulation of extracellular matrix (ECM) and remodeling of the lung architecture, which finally result in respiratory failure. Currently, there is no satisfactory treatment for PF， therefore, the development of effective agents is urgently needed. Lotus plumule, the green embryo of Nelumbo nucifera Gaertn., a plant of the Nymphaeaceae family, is a traditional Chinese food with exceptional nutritional value and its extracts exert prominent anti-inflammatory and anti-fibrotic effects. The aim of the present study was to investigate the inhibitory effects of lotus plumule extracts (LPEs) on bleomycin (BLM)-induced PF in mice. Therefore, enzyme-linked immunosorbent assay, RT-PCR, and western blot analysis were performed. The histopathological examination demonstrated that LPEs could obviously decrease the degree of alveolitis, deposition of ECM and the production of collagen I (Col-I) in the pulmonary interstitium. In addition, the results showed that LPEs markedly alleviated the expression of interleukin (IL)-6, IL-17, transforming growth factor (TGF)-ß, and α-smooth muscle actin (α-SMA). Additionally, the content of Col-I and hydroxyproline (HYP) was also attenuated. In conclusion, LPEs could ameliorate the BLM-induced lung fibrosis, thus suggesting that LPEs could serve as a potential therapeutic approach for PF.

Protective effect of sterols extracted from Lotus plumule on ethanol-induced injury in GES-1 cells in vitro.

In this study, sterols were isolated from Lotus plumule by Soxhlet extraction and saponification and were further characterized by GC-MS analysis. The results showed that the sterols extracted from Lotus plumule mainly contained ß-sitosterol, fucosterol, and campesterol. Models were established in vitro to investigate the protective effects of Lotus plumule sterols (LPSs) on ethanol-induced injury in human gastric epithelium (GES-1) cells. The results showed that appropriate concentrations of LPSs and ß-sitosterol could protect GES-1 cells from ethanol-induced injury by reducing ROS levels, reducing calcium ion release, increasing antioxidant enzyme activity and maintaining mitochondrial membrane potential. Western blot experiment results also showed that appropriate concentrations of LPSs and ß-sitosterol could up-regulate the expression of the anti-apoptotic protein Bcl-2 and down-regulate the pro-apoptotic proteins Bax and caspase-3 in GES-1 cells. Meanwhile, sterol pretreatment groups down-regulated the protein expression levels of p-P38 and p-JNK in ethanol-damaged GES-1 cells and up-regulated the expression level of p-ERK, suggesting that sterols protect GES-1 cells from ethanol-induced damage by regulating the MAPK signaling pathway. Taken together, Lotus plumule sterols could effectively prevent gastric cell damage in vitro and suggest the potential application of LPSs as bioactive ingredients for healthy foods.

Metabolomics Analyses of Cotyledon and Plumule Showing the Potential Domestic Selection in Lotus Breeding.

Lotus (Nelumbo nucifera) seeds are widely consumed as functional food or herbal medicine, of which cotyledon (CL) is the main edible part, and lotus plumule (LP) is commonly utilized in traditional Chinese medicine. However, few studies have been conducted to investigate the chemical components of CL and LP in dry lotus seeds, not to mention the comparison between wild and domesticated varieties. In this study, a widely targeted metabolomics approach based on Ultra Performance Liquid Chromatography-electrospray ionization-Tandem mass spectrometry (UPLC-ESI-MS/MS) was utilized to analyze the metabolites in CL and LP of China Antique ("CA", a wild variety) and Jianxuan-17 ("JX", a popular cultivar). A total of 402 metabolites were identified, which included flavonoids (23.08% to 27.84%), amino acids and derivatives (14.18-16.57%), phenolic acids (11.49-12.63%), and lipids (9.14-10.95%). These metabolites were classified into ten clusters based on their organ or cultivar-specific characters. Most of these metabolites were more abundant in LP than in CL for both varieties, except for metabolites belonging to organic acids and lipids. The analysis of differentially accumulated metabolites (DAMs) demonstrated that more than 25% of metabolites detected in our study were DAMs in CL and LP comparing "JX" with "CA", most of which were less abundant in "JX", including 35 flavonoids in LP, 23 amino acids and derivatives in CL, 7 alkaloids in CL, and 10 nucleotides and derivatives in LP, whereas all of 11 differentially accumulated lipids in LP were more abundant in "JX". Together with the fact that the seed yield of "JX" is much higher than that of "CA", these results indicated that abundant metabolites, especially the functional secondary metabolites (mainly flavonoids and alkaloids), were lost during the process of breeding selection.

Folium nelumbinis (Lotus leaf) volatile-rich fraction and its mechanisms of action against melanogenesis in B16 cells.

This study was aimed at determining the influence of Folium nelumbinis (Lotus leaf) extracts on melanogenesis in vitro models of melanoma cell line. The anticancer activity of four fractions, including petroleum ether (PEE), n-hexane (HE), ethanol (EE), and ethyl acetate (EAE) from F. nelumbinis on B16 cell lines (C57BL/6J melanoma cell), were evaluated after 24 and 48 h treatment. Results showed that PEE as well as volatile-rich fractions of linolenic acid and linolenic acid ethyl ester significantly (p < 0.05) reduced tyrosinase activity and melanin content in B16 melanoma cells model. Meanwhile, PEE and its primarily contained compound triggered apoptosis of B16 cells in a dose-dependent way. These results demonstrated that PEE possessed effective activities against melanin and tyrosinase generations through the induction of apoptosis. Moreover, a relation between the volatile-rich fractions of F. nelumbinis and the anticancer effects was demonstrated as well.

Optimization of lactic acid production using immobilized Lactobacillus Rhamnosus and carob pod waste from the Lebanese food industry.

The valorization of a solid carob waste from the Lebanese industry was investigated by optimizing the production of lactic acid using immobilized Lactobacillus rhamnosus in alginate beads and response surface methodology. The results showed that pH and alginate concentration had a significant effect on the production of lactic acid. The fermentation of non-enriched carob waste juice needed an additional nitrogen source to improve lactic acid production and yield. From extracts with 65 g/L sugars, the optimum conditions were found to be 2% for the concentration of alginate, 4% bacteria cells entrapped in beads, 80 rpm agitation speed and pH 6.4. Lactic acid concentration obtained under these conditions was 22 g/L with a yield of 76.9 g/g consumed sugar and a productivity of 1.22 g/L/h. The use of invertase pretreatment increased lactic acid concentration from 22 to 40 g/L, but reduced yield at 66.6%. Finally, cells immobilized in alginate beads could be used for at least five successive cycles.

Insight into the characterization and digestion of lotus seed starch-tea polyphenol complexes prepared under high hydrostatic pressure.

Complex starch is gaining research attention due to its unique physicochemical and functional properties. In this study, the effects of green tea polyphenols on the properties and digestion of lotus seed starch under high hydrostatic pressure were investigated. The particle size, swelling power, solubility, crystallization, morphology and thermal properties of lotus seed starch were affected by green tea polyphenols. These may be due to the formation of non-inclusive complexes between lotus seed starch and green tea polyphenols. The morphology and green tea polyphenols distribution of the complexes were determined by scanning electron microscopy and confocal laser scanning microscopy. In addition, slow digestion properties of starch were realized under a dynamic in vitro rat stomach-duodenum model and the erosion of granules by amylase gradually decreased by scanning electron microscopy. Furthermore, green tea polyphenols were shown to be able to form V-type inclusion complex with amylose via high hydrostatic pressure.

The role of endogenous thiamine produced via THIC in root nodule symbiosis in Lotus japonicus.

Thiamine is a pivotal primary metabolite which is indispensable to all organisms. Although its biosynthetic pathway has been well documented, the mechanism by which thiamine influences the legume-rhizobium symbiosis remains uncertain. Here, we used overexpressing transgenic plants, mutants and grafting experiments to investigate the roles played by thiamine in Lotus japonicus nodulation. ljthic mutants displayed lethal phenotypes and the defect could be overcome by supplementation of thiamine or by overexpression of LjTHIC. Reciprocal grafting between L. japonicus wild-type Gifu B-129 and ljthic showed that the photosynthetic products of the aerial part made a major contribution to overcoming the nodulation defect in ljthic. Overexpression of LjTHIC in Lotus japonicus (OE-LjTHIC) decreased shoot growth and increased the activity of the enzymes 2-oxoglutarate dehydrogenase and pyruvate dehydrogenase. OE-LjTHIC plants exhibited an increase in the number of infection threads and also developed more nodules, which were of smaller size but unchanged nitrogenase activity compared to the wildtype. Taken together, our results suggest that endogenous thiamine produced via LjTHIC acts as an essential nutrient provided by the host plant for rhizobial infection and nodule growth in the Lotus japonicus - rhizobium interaction.

Differentiation between physical and chemical effects of oil presence in freshly spiked soil during rhizoremediation trial.

Petroleum contamination and its remediation via plant-based solutions have got increasing attention by environmental scientists and engineers. In the current study, the physiological and growth responses of two diesel-tolerant plant species (tolerance limit: 1500-2000 mg/kg), Italian ryegrass (Lolium multiflorum) and Birdsfoot trefoil (Lotus corniculatus), have been investigated in vegetable oil- and diesel oil-amended soils. A long-term (147-day) greenhouse pot experiment was conducted to differentiate the main focus of the study: physical and chemical effects of oil (vegetable and diesel) in freshly spiked soils via evaluating the plant performance and hydrocarbon degradation. Moreover, plant performance was evaluated in terms of seed germination, plant shoot biomass, physiological parameters, and root biomass. Addition of both diesel oil and vegetable oil in freshly spiked soils showed deleterious effects on seedling emergence, root/shoot biomass, and chlorophyll content of grass and legume plants. Italian ryegrass showed more sensitivity in terms of germination rate to both vegetable and diesel oil as compared to non-contaminated soils while Birdsfoot trefoil reduced the germination rate only in diesel oil-impacted soils. The results of the current study suggest that both physical and chemical effects of oil pose negative effects of plant growth and root development. This observation may explain the phenomenon of reduced plant growth in aged/weathered contaminated soils during rhizoremediation experiments.

Oligomer Procyanidins from Lotus Seedpod Regulate Lipid Homeostasis Partially by Modifying Fat Emulsification and Digestion.

Dietary polyphenols have shown hypolipidemic effects by reducing triglyceride absorption. The mechanisms may involve modifying fat emulsion during digestion in the gastrointestinal tract and suppressing lipase during hydrolysis in the small intestine. In an in vivo study, lotus seedpod oligomeric procyanidin (LSOPC) decreased total serum triglyceride and total cholesterol and elevated the high-density lipoprotein level in the hyperlipidemic rat model. In addition, LSOPC suppressed de novo lipogenesis-related gene expressions. In an in vitro study, the LSOPC-enriched emulsion decreased the mean droplet size from 0.36 to 0.33 µm and increased the viscosity of the emulsion. Moreover, the LSOPC-enriched emulsion improved the antioxidant properties. A digestion model was developed and showed that the particle size of the LSOPC-enriched emulsion increased in the oral cavity. However, an increase and then a significant drop of the particle size was measured in the stomach and small intestine. The free fatty acid release rate was decreased in the LSOPC-enriched emulsion partly ascribed to the inhibition of lipase by LSOPC.

Antioxidant and anti-inflammatory properties of flavonoids from lotus plumule.

The antioxidant and anti-inflammatory mechanisms of action of flavonoids in lotus plumule were systematically analyzed using radical scavenging assays and ELISA kits. By this means, flavonoids displayed significant antioxidant activity by donating electron, H atom as well as capturing DPPH and ABTS+ free radicals, and anti-inflammatory effect by inhibiting the production of the inflammatory mediators (NO radicals, PGE2 and TNF-α) and pro-inflammatory cytokines (IL-1ß and IL-6). Meanwhile, the bioactive components against inflammation targeting COX-2 were also revealed using ultrafiltration coupled to LC-MS (UF-LC/MS). In this way, 12 components showing specific binding to COX-2 were screened out and identified. The structure-activity relationships suggested that flavonoids O-glycosides displayed comparable binding affinities to COX-2 compared with flavonoids C-glycosides and could be considered as the main active components. This study will provide valuable information for the further exploration of lotus plumule as functional foods or in pharmaceutical industries in the near future.

Genome-wide transcriptional changes in type 2 diabetic mice supplemented with lotus seed resistant starch.

Resistant starch has been studied extensively for its hypoglycemic activity, while its underlying molecular mechanism is not fully understood. In this study, we investigated the hypoglycemic effect of different doses of lotus seed resistant starch (LSRS) supplementation on type 2 diabetic mice and elucidated the molecular basis of its hypoglycemic effect. LSRS supplementation significantly reduced blood glucose level by 16.0%-33.6%, recovered serum insulin level by 25.0%-39.0% and improved lipid metabolism disorder in the diabetic mice. The genome-wide expression patterns in pancreatic tissue were analyzed, and 511 differentially expressed genes (DEGs) were identified. The analysis results of gene ontology (GO) functions and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways indicated that the protective effect of LSRS supplementation was most likely driven by modulating expression levels of various key factors involved in insulin secretion, insulin signal transmission, cell apoptosis, antioxidant activity and p53 signaling pathways.

Genistein-Specific G6DT Gene for the Inducible Production of Wighteone in Lotus japonicus.

Prenylated isoflavonoids have been found in several legume plants, and they possess various biological activities that play important roles in both plant defense and human health. However, it is still unknown whether prenylated isoflavonoids are present in the model legume plant Lotus japonicus. In the present study, we found that the prenylated isoflavonoid wighteone was produced in L. japonicus when leaf was supplemented with genistein. Furthermore, a novel prenyltransferase gene, LjG6DT, was identified, which shared high similarity with and was closely related to several known prenyltransferase genes involved in isoflavonoid biosynthesis. The recombinant LjG6DT protein expressed in yeast exhibited prenylation activity toward genistein as an exclusive substrate, which produced wighteone, a prenylated genistein at the C-6 position that occurs normally in legume plants. The LjG6DT-green fluorescent protein (GFP) fusion protein is targeted to plastids. The transcript level of LjG6DT is induced by glutathione, methyl jasmonate and salicylic acid, implying that LjG6DT is involved in stress response. Overexpression of LjG6DT in L. japonicus hairy roots led to increased accumulation of wighteone when genistein was supplied, indicating that LjG6DT is functional in vivo. Feeding assays with the upstream intermediate naringenin revealed that accumulation of wighteone in L. japonicus was dependent on genistein supplementation, and accumulation of wighteone is competed by genistein methylation. This study demonstrated that phytoalexin wighteone is inducibly produced in L. japonicus, and it provides new insight into the biosynthesis and accumulation of prenylated isoflavonoids in legume plants.

Biosynthesis of Ag/reduced graphene oxide/Fe(3)O(4) using Lotus garcinii leaf extract and its application as a recyclable nanocatalyst for the reduction of 4-nitrophenol and organic dyes.

In current research, Ag/RGO/Fe3O4 nanocomposite was synthesized through applying the aqueous extract of Lotus garcinii leaves through a two-step method. The plant extract was utilized as agents for reduction and stabilization in biosynthesizing nanocomposite. The green synthesized nanocatalyst was characterized by FT-IR, FE-SEM, EDX, XRD, TEM, UV-Vis absorption spectroscopy. The catalytic activity of the biosynthesized nanocomposite was studied by reducing different colored solutions contained organic pollutants such as 4-nitrophenol (4-NP), Congo red (CR) and Rhodamine B (RhB). In order to verify commercial applications of the prepared heterogeneous nanocatalyst, reusability and recoverability tests were performed in five successive catalytic reactions.

Structural characteristics and crystalline properties of lotus seed resistant starch and its prebiotic effects.

Lotus seed resistant starch (LRS) is a type of retrograded starch that is commonly known as resistant starch type 3 (RS3). The structural and crystalline properties of unpurified LRS (NP-LRS3), enzyme purified LRS after drying (GP-LRS3), and enzyme purified LRS (ZP-LRS3) were characterized. The result showed that the molecular weights of NP-LRS3, GP-LRS3, and ZP-LRS3 were 0.102 × 10(6), 0.014 × 10(6), and 0.025 × 10(6)Da, respectively. Compared with native starch and high amylose maize starch (HAMS), LRS lacked the polarization cross and the irregularly shaped LRS granules had a rougher surface, B-type crystal structure, and greater level of molecular order. The FT-IR measurements indicated no differences in the chemical groups. Analysis by (13)C NMR indicated an increased propensity for double helix formation and higher crystallinity in LRS than in the two other types of starch. Moreover, LRS was more effective than either glucose or HAMS in promoting the proliferation of bifidobacteria.

The cis-acting CTTC-P1BS module is indicative for gene function of LjVTI12, a Qb-SNARE protein gene that is required for arbuscule formation in Lotus japonicus.

The majority of land plants live in symbiosis with arbuscular mycorrhizal fungi from the phylum Glomeromycota. This symbiosis improves acquisition of phosphorus (P) by the host plant in exchange for carbohydrates, especially under low-P availability. The symbiosome, constituted by root cortex cells accommodating arbuscular mycorrhizal fungal hyphae, is the site at which bi-directional exchange of nutrients and metabolites takes place. Uptake of orthophosphate (Pi) in the symbiosome is facilitated by mycorrhiza-specific plant Pi transporters. Modifications of the potato Pi transporter 3 (StPT3) promoter were analysed in transgenic mycorrhizal roots, and it was found that the CTTC cis-regulatory element is necessary and sufficient for a transcriptional response to fungal colonization under low-Pi conditions. Phylogenetic footprinting also revealed binary combination of the CTTC element with the Pi starvation response-associated PHR1-binding site (P1BS) in the promoters of several mycorrhiza-specific Pi transporter genes. Scanning of the Lotus japonicus genome for gene promoters containing both cis-regulatory elements revealed a strong over-representation of genes involved in transport processes. One of these, LjVTI12, encoding a member of the SNARE family of proteins involved in membrane transport, exhibited enhanced transcript levels in Lotus roots colonized with the arbuscular mycorrhizal fungus Glomus intraradices. Down-regulation of LjVTI12 by RNA interference resulted in a mycorrhiza-specific phenotype characterized by distorted arbuscule morphology. The results highlight cooperative cis-regulation which integrates mycorrhiza and Pi starvation signaling with vesicle trafficking in symbiosome development.

Water stress induces a differential and spatially distributed nitro-oxidative stress response in roots and leaves of Lotus japonicus.

Water stress is one of the most severe problems for plant growth and productivity. Using the legume Lotus japonicus exposed to water stress, a comparative analysis of key components in metabolism of reactive nitrogen and oxygen species (RNS and ROS, respectively) were made. After water stress treatment plants accumulated proline 23 and 10-fold in roots and leaves respectively, compared with well-watered plants. Significant changes in metabolism of RNS and ROS were observed, with an increase in both protein tyrosine nitration and lipid peroxidation, which indicate that water stress induces a nitro-oxidative stress. In roots, ·NO content was increased and S-nitrosoglutathione reductase activity was reduced by 23%, wherein a specific protein nitration pattern was observed. As part of this response, activity of NADPH-generating dehydrogenases was also affected in roots resulting in an increase of the NADPH/NADP(+) ratio. Our results suggest that in comparison with leaves, roots are significantly affected by water stress inducing an increase in proline and NO content which could highlight multiple functions for these metabolites in water stress adaptation, recovery and signaling. Thus, it is proposed that water stress generates a spatial distribution of nitro-oxidative stress with the oxidative stress component being higher in leaves whereas the nitrosative stress component is higher in roots.

Combined topical application of lotus and green tea improves facial skin surface parameters.

No study has yet determined the anti-wrinkle efficacy of green tea plus lotus in Asian subjects using skin image analysis technique. In this study, the efficacy of two cosmetic active formulations intended for the treatment of facial wrinkles (green tea and lotus extract) has been evaluated in healthy subjects using a non-invasive device, the Visioscan(®) VC, and software for surface evaluation of living skin (SELS). Thirty-three healthy Asian subjects, all men, were enrolled after consent in a placebo-controlled comparative study with a split face design. One group applied multiple emulsions with green tea. The second group applied multiple emulsions with lotus extract, while a third group applied a multiple emulsion with a combination of both extracts. In all three groups, active formulations were applied to one side of the face and the placebo to the other side, once daily over the 60-day treatment course. Non-invasive measurements were performed at baseline and on days 30 and 60. Interesting and significant improvements were observed for the treatment effects on skin roughness (SEr), scaliness (SEsc), smoothness (SEsm), and wrinkling (SEw). For example, a 49.99% improvement in skin smoothness (SEsm) from baseline value and -23.22% and perfection in facial wrinkles (SEw) substantiated that combined treatment is superior over single treatments. Green tea and lotus combined in multiple emulsions brought a superior synergistic anti-aging effect. We conclude that diverse anti-oxidant constituents in both plants have a potential influence on skin surface parameters, thus indicating these plants as the future of new anti-aging products.