Pyruvate stimulates transamination of leucine into α-ketoisocaproic acid and supports 3-methylbutanal production by Lactococcus lactis.

AIM: To investigate the effect of pyruvate and glucose on leucine transamination and 3-methylbutanal production by Lactococcus lactis, including the comparison with cells possessing glutamate dehydrogenase (GDH) activity. METHODS AND RESULTS: L. lactis cells were incubated in chemically defined medium (CDM) with the pH controlled at 5.2 to mimic cheese conditions. Pyruvate supplementation stimulated the production of the key flavour compound 3-methylbutanal by 3 to 4 times after 72 hours of incubation. Concurrently, alanine production increased, demonstrating the involvement of pyruvate in transamination reactions. Glucose-metabolising cells excreted α-ketoisocaproic acid and produced even 3 times more 3-methylbutanal after 24 hours than pyruvate-supplemented cells. Conjugal transfer technique was used to transfer the plasmid pGdh442 carrying the gdh gene encoding for GDH to L. lactis. Introducing GDH did not stimulate the excretion of α-ketoisocaproic acid and the production of 3-methylbutanal. CONCLUSIONS: These results demonstrate that Lactococcus uses pyruvate to transaminate leucine into α-ketoisocaproic acid which supports 3-methylbutanal production. Surprisingly, GDH activity did not stimulate leucine transamination and 3-methylbutanal production.

Evaluation of the effect of dietary supplementation with Allium mongolicum regel bulb powder on the volatile compound and lipid profiles of the longissimus thoracis in Angus calves based on GC-IMS and lipidomic analysis.

The effect of A. mongolicum Regel bulb powder (AMRP) supplementation on the flavour of beef from Angus calves has not been investigated thus far. We used GC-IMS and untargeted lipidomics techniques to examine the volatile compound and lipid metabolic profiles and reveal the effects of dietary AMRP supplementation on the flavour of beef. A total of 6 characteristic volatile compounds and 30 key lipid compounds were identified in the AMRP treatment group. AMRP promoted the release of triglycerides and phosphatidylinositols from beef and accelerated the production of volatile compounds such as ethyl acetate, 1-penten-3-one, and tetrahydrofurane, and the production of these three characteristic volatile compounds was significantly correlated with the UFAs in triglycerides according to correlation analysis. In summary, dietary AMRP supplementation had a positive effect on the flavour of beef, and these findings provide a theoretical basis for the development and utilisation of AMRP as a feed additive.

Using machine-learning approaches to investigate the volatile-compound fingerprint of fishy off-flavour from beef with enhanced healthful fatty acids.

Machine learning classification approaches were used to discriminate a fishy off-flavour identified in beef with health-enhanced fatty acid profiles. The random forest approach outperformed (P < 0.001; receiver operating characteristic curve: 99.8 %, sensitivity: 99.9 % and specificity: 93.7 %) the logistic regression, partial least-squares discrimination analysis and the support vector machine (linear and radial) approaches, correctly classifying 100 % and 82 % of the fishy and non-fishy meat samples, respectively. The random forest algorithm identified 20 volatile compounds responsible for the discrimination of fishy from non-fishy meat samples. Among those, seven volatile compounds (pentadecane, octadecane, γ-dodecalactone, dodecanal, (E,E)-2,4-heptadienal, 2-heptanone, and ethylbenzene) were selected as significant contributors to the fishy off-flavour fingerprint, all being related to lipid oxidation. This fishy off-flavour fingerprint could facilitate the rapid monitoring of beef with enhanced healthy fatty acids to avoid consumer dissatisfaction due to fishy off-flavour.

Design of experiment approach to boost volatile production from kiwi byproducts.

Design of Experiments (DoE), is a tool to explore relationships between factors and responses of a system. DoE and response surface methodology are increasingly used in different fields, but their application are limited in the valorization of residual biomass and agro-industrial by-products. Agro-industrial biomass residues can be eco-friendly converted into high-value compounds through bioprocesses. This approach identified key factors and predicted optimal conditions for enhancing microbial growth and the production of specific compounds or volatile classes. Lactiplantibacillus plantarum 4193 and Lacticaseibacillus paracasei 2243, were identified as the best starters while the production of methyl heptenone is influenced by fermentation time and pH. This out-turn in the generation of aromatically rich biomass, which can be utilised as a food ingredient or for the extraction of specific volatile compounds, and employed as flavouring agents. This study underlines the potential of fermentation in maximizing the value of unripe kiwi biomass.

Mapping taste and flavour traits to genetic markers in lettuce Lactuca sativa.

Lettuce is the most highly consumed raw leafy vegetable crop eaten worldwide, making it nutritionally important in spite of its comparatively low nutrient density in relation to other vegetables. However, the perception of bitterness caused by high levels of sesquiterpenoid lactones and comparatively low levels of sweet tasting sugars limits palatability. To assess variation in nutritional and taste-related metabolites we assessed 104 members of a Lactuca sativa cv. Salinas x L. serriola (accession UC96US23) mapping population. Plants were grown in three distinct environments, and untargeted NMR and HPLC were used as a rapid chemotyping method, from which 63 unique Quantitative Trait Loci (QTL) were identified. We were able to identify putative regulatory candidate genes underlying the QTL for fructose on linkage group 9, which accounted for up to 36 % of our population variation, and which was stable across all three growing environments; and for 15-p-hydroxyyphenylacetyllactucin-8-sulfate on linkage group 5 which has previously been identified for its low bitterness, while retaining anti-herbivory field effects. We also identified a candidate gene for flavonoid 3',5'- hydroxylase underlying a polyphenol QTL on linkage group 5, and two further candidate genes in sugar biosynthesis on linkage groups 2 and 5. Collectively these candidate genes and their associated markers can inform a route for plant breeders to improve the palatability and nutritional value of lettuce in their breeding programmes.

Textured vegetable protein as a partial replacement for lean meat in salami analogues: Perspectives on physicochemical properties, flavour and proteome changes.

Integrating plant proteins into meat products offers a sustainable way to reduce the environmental impact of meat consumption while satisfying the growing flexitarian population. This study explored the effects of textured vegetable proteins (TVPs) on the physico-chemical attributes and flavour profile of hybrid salamis using 4D label-free proteomics. Results showed that hybrid salamis had lower pH, reduced water activity and increased weight loss compared with traditional salamis, along with greater hardness and a slightly rough, porous texture with a filamentous structure. TVPs substantially modified crucial meaty flavour compounds (nitrogen oxides, sulfides and pyrazine), increasing heightening sourness and bitterness while diminishing umami. Proteomic analysis revealed significant upregulation of myosin and actin in hybrid salamis; notably, these proteins were involved in glycerol-3-phosphate dehydrogenase activity and calcineurin-mediated signalling, underscoring their role in flavour enhancement. Therefore, hybrid salamis offer an attractive alternative to traditional salamis by merging meat-like taste and texture with plant protein.

Advantages of UHT in retaining coconut milk aroma and insights into thermal changes of aroma compounds.

Coconut milk products are susceptible to bacterial damage, necessitating sterilization methods that often compromise nutrient and aroma integrity. This study investigates the effects of different thermal sterilisation methods on coconut milk aroma using headspace gas chromatography-ion mobility spectrometry (HS-GC-IMS) and headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS). We assessed the impact of pasteurisation (PAS, 70 °C, 25 min), high-temperature sterilisation (HTS, 121.1 °C, 15 min), and ultra-high temperature sterilisation (UHT, 130 °C, 5 s) through clustered heat maps and correlation analyses. Significant differences were observed (p < 0.05), with 37 and 52 substances detected by HS-GC-IMS and HS-SPME-GC-MS, respectively, identifying 12 key aroma compounds. UHT treatment primarily reduced 8 acids, maintaining a compositional structure and sensory profile similar to raw coconut milk. PAS and HTS treatments decreased the sensory intensity of overall coconut milk aroma, creamy, and floral notes, correlating with the presence of 2-heptanol, nonanal, 4-methylvaleric acid, and 2-tridecanone. These methods increased cooked notes, associated with 5-methyl-3-heptanone, 3-butyn-1-ol, hydroxyacetone, and acetoin. Rancidity was linked to acids such as isobutyric acid, isovaleric acid, and heptanoic acid, with high temperatures effectively reducing these compounds. Prolonged temperature changes in PAS and HTS accelerated lipid oxidative degradation and the Maillard reaction, involving free fatty acids in the formation of alcohols, aldehydes, esters, and lactones. These findings provide a theoretical basis for studying coconut milk flavour deterioration.

Omics analysis of key pathway in flavour formation and B vitamins synthesis during chickpea milk fermentation by Lactiplantibacillus plantarum.

Chickpea milk is a nutrient-rich plant-based milk, but its pronounced beany flavour limits consumer acceptance. To address this issue, chickpea milk was fermented using two strains of Lactiplantibacillus plantarum, FMBL L23251 and L23252, which efficiently utilize chickpea milk. L. plantarum FMBL L23251 demonstrated superior fermentation characteristics. Fermentation with L. plantarum FMBL L23251 resulted in a 1.90-fold increase in vitamin B3 (271.66 ng/ml to 516.15 ng/ml) and a 1.58-fold increase in vitamin B6 (91.24 ng/ml to 144.16 ng/ml) through the L-aspartic acid pathway and the 1-deoxy-D-xylulose-5-phosphate (DXP)-independent pathway, respectively. Furthermore, L. plantarum FMBL L23251 effectively removed beany flavours due to its enhanced pathway for pyruvate metabolism. The main aldehydes are converted into corresponding alcohols or acids, resulting in 87.74 % and 96.99 % reductions in hexanal and 2-pentyl-furan, respectively. In summary, the fermentation of L. plantarum FMBL L23251 generated fermented chickpea milk that is rich in B vitamins and provides a better flavour.

Effect of yeast (Saccharomyces cerevisiae) fermentation on conformational changes in pig liver proteins and their ability to bind to characteristic aldehydes.

This study aimed to explore the potential of a fermentation technology to reduce off-flavour perception and its underlying mechanisms. Results revealed that yeast fermentation (YF) significantly ameliorated the off-flavour of pig liver (p < 0.05). Specifically, YF pre-treatment decreased the relative abundance of α-helix and fluorescence intensity while increasing the surface hydrophobicity and SS level and loosening the microstructure of myofibrillar proteins (MPs) in pig liver. Additionally, the appropriate fermentation treatments enhanced the MP-aldehyde binding capacity by 0.25-1.30 times, demonstrating that YF-induced conformational modifications in pig liver proteins made them more prone to interacting with characteristic aldehydes. Moreover, molecular docking results confirmed that hydrophobic interactions are the primary drivers of MP-aldehyde binding. These findings suggest that YF technology holds immense promise for modulating off-flavour perception in liver products by altering protein conformation.

Evaluation of ultrasound-assisted tomato sour soup marination on beef: Insights into physicochemical, sensory, microstructural, and flavour characteristics.

This study evaluated the quality attributes of tomato sour soup marinade and investigated the effects of ultrasound-assisted marination on the physicochemical properties, microstructure, texture, sensory quality, and flavour profile of beef. The results showed that tomato sour soup significantly increased the marinade absorption rate and improved beef tenderloin's physicochemical properties, texture, and flavour attributes compared to static brine (P < 0.05), with organic acids playing an essential role in the marinade tenderisation process. Compared to static sour soup marination, ultrasound treatment significantly accelerated the marination process, reducing beef's shear force, hardness, and chewiness while increasing its tenderness. Microstructural observations revealed that sour soup marination induced a fragmented and irregular muscle fibre structure. Furthermore, sour soup marination significantly increased the relative concentrations of volatile flavour compounds, including alkanes, organic sulphides, alcohols, aldehydes, and aromatic compounds. Appropriate ultrasound treatment positively affects the texture and flavour characteristics of beef marinated with tomato sour soup, and the optimal approach was 320 W ultrasound treatment for 60 min. Overall, tomato sour soup improved beef's textural and flavour attributes, while ultrasound-assisted marination is an effective processing method to improve the quality of meat products.

Preparation of ß-Cyclodextrin(CD)/Flavour CD Powder and Its Application on Flavour Improvement of Regular Coffee.

To improve the overall sensory evaluation of regular coffee, a mixture of ß-CD/flavour CD powder was prepared by a freeze-drying method. Cyclodextrin inclusion complexes consist of eight compounds that are naturally present in coffee, specifically: 2,5-dimethylpyrazine, benzaldehyde, citral, linalool, limonene, phenethyl acetate, furfural, and ethyl acetate. These eight compounds naturally occur in coffee, making them safer than using other compounds. Moreover, these eight compounds are the primary active ingredients in coffee, significantly influencing its flavour profile. Therefore, choosing to complex these eight compounds with cyclodextrins can effectively enhance the taste of the coffee. XRD, FT-IR, and SDE-GC-FID were presented to study the formation of inclusion CD powder, the storage stability, chemical composition changes, and safety. Results show that by the cyclodextrin method of freeze-drying, the CD powder showed a stable encapsulated structure and increased stability of flavour compounds. Based on the coffee aroma analysis results, prepared CD powder can enhance the coffee's aroma score by 3.0-4.0 points and increase the flavour score by 2.1-3.5 points, and it can achieve preservation for a minimum of 181 days at 25 °C. Furthermore, under the requirements of the China national standard for additives, the mixture of ß-CD/flavour CD powder was used for the cup testing with four regular coffees to obtain improved coffees. With the full score is 10, improved coffees could score extra 3.0-4.0 points on aroma and 2.1-3.5 on flavour compared to regular coffee. In addition, the CD powder also improves the quality of the coffee in terms of aftertaste, body, and sweetness. Overall, ß-CD/flavour CD powders provide several advantages over the currently popular coffee bean processing methods, including improved reproducibility, enhanced controllability, and increased flexibility, while prioritizing safety. And it should be explored further with appropriate compounds given its potential for coffee aroma modulation.

Improvement of rheological and sensory properties of Lactobacillus helveticus fermented milk by prebiotics.

The fermentation characteristics and aroma-producing properties of Lactobacilli could influence the flavour quality of fermented milk, an important influencing factor of consumers' preference. In this study, fermented milk was prepared using Lactobacillus helveticus and the dynamic changes in the sensory quality of fermented milk throught fermentation were to assess the dynamic changes in sensory quality of fermented milks throughout the fermentation process, including rheological properties and flavour profiles. Styrene, linalool, octanoic acid, and 1-nonanol were considered as the key flavour components during fermentation. The quality of the fermented milk tends to be stabilized after 24-h, showing the minimal off-flavour at 48-h and optimal fermented aroma at 72-h. Three prebiotics (inulin, Galactooligosaccharides and inulin mixed with Galactooligosaccharides) were added to Lactobacillus spiralis fermented milk separately, and the results showed that inulin mixed with Galactooligosaccharides was the most effective group in improving the organoleptic quality of the fermented milk. Overall, the experimental results provide deeper insights into the release and retention of aroma compounds during fermentation and scientific reference for broadening the application of prebiotics and flavour-producing Lactobacilli in fermented milk processing.

Enhancement of culled ewes' meat quality: Effects of aging method and time.

This study assesses the impact of wet and dry aging, over 35 days, on various physico-chemical, colorimetric, oxidative, volatolomic, and sensory attributes of meat from culled ewes. Water holding capacity of dry-aged (DA) meat increased from day 28 and was significantly higher than wet-aged (WA) meat. Cooking loss of DA meat decreased, and it was lower than that of WA meat. Warner Bratzler shear force increased in DA meat but decreased in WA meat during aging. Higher oxidation product concentration in DA meat likely results from oxygen exposure. Some aldehydes and ketones peaked at day 7 in DA meat, surpassing levels in WA meat. Overall liking scores favored DA meat at day 14 and 21 but declined from day 14 to 35, coinciding with increased pentanal content. Dry aging could improve the acceptability of culled ewes' meat more than wet aging, but in short aging time (14 days).

Reduction of antinutrients and off-flavour in kidney bean flour by acidic and alkaline reactive extrusion.

The presence of antinutrients and undesirable flavours in kidney bean flour poses challenges to consumer acceptance. Although extrusion can mitigate antinutrients to some extent, its impact on reducing beany flavour in bean flour remains underexplored. This study investigated the effects of injecting acetic acid or sodium carbonate solutions at three concentration levels (0.05, 0.1, 0.15 mol/L), in conjunction with three temperature profiles (40/60/80/80/90, 40/60/80/90/110, 50/70/90/110/130 °C) and two feed moisture levels (25, 30 %), on the removal of antinutrients (condensed tannins, trypsin inhibitor activity, phytic acid, raffinose family oligosaccharides) and reduction of volatile compounds that contribute to beany flavour in whole kidney bean flour. The results showed that all concentrations of acetic acid and sodium carbonate solutions effectively reduced condensed tannins compared to water, especially at 130 °C extrusion temperature. Introducing acetic acid and sodium carbonate solutions at a concentration of 0.15 mol/L led to 72 and 90 % reduction of total raffinose family oligosaccharide content, respectively, in contrast to the 17 % reduction observed with water alone. The incorporation of sodium carbonate solution reduced the total volatile compounds by 45-58 % as compared with water (23-33 %) and acetic acid (11-27 %). This reduction was primarily due to the reduction of aldehydes, alcohols, and aromatic hydrocarbons. These results indicate that injecting sodium carbonate solution during extrusion can effectively reduce antinutrients and beany flavour compounds in kidney bean flour.

Characterisation of Pea Milk Analogues Using Different Production Techniques.

Research background: Among legumes, peas are characterised by their high protein content, low glycaemic index and exceptional versatility. However, their potential as a food is often compromised by their undesirable off-flavour and taste. Hence, this study focuses on minimising off-flavours through simple pretreatments with the aim of improving the potential for the production of pea milk analogues. Pea milk analogues are a burgeoning type of plant-based milk alternatives in the growing plant-based market. Experimental approach: Pea seeds were subjected to different pretreatments: (i) dry milling, (ii) blanching followed by soaking in alkaline solution and subsequent dehulling and (iii) vacuum. Typical physicochemical properties such as pH, viscosity, colour, titratable acidity and yield were measured to obtain a brief overview of the products. Consumer acceptance test, descriptive sensory analysis, gas chromatography-mass spectrometry and gas chromatography-olfactometry were used to map the complete sensory profile and appeal of the pea milk substitutes. Results and conclusions: The L* values of the pea milk analogues were significantly lower than those of cow's milk, while a*, b*, viscosity and pH were similar. In the descriptive sensory analysis, sweet, astringent, pea-like, cooked, hay-like, boiled corn and green notes received relatively higher scores. The vacuum-treated pea milk analogues received higher scores for flavour and overall acceptability in the consumer acceptance test. The pretreatments resulted in significant changes in the volatile profiles of the pea milk analogues. Some volatiles typically associated with off-flavour, such as hexanal, were found in higher concentrations in blanched pea milk analogues. Among the applied pretreatments, vacuum proved to be the most effective method to reduce the content of volatile off-flavour compounds. Novelty and scientific contribution: This study stands out as a rare investigation to characterise pea milk analogues and to evaluate the impact of simple pretreatments on the improvement of their sensory properties. The results of this study could contribute to the development of milk alternatives that offer both high nutritional value and strong appeal to consumers.

Pomegranate Quality from Consumers' Perspective: Drivers of Liking, Preference Patterns, and the Relation between Sensory and Physico-Chemical Properties.

Acquiring information on consumer preferences for the sensory properties of pomegranates is a cue for breeding programmes to set their quality targets and promote the consumption of this particularly healthy fruit. In this study, a total of 12 pomegranate varieties were evaluated, including commercial (Valenciana, Illina, Tastem, Rugalate, Wonderful, Mollar 49 y Mollar 45) and new varieties (Ref 102, Ref 383, H3/27, and D27/12). For the first time, consumers not only scored their acceptance of different pomegranate varieties but also described their sensory properties using CATA questions. This approach allowed us to identify the main drivers of liking, preference patterns, and the relationship between sensory and physico-chemical properties. Of all the sensory attributes, acidity intensity was revealed as the main driver of liking/disliking, and two different consumer preference patterns were identified: 'low acid pomegranate lovers' and 'acid pomegranate lovers'. Seed properties like the intensity of woody flavour and seed/aril ratio were also key drivers for preferences. A relationship between sensory and physico-chemical properties was also established. Interestingly, sweetness perception correlated more strongly with low titratable acidity levels than with high total soluble solids levels, corroborating that acidity level is a key measurement for quality assessments. 'H3/27' was the most promising of the new varieties for having the well-appreciated internal properties of the 'Mollar' varieties and external and internal red colouration, which makes it much more appealing to consumers. This study shows that there is still room on the market for pomegranate varieties with very different sensory properties and highlights the need to develop sensory labels that help consumers make the right choices.

Analysis of differences in physicochemical properties of different sorghum varieties and their influence on the selection of raw materials for winemaking.

Sorghum is one of the oldest crops in the world, an important grain crop in northern China, and a major raw material in the liquor-brewing industry. The physicochemical properties, cooking characteristics, and starch quality of sorghum seeds considerably affect the liquor-brewing process.To select suitable sorghums for liquor brewing and to determine the cooking characteristics and starch physicochemical properties of different sorghum varieties, 30 types of sorghum were used in this study, and their compositions were compared; six types of sorghum were further studied for their cooking quality and starch physicochemical and pasting characteristics. Gas chromatography time of flight mass spectrometry was used to analyse the cooking aroma of sorghum seeds. Additionally, scanning electron microscopy, a rapid visco analyser, and a differential calorimetric scanner were used to analyse the microstructure of sorghum starch, starch pasting characteristics, and thermodynamic properties, respectively. The results revealed that the water absorption and saccharification forces of glutinous sorghum were higher than those of japonica sorghum and that the aroma substances were significantly different. Glutinous sorghum starch had high crystallinity, freeze-thaw stability, and enthalpy, thus indicating its structural stability. This study provides a theoretical basis for the selection of wine raw materials in the future.

Targeted-metabolomic and untargeted-proteomic approaches reveal the effects of muscle fibre type and postmortem ageing on taste-active compounds in beef.

The taste of beef is caused by taste-active compounds detected in the mouth during mastication. We hypothesised that the concentration of taste-active compounds in beef is influenced by muscle-fibre-type and postmortem ageing. To test this, and unravel the underlying mechanisms, we investigated the taste-active compounds, and proteomic profiles, in beef masseter [oxidative muscle, all type I fibres) and cutaneous trunci (glycolytic muscle, mostly type II fibres) before and after 14-days postmortem ageing. Our results showed that nucleotides were initially higher and degraded slower in cutaneous trunci (P < 0.05 for both), which could be explained by the profile of nucleotide metabolism enzymes. In contrast, free amino acids were initially higher and increased more in masseter compared to cutaneous trunci (P < 0.05 for all), which might be explained by the profile and activity of proteases in these two muscles. Our results indicate the taste of beef is affected by the muscle-fibre-type and postmortem ageing.

Characterization of flavour components and identification of lipid flavour precursors in different cuts of pork by phospholipidomics.

The lipids and volatile compounds in pork from different parts, including the loin, belly, shoulder and hind leg were analyzed by triple quadrupole tandem time-of-flight mass spectrometer (Q-TOF/MS) and gas chromatography-olfactometry-mass spectrometry (GC-O-MS), respectively. Partial least squares regression (PLSR) and Pearson correlation analysis were utilized to establish the relationship between the lipids and volatile compounds. A total of 8 main flavour substances, 38 main phospholipids, and 32 main fatty acids were identified. The results showed that the key flavour compounds were mainly derived from unsaturated fatty acids and phospholipids containing unsaturated fatty acids, including oleic acid (C18:2n6c), α-Linolenic acid (C18:3n3), arachidonic acid (C20:4n6), PE O (18:1/20:4), PE O (18:2/20:4), and PE O (18:2/18:2), etc. Understanding the relationship between flavour compounds and lipids of pork will be helpful to control the quality of pork.

The flavour of wheat gluten hydrolysate after Corynebacterium Glutamicum fermentation: Effect of degrees of hydrolysis and fermentation time.

Corynebacterium glutamicum was used to ferment wheat gluten hydrolysates (WGHs) to prepare flavour base. This study investigated the effect of hydrolysis degrees (DHs) and fermentation time on flavour of WGHs. During fermentation, the contents of amino nitrogen, total acid and small peptides increased, while the protein and pH value decreased. Succinic acid, GMP, and Glu were the prominent umami substances in fermented WGHs. The aromas of WGHs with different DHs could be distinguished by electronic nose and GC-IMS. Based on OAV of GC-MS, hexanal was the main compound in WGHs, while phenylethyl alcohol and acetoin were dominant after fermentation. WGHs with high DHs accumulated more flavour metabolites. Correlation analysis showed that small peptides (<1 kDa) could promote the formation of flavour substances, and Asp was potentially relevant flavour precursor. This study indicated that fermented WGHs with different DHs can potentially be used in different food applications based on flavour profiles.