Zero-calorie Sweetener Research Articles

Sandy loam soil maintains better physicochemical parameters and more abundant beneficial microbiomes than clay soil in Stevia rebaudiana cultivation.

Depending on the texture of soil, different physicochemical and microbiological parameters are characterized, and these characteristics are influenced by crop cultivation. Stevia, a popular zero-calorie sweetener crop, is widely cultivated around the world on various soil textures. Sandy loam and clay soil show great differences in physicochemical and biological parameters and are often used for Stevia cultivation. To understand the effects of Stevia cultivation on soil physicochemical and biological features, we investigated the changes of physicochemical and microbiological parameters in sandy loam and clay soil following Stevia cultivation. This study was carried out through different physiological and biochemical assays and microbiomic analysis. The results indicated that the sandy loam soil had significantly lower pH and higher nutrient content in the rhizosphere and bulk soils after the Stevia cultivation. The sandy loam soil maintained higher bacterial diversity and richness than the clay soil after Stevia harvest. Beneficial bacteria such as Dongia, SWB02, Chryseolinea, Bryobacter and Devosia were enriched in the sandy loam soil; however, bacteria such as RB41, Haliangium and Ramlibacter, which are unfavorable for nutrient accumulation, predominated in clay soil. Redundancy analysis indicated that the variation in the composition of bacterial community was mainly driven by soil pH, organic matter, total nitrogen, available phosphorus, and microbial biomass phosphorus. This study provides a deeper understanding of physicochemical and microbiological changes in different soil textures after Stevia cultivation and guidance on fertilizer management for Stevia rotational cultivation.

Development of New Multi-Glycosylation Routes to Facilitate the Biosynthesis of Sweetener Mogrosides from Bitter Immature Siraitia Grosvenorii Using Engineered Escherichia coli.

Mogrosides, which have various pharmacological activities, are mainly extracted from Siraitia grosvenorii (Luo Han Guo) and are widely used as natural zero-calorie sweeteners. Unfortunately, the difficult cultivation and long maturation time of Luo Han Guo have contributed to a shortage of mogrosides. To overcome this obstacle, we developed a highly efficient biosynthetic method using engineered Escherichia coli to synthesize sweet mogrosides from bitter mogrosides. Three UDP-glycosyltransferase (UGT) genes with primary/branched glycosylation catalytic activity at the C3/C24 sites of mogrosides were screened and tested. Mutant M3, which could catalyze the glycosylation of nine types of mogrosides, was obtained through enhanced catalytic activity. This improvement in β-(1,6)-glycosidic bond formation was achieved through single nucleotide polymorphisms and direct evolution, guided by 3D structural analysis. A new multienzyme system combining three UGTs and UDP-glucose (UDPG) regeneration was developed to avoid the use of expensive UDPG. Finally, the content of sweet mogrosides in the immature Luo Han Guo extract increased significantly from 57% to 95%. This study not only established a new multienzyme system for the highly efficient production of sweet mogrosides from immature Luo Han Guo but also provided a guideline for the high-value utilization of rich bitter mogrosides from agricultural waste and residues.

Consumer acceptability of no added sugar pound cakes under blind and informed testing conditions.

Stevia is an emerging natural high-intensity sweetener. There are negative perceptions of zero-calorie sweeteners, but studies that provide knowledge of these sweeteners improve their perception. This study evaluated consumer acceptability of a zero-sugar bakery product under blind and informed conditions (n=96) along with physicochemical analysis of the products. Rebaudioside A (Reb A) and the new types of stevia (Rebs D and M) with sugar as a control were used to formulate pound cakes. Panelists evaluated the overall hedonic impressions(aroma, texture, flavor, and aftertaste) and intensity (sweetness and bitterness) of the cakes under blind and informed conditions with an enforced 2-week break between evaluations. During the informed session, a document was provided prior to evaluating samples that included stevia's health benefits and the nutritional facts panels for the cakes. The cakes underwent volatile profile (electronic nose [e-nose]) and water activity (aw ) analysis. Overall, stevia cakes showed an increase in flavor and texture liking during the informed session when compared to the blind session, but only Reb A showed a significant difference (p <0.05). The increase in liking scores indicated that information positively affected the consumer's perception of the stevia-sweetened cakes attributes. The e-nose confirmed differences in aroma. There was a significant difference in aw of the samples Rebs A, D, M versus sucrose (p <0.05). No significant differences were observed among the Rebs (p >0.05). This study illustrates that stevia, despite non-browning or fermenting, can be used in a practical baking application, and product-related information impacts consumer acceptability. PRACTICAL APPLICATION: This study demonstrates that product-related information may have an impact on the consumer acceptability of the product. Through potential labeling improvements, overall consumer perception and acceptability of zero-sugar added or low-sugar products could be improved. This study also illustrates that stevia, despite being a non-browning or fermenting sugar alternative, can be used in a practical baking application.

Recently evolved combination of unique sulfatase and amidase genes enables bacterial degradation of the wastewater micropollutant acesulfame worldwide.

Xenobiotics often challenge the principle of microbial infallibility. One example is acesulfame introduced in the 1980s as zero-calorie sweetener, which was recalcitrant in wastewater treatment plants until the early 2010s. Then, efficient removal has been reported with increasing frequency. By studying acesulfame metabolism in alphaproteobacterial degraders of the genera Bosea and Chelatococcus, we experimentally confirmed the previously postulated route of two subsequent hydrolysis steps via acetoacetamide-N-sulfonate (ANSA) to acetoacetate and sulfamate. Genome comparison of wildtype Bosea sp. 100-5 and an acesulfame degradation-defective mutant revealed the involvement of two plasmid-borne gene clusters. The acesulfame-hydrolyzing sulfatase is strictly manganese-dependent and belongs to the metallo beta-lactamase family. In all degraders analyzed, it is encoded on a highly conserved gene cluster embedded in a composite transposon. The ANSA amidase, on the other hand, is an amidase signature domain enzyme encoded in another gene cluster showing variable length among degrading strains. Transposition of the sulfatase gene cluster between chromosome and plasmid explains how the two catabolic gene clusters recently combined for the degradation of acesulfame. Searching available genomes and metagenomes for the two hydrolases and associated genes indicates that the acesulfame plasmid evolved and spread worldwide in short time. While the sulfatase is unprecedented and unique for acesulfame degraders, the amidase occurs in different genetic environments and likely evolved for the degradation of other substrates. Evolution of the acesulfame degradation pathway might have been supported by the presence of structurally related natural and anthropogenic compounds, such as aminoacyl sulfamate ribonucleotide or sulfonamide antibiotics.

Heterologous mogrosides biosynthesis in cucumber and tomato by genetic manipulation

Mogrosides are widely used as high-value natural zero-calorie sweeteners that exhibit an array of biological activities and allow for vegetable flavour breeding by modern molecular biotechnology. In this study, we developed an In-fusion based gene stacking strategy for transgene stacking and a multi-gene vector harbouring 6 mogrosides biosynthesis genes and transformed it into Cucumis sativus and Lycopersicon esculentum. Here we show that transgenic cucumber can produce mogroside V and siamenoside I at 587 ng/g FW and 113 ng/g FW, respectively, and cultivated transgenic tomato with mogroside III. This study provides a strategy for vegetable flavour improvement, paving the way for heterologous biosynthesis of mogrosides.

Enhancing erythritol production from crude glycerol in a wild-type Yarrowia lipolytica by metabolic engineering.

Background: Erythritol is a zero-calorie sweetener that is widely used in the food, pharmaceutical, and medical industries. Crude glycerol is the main by-product of biodiesel, and the effective utilization of crude glycerol will help to improve biodiesel viability. Previous studies on the production of erythritol from Y. lipolytica using crude glycerol as a carbon source have focused on optimizing the fermentation process of the mutant Y. lipolytica Wratislavia K1, while metabolic engineering has not been successfully applied. Results: To this end, we engineered the yeast Y. lipolytica to increase the productivity of this strain. Wild strains tolerant to high concentrations of crude glycerol were screened and identified. A series of rational metabolic approaches were employed to improve erythritol production. Among them, the engineered strain Y-04, obtained by tandem overexpression of GUT1 and GUT2, significantly increased glycerol assimilation by 33.3%, which was consistent with the results of RT-qPCR analysis. The effects of tandem overexpression of GUT1, GUT2, TKL1, and TAL1 on erythritol synthesis were also evaluated. The best results were obtained using a mutant that overexpressed GUT1, GUT2, and TKL1 and knocked out EYD1. The final Y-11 strain produced 150 g/l erythritol in a 5-L bioreactor with a yield and productivity of 0.62 g/g and 1.25 g/l/h, respectively. To the best of our knowledge, this is the highest erythritol yield and productivity from crude glycerol ever reported in Y. lipolytica. Conclusion: This work demonstrated that overexpression of GUT1, GUT2, and TKL1 and knockdown of EYD1 could be used to improve crude glycerol utilization and erythritol synthesis in Y. lipolytica. The process parameters such as erythritol yield and productivity were significantly elevated, which is valuable for industrial applications. Crude glycerol, as a carbon source, could efficiently restrict the synthesis of by-products while enhancing the generation of erythritol, compared to glucose. This indicates considerable potential for synthesizing value-added products from crude glycerol by Y. lipolytica.

Development and Quality Assessment of Probiotic Ice-cream using Stevia as Sweetener

Probiotics is defined as a live microorganism that, when consumed in adequate quantities, confers a healthbenefit to the host. Stevia is a zero-calorie sweetener does not increase the blood glucose level, making it apossible alternative for diabetics and replacing sugar with different concentrations. Stevia incorporated atfour different levels (0.1%, 0.2%, 0.3% and 0.4%) and sugar reduction levels (6.5%, 13%, 19% and 25%).Based on the organoleptic evaluation highest overall acceptability for the Ice-cream incorporated with 0.2%stevia and sugar replacement (13%) was finalised. The 12 hrs incubated milk with Probiotic bacteria cellcount of 107cfu/ml was added to Ice-cream (T1). After inoculation of purified probiotic into the Ice-creamdoes not affect the sensory properties of probiotic Ice-cream with stevia (T1). Final product (T1) containsviable cell count of 107cfu/ml remained same count thus proving the probiotic bacterial viability. The selectedsample was analysed for Physico-Chemical properties. Overrun percentage was high in T1 sample (35.8%)and T1 (72.5 min) had significantly higher meltdown time than control (55.4 min). The selected samplecontains the moisture of 61.8%, Protein-5.6%, Fat-11.0%, Ash-0.92%, carbohydrate-20.7% and total solids of38.2%. Energy values of selected Ice-cream sample were found to be 217.1 kcal/g for control sample and204.2 kcal/g for T1 sample.

Glycosyltransferase engineering and multi-glycosylation routes development facilitating synthesis of high-intensity sweetener mogrosides

SummaryMogrosides are widely served as natural zero-calorie sweeteners. To date, the biosynthesis of high-intensity sweetness mogrosides V from mogrol has not been achieved because of inefficient and uncontrollable multi-glycosylation process. To address this challenge, we reported three UDP-glycosyltransferases (UGTs) catalyzing the primary and branched glycosylation of mogrosides and increased the catalytic efficiency by 74–400-folds toward branched glycosylation using an activity-based sequence conservative analysis engineering strategy. The computational studies provided insights into the origin of improved catalytic activity. By virtue of UGT mutants, we provided regio- and bond-controllable multi-glycosylation routes, successfully facilitating sequential glycosylation of mogrol to three kinds of mogroside V in excellent yield of 91–99%. Meanwhile, the feasibility of the routes was confirmed in engineered yeasts. It suggested that the multi-glycosylation routes would be combined with mogrol synthetic pathway to de novo produce mogrosides from glucose by aid of metabolic engineering and synthetic biology strategies in the future.

Critical Review on Key Approaches to Enhance Synthesis and Production of Steviol Glycosides: A Blueprint for Zero-Calorie Sweetener

Steviol glycosides (SvGls) are plant secondary metabolites belonging to the class diterpenes. SvGls naturally derived from non-calorie sugar have therapeutic properties for diabetics, carcinogens, and anti-microbials. Over the past few years, SvGls have been extensively researched because of their extraordinary sweetness without side effects. SvGls are classified into several types based on the number of sugar bases attached to the steviol. Due to the difference in glycosylation, different SvGls have different sensory properties. One of the main obstacles is the mass production of SvGls. Novel physical and chemical treatments and advanced biotechnological approaches are introduced to increase the production of SvGls. Extraction of high-purity SvGls from plants is difficult, so researchers have manipulated the metabolic pathways of engineered microorganisms for bulk production of SvGls. There are many aspects related to the biochemistry and metabolism of SvGls, but their functional relationship with the S. rebaudiana is not well understood. Thus there is a need for in-depth research in this area. This review provides the readers with an overview of the research gaps and possible promising methodologies that can be utilized in the future. To trigger more research, this review encompasses the recent developments in SvGls production and marketing.

Fermentative Production of Erythritol from Cane Molasses Using Candida magnoliae: Media Optimization, Purification, and Characterization

Erythritol is a natural, zero-calorie sweetener that can be used as a sugar substitute and humectant for different products such as confectionaries, tablets, etc. Methods such as extraction and chemical synthesis for erythritol synthesis are not feasible or sustainable due to lower yield and higher operating costs. In the present study, erythritol is produced through the submerged fermentation of cane molasses, a by-product of the cane sugar industry, in the presence of the osmophilic yeast Candida magnoliae. Erythrose reductase enzyme assay was used for quantifying erythritol yield. Plackett–Burman’s design screened the three most influential factors viz. molasses, yeast extract, and KH2PO4 out of 12 contributing factors. Further, the concentration of molasses (200–300 g/L), yeast extract (9–12 g/L), and KH2PO4 (2–5 g/L) were optimized using response surface methodology coupled with numerical optimization. The optimized erythritol yield (99.54 g·L−1) was obtained when the media consisted of 273.96 g·L−1 molasses, 10.25 g·L−1 yeast extract, and 3.28 g·L−1 KH2PO4 in the medium. After purification, the liquid chromatography–mass spectrometry (LC-MS) analysis of erythritol crystals from this optimized fermentation condition showed 94% purity. Glycerol was produced as the side product (5.4%) followed by a trace amount of sucrose and mannitol. The molecular masses of the erythritol were determined through mass spectrometry by comparing [M + Na] + ions. Analysis in electrospray (ES) positive mode gave (m/z) of 145.12 [M + 23]. This study has reported a higher erythritol yield from molasses and used osmotolerant yeast Candida magnoliae to assimilate the sucrose from molasses.

Chemical profile and beneficial effect of standardized extract of Stevia rebaudiana Bertoni leaves on metabolic syndrome in high fat diet streptozotocin-induced diabetic rats

Stevia (Stevia rebaudiana Bertoni) is a natural zero calorie sweetener with significant economic and medicinal values due to its high contents of steviosides (SVGs) in the leaves. The aqueous extract of Stevia leaves (TAqE) was standardized to contain 8.5% w/w of SVGs (HPLC), total phenolics (164.63 ± 1.39 µg Gallic acid/mg extract) and total flavonoids of 100.5 ± 0.79 µg QE/mg extract. Twenty-one compounds were tentatively identified in the leaves via UPLC-Orbitrap HRMS and stevioside, rebaudioside A, and quercetrin were isolated from TAqE by repeated column chromatography. Stevioside showed significant inhibition of pancreatic lipase, α-amylase, and α-glucosidase enzymes. The effect of a standardized TAqE on high fat diet (HFD)-streptozotocin (STZ)-induced diabetic rats was investigated. Thirty-six animals were divided into 6 groups (each of 6). Rats in group I (control) and group II (control/HFD-STZ) received distilled water, and rats in groups III and IV received TAqE for 4 weeks in two doses; 300 mg/kg b.wt., and 500 mg/kg b.wt., respectively. Rats in group V received metformin (200 mg/kg), while those in group VI received statin (1 mg/kg). Body weight, fasting blood glucose, lipid profile (total cholesterol and triglycerides), liver enzymes (alanine transaminase and aspartic transaminase), and serum kidney parameters (urea and creatinine) were decreased in rats treated with TAqE (300 mg/kg b.wt.), while insulin sensitivity was enhanced, when compared to that in group II. These findings could justify the use of Stevia as a complementary medicine for the prevention and treatment of metabolic changes associated with diabetes mellitus type 2.

The chromosome-level Stevia genome provides insights into steviol glycoside biosynthesis

Stevia (Stevia rebaudiana Bertoni) is well known for its very sweet steviol glycosides (SGs) consisting of a common tetracyclic diterpenoid steviol backbone and a variable glycone. Steviol glycosides are 150–300 times sweeter than sucrose and are used as natural zero-calorie sweeteners. However, the most promising compounds are biosynthesized in small amounts. Based on Illumina, PacBio, and Hi-C sequencing, we constructed a chromosome-level assembly of Stevia covering 1416 Mb with a contig N50 value of 616.85 kb and a scaffold N50 value of 106.55 Mb. More than four-fifths of the Stevia genome consisted of repetitive elements. We annotated 44,143 high-confidence protein-coding genes in the high-quality genome. Genome evolution analysis suggested that Stevia and sunflower diverged ~29.4 million years ago (Mya), shortly after the whole-genome duplication (WGD) event (WGD-2, ~32.1 Mya) that occurred in their common ancestor. Comparative genomic analysis revealed that the expanded genes in Stevia were mainly enriched for biosynthesis of specialized metabolites, especially biosynthesis of terpenoid backbones, and for further oxidation and glycosylation of these compounds. We further identified all candidate genes involved in SG biosynthesis. Collectively, our current findings on the Stevia reference genome will be very helpful for dissecting the evolutionary history of Stevia and for discovering novel genes contributing to SG biosynthesis and other important agronomic traits in future breeding programs.

Tomatoes expressing thaumatin II retain their sweet taste after salting and pickling processing.

Thaumatin II, a supersweet protein from the African plant katemfe (Thaumatococcus daniellii Benth.), shows promise as a zero-calorie sweetener for use in the food and pharmaceutical industries and for improving the taste of fruit. We report on the stability of thaumatin in salted and pickled tomatoes, as well as on the effect of thaumatin on the taste quality of processed tomatoes. Fruit of tomato cv. Yalf, transformed with the thaumatin II gene were salted and pickled and then stored for 6months. Western blot analysis showed relative thaumatin II stability at salting; its content in processed fruits was 62-83% of the initial level depending in the studied line. In pickled tomatoes, thaumatin II content was decreased by up to 25% of the initial amount. Both salted and pickled tomatoes had a sweet taste with a typical thaumatin aftertaste. Salted tomatoes were characterized as being sweeter than pickled tomatoes. The overall taste of pickled tomatoes was rated by panellists as significantly better compared to that of salted or non-processed ones. In the present study, we have shown that tomatoes expressing supersweet protein thaumatin II can be used for processing under mild conditions, including salting and pickling. © 2021 Society of Chemical Industry.

Components of the Gut Microbiome That Influence Bone Tissue-Level Strength.

Modifications to the constituents of the gut microbiome influence bone density and tissue-level strength, but the specific microbial components that influence tissue-level strength in bone are not known. Here, we selectively modify constituents of the gut microbiota using narrow-spectrum antibiotics to identify components of the microbiome associated with changes in bone mechanical and material properties. Male C57BL/6J mice (4 weeks) were divided into seven groups (n=7-10/group) and had taxa within the gut microbiome removed through dosing with: (i) ampicillin; (ii) neomycin; (iii) vancomycin; (iv) metronidazole; (v) a cocktail of all four antibiotics together (with zero-calorie sweetener to ensure intake); (vi) zero-calorie sweetener only; or (vii) no additive (untreated) for 12 weeks. Individual antibiotics remove only some taxa from the gut, while the cocktail of all four removes almost all microbes. After accounting for differences in geometry, whole bone strength was reduced in animals with gut microbiome modified by neomycin (-28%, p=0.002) and was increased in the group in which the gut microbiome was altered by sweetener alone (+39%, p < 0.001). Analysis of the fecal microbiota detected seven lower-ranked taxa differentially abundant in animals with impaired tissue-level strength and 14 differentially abundant taxa associated with increased tissue-level strength. Histological and serum markers of bone turnover and trabecular bone volume per tissue volume (BV/TV) did not differ among groups. These findings demonstrate that modifications to the taxonomic components of the gut microbiome have the potential to decrease or increase tissue-level strength of bone independent of bone quantity and without noticeable changes in bone turnover. © 2021 American Society for Bone and Mineral Research (ASBMR).

Effect of the bio-fertilizers on the steviol glycosides (SGs) content and biomass in Stevia rebaudiana (Bert.) Bertoni at vegetative and flowering stages

Stevia rebaudiana Bertoni is a unique medicinal herb mostly utilized for substitution of the sugar, producing zero-calorie sweeteners known as steviol glycosides (SGs), including stevioside (ST), rebaudioside A (RA), rebaudioside F (RF), rebaudioside C (RC) and dulcoside (DU). While bio-fertilizers have been demonstrated to enhance the production of secondary metabolites in many plant species, their effects on S. rebaudiana have not been thoroughly studied. This paper presents a study on the effects of bio-fertilizers and their interactions on modifying and improving the SGs content and biomass production in S. rebaudiana at two growth stages, vegetative and flowering. Vermicompost (V) and cow manure (M) (as soil amendments), and garden soil (S) were used separately, and also in combination to fill pots. In this research two factors included cultivation bed [S (control), SM (garden soil + 15 % cow manure + 0% vermicompost), SMV10 (garden soil + 15 % cow manure + 10 % vermicompost), and SMV20 (garden soil + 15 % cow manure + 20 % vermicompost) by %weight in pots (w/w)], and fungi application (two species of fungi Funneliformis mosseae (Fm), Serendipita indica (Si) inoculation and without fungi inoculation) were investigated on the percent, yield of SGs and biomass at vegetative and flowering stages. The percentage and yield of total SGs, SGs elements [(ST, RA, RF, RC, DU, and RA/SGs ratio] and aerial dry biomass of S. rebaudiana were significantly affected by all treatments and their interactions (P < 0.01). Generally, the highest SGs elements percentage was obtained in the control plant at the vegetative and flowering stages (S). Also, the highest SGs yield was observed in SMV20Fm in both vegetative and flowering stages. Funneliformis mosseae, in combination with SMV20, had shown significant effects on RA/SGs ratio, SGs yield, and plant aerial dry biomass in vegetative and flowering stages.

Rapid and Economic Determination of 13 Steviol Glycosides in Market-Available Food, Dietary Supplements, and Ingredients: Single-Laboratory Validation of an HPLC Method.

Steviol glycosides, obtained from leaves of Stevia rebaudiana Bertoni (stevia) or produced via bioconversion and biosynthesis, are diterpenes used by the food/dietary supplement industry as zero-calorie sweeteners derived from natural sources. JECFA 2017 is the most updated international standardized method but it runs for 80 min per sample with suboptimal separations on several critical pairs for its high-performance liquid chromatography-ultraviolet (HPLC-UV) determination. We developed and validated a rapid and economic HPLC-UV method using the superficially porous particle column to determine 13 steviol glycosides (stevioside, dulcoside A, rubusoside, steviobioside, and rebaudioside A-F, I, M, and N). Baseline separation with a minimum resolution of 1.5 for 13 steviol glycosides was achieved within only 14 min of separation time. The hydrocarbon stationary phase with additional steric interactions from the isobutyl side chains on the C18 ligand was shown to be an important contributor to chromatographic selectivity of several critical pairs of steviol glycosides. The method was proven to perform suitably on columns from three different manufacturers and two HPLC instruments. The method was further used to perform a single-lab validation on eight food and supplement products with multiple matrices. The results ranged from 0.05% w/w rebaudioside A for a hard-candy finished product to 100.8% w/w purity for a rebaudioside M raw ingredient. The validation test results showed that the method was linear, suitable, specific, accurate, and precise. The method is therefore suitable to be considered as a new industrial standard for quality control analysis for stevia products.

Contribution of Katsura-uri (Japan's Heirloom Pickling Melon, Cucumis melo var. conomon) at the Completely Ripe Stage to Diabetes Control.

The fruit of Katsura-uri (Japan's heirloom pickling melon, Cucumis melo var. conomon) possesses a fruity aroma and moderate sweetness. The fruit juice has potential to minimize human postprandial blood glucose levels. This study provides information regarding the health benefits of Katsura-uri and its utility in treating diabetes. The study methodology involved measuring the color and firmness of Katsura-uri fruit at five ripening stages, and quantitation of the aroma substances, proximate composition, and sugars. Significant changes were detected in the color, firmness, and level of aroma substances with ripening of Katsura-uri fruit, albeit with no major changes in proximate composition, with the exception of dietary fiber, and sugars. To determine the effects of Katsura-uri juice, the blood glucose levels of ten diabetic volunteers aged 46-75 y were monitored after its consumption, and compared with after consumption of muskmelon juice equivalent to the total weight of Katsura-uri juice. The blood glucose area under the curve level was significantly lower after consumption of Katsura-uri juice (16±5 h • mg/dL) than after consumption of muskmelon juice (55±17 h • mg/dL; p<0.05). The level of the glucose spike was also significantly lower after consumption of Katsura-uri juice (22±5 mg/dL) than after consumption of muskmelon juice (64±6 mg/dL; p<0.05). The completely ripe Katsura-uri fruit provides the best results for diabetic subjects, which is the first case of fruits sweetened with the addition of zero-calorie sweeteners.

Obtención de ciruelas osmodeshidratadas bajas en calorías mediante un proceso dual de sustitución de azúcares

&#x0D; &#x0D; &#x0D; &#x0D; Se llevó a cabo el desarrollo de un alimento saludable reducido en calorías y de calidad diferencial: ciruelas D’Agen deshidratadas. Esto mediante un proceso osmótico dual (D3S) de sustitución de los azúcares naturales de la fruta por edulcorante no calórico Stevia complementado por secado. Se utilizó un diseño experimental factorial bajo la consideración del tiempo de pretratamiento en Etapa i (10, 20, 30 minutos), frecuencia de ultrasonido en Etapas i y ii (0,40 kHz), tiempo de tratamiento en Etapa ii (10, 20, 30 minutos), y concentración de solución de edulcorante Stevia (20, 25% p/p). El monitoreo del proceso se realizó al evaluar la evolución del contenido de sólidos solubles (SS), humedad (M) y peso de las muestras (W), así como las variables pérdida de sólidos solubles (SG), pérdida de agua (WL), y reducción de peso (WR), en cada etapa. Los resultados se analizaron mediante Anova (análisis de la varianza) y RSM (metodología de superficie de respuesta). El proceso dual óptimo que removió la mayor cantidad de azúcares calóricos e incorporó más edulcorante natural Stevia cero calorías (10,67%) a las ciruelas consistió en un pretratamiento de 22,8 minutos asistido por ultrasonido, seguido de inmersión en solución de Stevia del 20% p/p durante 22 minutos con aplicación de ultrasonido, mientras que para la optimización múltiple de SG y WL las condiciones operativas fueron solución al osmótica 23% p/p durante 24,6 minutos.&#x0D; &#x0D; &#x0D; &#x0D;

Knock Down of Erythrulose Kinase (eyk1) Leads to the Enhancement of Erythritol Production in Yarrowia lipolytica Mutant Strain YE4-2

Erythritol is a nearly zero-calorie sweetener, which is in great demand in food and health care industry. Our previous work obtained a Yarrowia lipolytica mutant strain YE4-2 with high erythritol yield. However, its conversion rate of glucose needs further improvement. Erythrulose kinase (eyk1) participates in the early stages of erythritol catabolism; the existence of eyk1 prevents the accumulation of erythritol. In this study, the genetic transformation system of Y. lipolytica YE4-2 was firstly established and the Cre/loxp homologous recombination knockout system was constructed. By using the Cre/loxp system, eyk1 gene was deleted. According to the PCR verification of eyk1 gene, it was preliminarily determined that one allele of eyk1 in this recombinant strain was knocked out. The recombination strain showed an average conversion rate of 48.51%, which was 49.26% higher than that of YE4-2 and the erythritol productivity is increased by 62.26%, which would be a significant improvement in the industrial scale.

Effects of Septoglomus viscosum inoculation on biomass yield and steviol glycoside concentration of some Stevia rebaudiana chemotypes

There is growing interest in the medicinal properties of Stevia rebaudiana Bertoni, due to its ability to produce zero-calorie sweeteners, steviol glycosides (SGs). Inoculation of the roots with arbuscular mycorrhizal fungi (AMF) is a method used to improve the production of secondary metabolites by many plant species, but its effects on S. rebaudiana have not been extensively studied, especially in open field conditions in Italy. Therefore, a field experiment was conducted to study the effects of the fungus Septoglomus viscosum on the leaves and secondary metabolites of three micropropagated stevia chemotypes (L1, L2 and L3) over three years. Our results indicate that the establishment of symbiotic relations between AMF and stevia plants of selected chemotypes improved leaf yield, because the mycorrhizal (MIC) treated plants had a better tendency to regrowth, especially L1 and L3, which had survival rates of 80 % and 90 %, respectively. MIC treatment also had positive effects in terms of SG production, which was not significant in terms of concentration, but was significant in terms of production.