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After publication of the draft Generally Regarded As Safe (GRAS) rule in 1997, the United States (US) Food and Drug Administration (FDA) initiated an Interim Pilot Program encouraging the notification to FDA of GRAS determinations. This paper analyzes GRAS notifications submitted during the Interim Pilot Program along with warning letters issued during the same time period to better understand the evolution of the program and anticipate the future GRAS landscape. The success of the GRAS Notification program is demonstrated by the increasing rate of GRAS Notifications submitted to the FDA during the Interim Pilot Program, as well as the shift from a primarily domestic process to a process featuring an equal to greater contribution of GRAS Notifications from companies outside the US. Analysis of the first 600 GRAS Notifications revealed a number of interesting trends regarding the inclusion and composition of GRAS Expert Panels; differences in notifications for substances with nutritive, processing aid, or effect; and the duration of GRAS Notifications. The review of FDA warning letters associated with GRAS issues provides additional insight into GRAS notices, from the perspective of ongoing post-market emphasis on food safety with the implementation of the GRAS Final Rule. [ABSTRACT FROM AUTHOR]

Salt forms of pharmaceutical compounds can have unique pharmacokinetic and toxicity properties. MDV1634 was evaluated for neurology indication and also demonstrated blood pressure (BP)-lowering effects in nonclinical studies. During the chemistry manufacturing campaign, 2 salt forms, dihydrochloride (2HCl) and maleate (MAL), which improved chemical stability and water solubility of the free base were identified. MDV1634.MAL showed better chemical attributes and was evaluated in toxicology studies for further development. A 28-day oral toxicity study in dogs with MDV1634.MAL demonstrated partially reversible renal toxicity. Although MAL salt is generally regarded as safe, renal toxicity is sometimes observed in rats and dogs. To evaluate contribution of each salt form to renal toxicity and BP lowering, an additional 28-day study was conducted with MDV1634.2HCL and MDV1634.MAL, which included toxicokinetics, continuous BP measurement in a subset of dogs, and sensitive urinary biomarker evaluation for temporal monitorability and reversibility of potential renal findings. In the repeat study, both salt forms showed similar exposures during the dosing period, but renal tubular toxicity was observed only with MDV1634.MAL and not with MDV1634.2HCl. The renal findings with MDV1634.MAL included early urinary biomarker changes (increase in albumin, clusterin, β2 microglobulin, and neutrophil gelatinase-associated lipocalin); elevations in serum blood urea nitrogen and creatinine; and microscopic findings of partially reversible tubular basophilia, single cell necrosis, pigmentation, and mineralization. The renal findings in contrast to the BP findings were MAL-specific and considered not related to MDV1634, thereby under scoring the importance of salt forms in pharmaceutical development. [ABSTRACT FROM AUTHOR]

GRAS genes encode plant-specific transcription factors that play important roles in plant growth and development. However, little is known about the GRAS gene family in apple. In this study, 127 GRAS genes were identified in the apple (Malus domestica Borkh.) genome and named MdGRAS1 to MdGRAS127 according to their chromosomal locations. The chemical characteristics, gene structures and evolutionary relationships of the MdGRAS genes were investigated. The 127 MdGRAS genes could be grouped into eight subfamilies based on their structural features and phylogenetic relationships. Further analysis of gene structures, segmental and tandem duplication, gene phylogeny and tissue-specific expression with ArrayExpress database indicated their diversification in quantity, structure and function. We further examined the expression pattern of MdGRAS genes during apple flower induction with transcriptome sequencing. Eight higher MdGRAS (MdGRAS6, 26, 28, 44, 53, 64, 107, and 122) genes were surfaced. Further quantitative reverse transcription PCR indicated that the candidate eight genes showed distinct expression patterns among different tissues (leaves, stems, flowers, buds, and fruits). The transcription levels of eight genes were also investigated with various flowering related treatments (GA3, 6-BA, and sucrose) and different flowering varieties (Yanfu No. 6 and Nagafu No. 2). They all were affected by flowering-related circumstance and showed different expression level. Changes in response to these hormone or sugar related treatments indicated their potential involvement during apple flower induction. Taken together, our results provide rich resources for studying GRAS genes and their potential clues in genetic improvement of apple flowering, which enriches biological theories of GRAS genes in apple and their involvement in flower induction of fruit trees. [ABSTRACT FROM AUTHOR]

The taxonomic differentiation among lactic acid bacteria (LAB) using only phenotypic methods is inadequate. In the past, several phenotypic techniques have been applied in identification of different species of LAB from Nigerian indigenous fermented foods. In this study, two less-expensive and rapid molecular techniques were employed in the identification and differentiation of LAB species isolated from some Nigerian fermented foods: gari (cassava mash), fufu (cassava dough) and ogi (maize gruel) to avoid the common errors that are synonymous with the use of conventional methods. The size of the amplified products for the LAB species using universal primers fD1 and rD1 was about 1500 bp. The differentiation of the three predominant LAB, Lactobacillus plantarum , Pediococcus pentosaceus and Weissella confusa to species level was performed by intergenic transcribed spacer (ITS-PCR) and amplified ribosomal DNA restriction analysis (ARDRA) using three restriction endonucleases: Hinf I, Hae III and Rsa I. In summary, the study demonstrated that the ARDRA and ITS-PCR analysis are simple, fast and efficient methods that can be used for the routine identification and rapid differentiation of LAB species which are often regularly used as probiotics, starter culture and biopreservative in fermented foods consumed since they are generally regarded as safe (GRAS). [ABSTRACT FROM AUTHOR]

Postharvest losses on table grapes caused by Botrytis cinerea , are controlled with SO 2 fumigations carried out every 7 or 10 d. The use of this gas is becoming more difficult to justify because of undesirable effects on the fruit and the increasing concern for human health. Objectives of the paper were to evaluate if repeated treatments with acetic acid (AC) during storage, were effective in preserving table grapes quality, comparing in addition the effects of AC and SO 2 treatments. Experiments carried out in vitro on B. cinerea proved that the effect of AC on mycelia growth and conidia germination was related not only to the dose and exposure period, but also to the elapsed time between fungal inoculation and treatment. The reinoculum test demonstrated that a treatment with 20 μL L −1 of AC for 15 min had a fungicidal effect. A laboratory test was performed, to evaluate in vivo the effectiveness of AC on B. cinerea . Results suggested that higher doses were needed to control the pathogen. On naturally infected table grapes two storage experiments were carried out: in the first trial a single AC concentration of 50 μL L −1 was used to perform one or two fumigations after 4 or 8 weeks (w), while three different AC concentrations (30, 50 and 75 μL L −1 ) were used in the second trial and fumigations were repeated 5, 3 and 2 times respectively. Treatments lasted 15 min and fruit was stored for 8 w at 5 °C and 90% RH, followed by 3 d of a simulated marketing period. All treatments reduced gray mold incidence, with respect to untreated fruit, after 8 w of storage, but repeated treatments resulted the most effective. Two fumigations at 50 μL L −1 or 5 fumigations at 30 μL L −1 reduced gray mold incidence by 63.6 or 57.1% respectively. Fruit weight loss was significantly reduced by all treatments, while quality parameters resulted not to be affected by any of the treatments. [ABSTRACT FROM AUTHOR]

Conventional fungicides are used in IPM programs to manage fungal plant pathogens, but there are concerns about resistance development in target organisms, environmental contamination, and human health risks. This study explored the potential of calcium propionate (CaP), a common food preservative generally recognized as safe (GRAS) to control fungicide-resistant plant pathogens, mainly Botrytis cinerea , and botrytis blight in ornamentals. In-vitro experiments using mycelium growth inhibition indicated a mean EC 50 value for CaP (pH 6.0) of 527 mg/L for six isolates of Botrytis cinerea as well as 618, 1354, and 1310 mg/L for six isolates each of Monilinia fructicola, Alternaria alternata , and Colletotrichum acutatum. In vitro efficacy tests indicated CaP equally inhibited mycelium growth of fungal isolates sensitive and resistant to FRAC codes 1, 2, 3, 7, 9, 11, 12, and 17 fungicides. CaP at 0.1% (pH 6.0–6.5) reduced infection cushion (IC) formation in vitro, botrytis blight on petunia flowers, and botrytis blight of cut flower roses with little to no visible phytotoxicity. Although higher concentrations strongly inhibited infection cushion formation, they did not improve efficacy and exhibited phytotoxicity. We hypothesize that high concentrations may create tissue damage that facilitates direct fungal penetration without the need for infection cushion and subsequent appressoria formation. This study indicates the potential usefulness of CaP for blossom blight disease management in ornamentals if applied at concentrations low enough to avoid phytotoxicity. [Display omitted] • CaP inhibited B. cinerea isolates sensitive and resistant to FRAC codes 1, 2, 3, 7, 11, 12, and 17 • CaP reduced botrytis blight of petunias and cut flower roses at 0.1% (pH 6.0–6.5) with little to no phytotoxicity • At 0.1% CaP inhibited B. cinerea infection cushion formation by about 50% [ABSTRACT FROM AUTHOR]

2′-Fucosyllactose (2′-FL) is a major oligosaccharide found in human breast milk. It is produced from GDP- L -fucose and D -lactose by ⍺1,2-fucosyltransferase (⍺1,2-fucT), but the enzyme has been identified mostly in pathogens. In this study, an ⍺1,2-fucT was isolated from a Generally Recognized as Safe (GRAS) Bacillus megaterium strain. The enzyme was successfully expressed in metabolically-engineered Escherichia coli. Furthermore, replacement of non-conserved amino acid residues with conserved ones in the protein led to an increase in the rate of 2′-FL production. As a result, fed-batch fermentation of E. coli produced 30 g/L of 2′-FL from glucose and lactose. Thus, the overproduction of 2′-FL using a novel enzyme from a GRAS bacteria strain was successfully demonstrated. • To produce 2′-FL, ⍺1,2-fucosyltransferase was identified from Bacillus megaterium. • The enzyme from a GRAS strain was active in 2′-FL production in engineered E. coli. • The engineering of its non-conserved residues improved solubility and activity. • The 2′-FL production using the enzyme was comparable to the one from a pathogen. [ABSTRACT FROM AUTHOR]

The Federal Food, Drug, and Cosmetic Act (FFDCA) provides that a food is misbranded if the label accompanying the product is false or misleading in any particular. Congress provided minimal guidance to assist the FDA in making these determinations. When challenged, courts have granted substantial deference to FDA’s various interpretations of what constitute a material fact. However, when confronted with the regulatory question of whether or how to label products derived from genetically engineered (GE) crops, the FDA adopted a narrow reading of the statute that focused on changes in the product itself, rather than the act of genetic engineering. Only those GE products that possessed characteristics significantly different from their conventional counterparts would require labels. This “process versus product” distinction in food labels lies at the heart of the FDA’s resistance to repeated calls for mandatory labeling of foods derived from genetic engineering. Consumer interest in GE food, according to the agency, is not a material fact to trigger mandatory labeling under the statute. In contrast to its approach to GE labels, the agency has long required (since 1966) process-based labels for foods treated with irradiation. As recently as 1986, the FDA affirmed that materiality of information under it misbranding analysis is not limited to product safety or even the abstract worth of the information, but whether consumers view the information as important and whether the omission of a labeling statement would mislead the consumer. Accordingly, mere consumer interest can give rise to a mandatory labeling regime under the FFDCA. In the irradiation context, whole foods and single-ingredient products treated with irradiation must bear a label indicating the process. The irradiation of components in a multi-ingredient food product, however, need not bear a label. This distinction between processed, multi-ingredient and whole or single-ingredient foods provides a potential pathway for the agency to revise its approach to mandatory GE labeling. Exempting highly processed, multi-ingredient foods from a labeling regime would minimize traceability and segregation-generated disruptions in the commodity supply chain, thereby minimizing potential compliance costs, while also empowering consumers to express their preferences for non-GE whole and single-ingredient food products.

The GRAS gene family is one of the most important plant-specific gene families, which encodes transcriptional regulators and plays an essential role in plant development and physiological processes. The GRAS gene family has been well characterized in many higher plants such as Arabidopsis, rice, Chinese cabbage, tomato and tobacco. In this study, we identified 38 GRAS genes in sacred lotus (Nelumbo nucifera), analyzed their physical and chemical characteristics and performed phylogenetic analysis using the GRAS genes from eight representative plant species to show the evolution of GRAS genes in Planta. In addition, the gene structures and motifs of the sacred lotus GRAS proteins were characterized in detail. Comparative analysis identified 42 orthologous and 9 co-orthologous gene pairs between sacred lotus and Arabidopsis, and 35 orthologous and 22 co-orthologous gene pairs between sacred lotus and rice. Based on publically available RNA-seq data generated from leaf, petiole, rhizome and root, we found that most of the sacred lotus GRAS genes exhibited a tissue-specific expression pattern. Eight of the ten PAT1-clade GRAS genes, particularly NnuGRAS-05, NnuGRAS-10 and NnuGRAS-25, were preferentially expressed in rhizome and root. In summary, this is the first in silico analysis of the GRAS gene family in sacred lotus, which will provide valuable information for further molecular and biological analyses of this important gene family.

The safety of long-term, high-dose amino acid consumption marketed as dietary supplements or functional or medical foods requires regulatory clearance in the European Union through the novel food process or through the Dietary Supplement Health and Education Act or the GRAS (Generally Recognized As Safe) route in the United States. The safety assessment of high daily doses of amino acids for bodybuilding or other health benefits is expected to require human studies to support tolerability and safety. The need for human studies is based on the fact that there is little or no evidence of toxicity from the conduct of animal toxicity studies and because standard animal testing would be inappropriate because of the large dosages required to provide a suitable margin of safety when extrapolating from animals to humans. Furthermore, the large dosages in animals required to provide a substantial margin of safety could lead to nutritional and physiologic imbalances, potentially confounding an amino acid safety assessment. [ABSTRACT FROM AUTHOR]

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Inclusion bodies (IBs) are protein-based nanoparticles formed in Escherichia coli through stereospecific aggregation processes during the overexpression of recombinant proteins. In the last years, it has been shown that IBs can be used as nanostructured biomaterials to stimulate mammalian cell attachment, proliferation, and differentiation. In addition, these nanoparticles have also been explored as natural delivery systems for protein replacement therapies. Although the production of these protein-based nanomaterials in E. coli is economically viable, important safety concerns related to the presence of endotoxins in the products derived from this microorganism need to be addressed. Lactic acid bacteria (LAB) are a group of food-grade microorganisms that have been classified as safe by biologically regulatory agencies. In this context, we have demonstrated herein, for the first time, the production of fully functional, IB-like protein nanoparticles in LAB. These nanoparticles have been fully characterized using a wide range of techniques, including field emission scanning electron microscopy (FESEM), transmission electron microscopy (TEM), dynamic light scattering (DLS), Fourier transform infrared (FTIR) spectroscopy, zymography, cytometry, confocal microscopy, and wettability and cell coverage measurements. Our results allow us to conclude that these materials share the main physico-chemical characteristics with IBs from E. coli and moreover are devoid of any harmful endotoxin contaminant. These findings reveal a new platform for the production of protein-based safe products with high pharmaceutical interest. Statement of Significance The development of both natural and synthetic biomaterials for biomedical applications is a field in constant development. In this context, E. coli is a bacteria that has been widely studied for its ability to naturally produce functional biomaterials with broad biomedical uses. Despite being effective, products derived from this species contain membrane residues able to trigger a non-desired immunogenic responses. Accordingly, exploring alternative bacteria able to synthesize such biomaterials in a safe molecular environment is becoming a challenge. Thus, the present study describes a new type of functional protein-based nanomaterial free of toxic contaminants with a wide range of applications in both human and veterinary medicine. [ABSTRACT FROM AUTHOR]

Neutral Ion Mass spectrometer (NIM) is one of the instruments in the Particle Environmental Package (PEP) designed for the JUICE mission of ESA to the Jupiter system. NIM, equipped with a sensitive MCP ion detector, will conduct detailed measurements of the chemical composition of Jovian icy moons exospheres. To achieve high sensitivity of the instrument, radiation effects due to the high radiation background (high-energy electrons and protons) around Jupiter have to be minimised. We investigate the performance of an Al–Ta–Al composite stack as a potential shielding against high-energy electrons. Experiments were performed at the PiM1 beam line of the High Intensity Proton Accelerator Facilities located at the Paul Scherrer Institute, Villigen, Switzerland. The facility delivers a particle beam containing e − , μ − and π − with momentum from 17.5 to 345 MeV/c (Hajdas et al., 2014). The measurements of the radiation environment generated during the interaction of primary particles with the Al–Ta–Al material were conducted with dedicated beam diagnostic methods and with the NIM MCP detector. In parallel, modelling studies using GEANT4 and GRAS suites were performed to identify products of the interaction and predict ultimate fluxes and particle rates at the MCP detector. Combination of experiment and modelling studies yields detailed characterisation of the radiation fields produced by the interaction of the incident e − with the shielding material in the range of the beam momentum from 17.5 to 345 MeV/c. We derived the effective MCP detection efficiency to primary and secondary radiation and effective shielding transmission coefficients to incident high-energy electron beam in the range of applied beam momenta. This study shows that the applied shielding attenuates efficiently high-energy electrons. Nevertheless, owing to nearly linear increase of the bremsstrahlung production rate with incident beam energy, above 130 MeV their detection rates measured by the MCP detector compares to the MCP rate of the incident electron beam. We define key performance parameters for the shielding and show direction of its improvements by introducing additional layer of material to attenuate γ-rays and reduce the MCP sensitivity to the penetrating radiation. The experiments also verify the predictions by modelling tools used currently for optimisation of shielding against high-energy particles. [ABSTRACT FROM AUTHOR]

In multicellular organisms, controlling the timing and extent of asymmetric cell divisions (ACDs) is crucial for correct patterning. During post-embryonic root development in Arabidopsis thaliana , ground tissue (GT) maturation involves an additional ACD of the endodermis, which generates two different tissues: the endodermis (inner) and the middle cortex (outer). It has been reported that the abscisic acid (ABA) and gibberellin (GA) pathways are involved in middle cortex (MC) formation. However, the molecular mechanisms underlying the interaction between ABA and GA during GT maturation remain largely unknown. Through transcriptome analyses, we identified a previously uncharacterized C 2 H 2 -type zinc finger gene, whose expression is regulated by GA and ABA, thus named GAZ ( GA- AND ABA-RESPONSIVE ZINC FINGER ). Seedlings ectopically overexpressing GAZ ( GAZ-OX ) were sensitive to ABA and GA during MC formation, whereas GAZ-SRDX and RNAi seedlings displayed opposite phenotypes. In addition, our results indicated that GAZ was involved in the transcriptional regulation of ABA and GA homeostasis. In agreement with previous studies that ABA and GA coordinate to control the timing of MC formation, we also confirmed the unique interplay between ABA and GA and identified factors and regulatory networks bridging the two hormone pathways during GT maturation of the Arabidopsis root. [ABSTRACT FROM AUTHOR]

Current guidelines for cow's milk consumption in children older than age 2 years suggest 1% or 2% milk to reduce the risk of obesity. Given that milk is the main dietary source of vitamin D for North American children and that vitamin D is fat soluble, we hypothesized 25-hydroxyvitamin D (25(OH)D) concentration to be positively associated with the fat content of milk. The objective was to determine the relationship between the fat content of milk consumed and the serum 25(OH)D concentration; our secondary objective was to explore the role that the volume of milk consumed played in this relationship. We completed a cross-sectional study of children aged 12-72 months in the TARGetKids! research network. Multivariable linear regression was used to test the association between milk fat content and child 25(OH)D, adjusted for clinically relevant covariates. The interaction between volume of milk and fat content was examined. Two thousand eight hundred fifty-seven children were included in the analysis. The fat content of milk was positively associated with 25(OH)D (p = 0.03), and the interaction between the volume of milk consumed and the milk fat content was statistically significant (p = 0.005). Childrenwhodrank 1% milk needed 2.46 cups (95% confidence interval (CI) 2.38-2.54) of milk to have a 25(OH)D concentration similar to that of childrenwhodrank 1 cup of homogenized milk (3.25% fat). Childrenwhoconsumed 1% milk had 2.05 (95% CI 1.73-2.42) times higher odds of having a 25(OH)D concentration <50 nmol/L compared with children who consumed homogenized milk. In conclusion, recommendations for children to drink lower-fat milk (1% or 2%) may compromise serum 25(OH)D levels and may require study to ensure optimal childhood health. [ABSTRACT FROM AUTHOR]

[Display omitted] • The melanin nanoparticles in cuttlefish ink have good Photothermal performance. • An edible antibacterial food packaging film with high safety was prepared. • The melanin nanoparticles improve the mechanical properties of pectin film. • The melanin nanoparticles also improved the antibacterial ability of the PC/NMPs films. Although nanocomposite films had shown excellent potential in antibacterial food packaging, their potential harmful impact limits their further application in reality. Therefore, exploring a Generally Recognized As Safe (GRAS) nanomaterial that has antibacterial ability is the pioneer for the fabrication of a real edible nanocomposite-based antibacterial packaging film. Herein, for the first time by using the natural nanostructure extracted from cuttlefish ink, an edible antibacterial food packaging with high safety were constructed. The natural melanin nanoparticles (NMPs) in cuttlefish ink have good photothermal conversion ability. As such, by incorporating with natural pectin (PC) film and with near infrared (NIR) irradiation triggering, the results show that PC/NMPs films perform high-efficiency and short-term bactericidal activity against foodborne pathogenic bacteria, including thermotolerant Listeria monocytogenes. The sterilization rate could reach more than 90 % within only 5 min. Also, this nanocomposite film showed better mechanical properties, thermal stability and barrier properties than the neat pectin film. Antibacterial test on food sample also proved the good antibacterial ability of the PC/NMPs films. Therefore, exploring GRAS natural functional nanocomposite film is expected to be the effective way to promote edible nano-antibacterial packaging. [ABSTRACT FROM AUTHOR]

Background: GRAS transcription factors play a variety of functions in plant growth and development and are named after the first three transcription factors GAI (GIBBERRELLICACIDINSENSITIVE), RGA (REPRESSOROFGAI), and SCR (SCARECROW) found in this family. Oat ( Avena sativa ) is one of the most important forage grasses in the world. However, there are few reports on the GRAS gene family in oat., Methods: In order to understand the information and expression pattern of oat GRAS family members, we identified the GRAS members and analyzed their phylogenetic relationship, gene structure, and expression pattern in oat by bioinformatics technology., Results: The results showed that the oat GRAS family consists of 30 members, and most of the AsGRAS proteins were neutral or acidic proteins. The phylogenetic tree divided the oat GRAS members into four subfamilies, and each subfamily has different conservative domains and functions. Chromosome location analysis suggested that 30 GRAS genes were unevenly distributed on five chromosomes of oat. The results of real-time quantitative reverse transcription-PCR (qRT-PCR) showed that some AsGRAS genes ( AsGRAS12 , AsGRAS14 , AsGRAS21 , and AsGRAS24 ) were all up-regulated with increasing stress treatment time.The results of this study provide a theoretical basis for further research into the corresponding stress of oat. Therefore, further studies concentrating on these AsGRAS genes might reveal the many roles played by GRAS genes in oat., Competing Interests: The authors declare that they have no competing interests., (© 2023 Pan et al.)

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a program for the re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavor ingredients. This publication, eleventh in the series, evaluates the safety of NFCs characterized by primary alcohol, aldehyde, carboxylic acid, ester and lactone constituents derived from terpenoid biosynthetic pathways and/or lipid metabolism. The scientific-based evaluation procedure published in 2005 and updated in 2018 that relies on a complete constituent characterization of the NFC and organization of the constituents into congeneric groups. The safety of the NFCs is evaluated using the threshold of toxicological concern (TTC) concept in addition to data on estimated intake, metabolism and toxicology of members of the congeneric groups and for the NFC under evaluation. The scope of the safety evaluation does not include added use in dietary supplements or any products other than food. Twenty-three NFCs, derived from the Hibiscus, Melissa, Ricinus, Anthemis, Matricaria, Cymbopogon, Saussurea, Spartium, Pelargonium, Levisticum, Rosa, Santalum, Viola, Cryptocarya and Litsea genera were affirmed as generally recognized as safe (GRAS) under their conditions of intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Drs. Cohen, Eisenbrand, Fukushima, Gooderham, Guengerich, Hecht, Rietjens and Rosol are members of the Expert Panel of the Flavor and Extract Manufacturers Association. Authors Davidsen, Harman, Kelly, Ramanan and Taylor are employed by Verto Solutions which provides scientific and management support services to FEMA. A full description of the conflict of interest protections and procedures used to ensure that the FEMA Expert Panel decisions are fully objective and based solely on the merits of the available information have been published (Marnett et al., 2013) and are available on the FEMA website at https://www.femaflavor.org/gras#conflict., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The nonproteinogenic cyclic metabolite l-pipecolic acid is a chiral precursor for the synthesis of various commercial drugs and functions as a cell-protective extremolyte and mediator of defense in plants, enabling high-value applications in the pharmaceutical, medical, cosmetic, and agrochemical markets. To date, the production of the compound is unfavorably fossil-based. Here, we upgraded the strain Corynebacterium glutamicum for l-pipecolic acid production using systems metabolic engineering. Heterologous expression of the l-lysine 6-dehydrogenase pathway, apparently the best route to be used in the microbe, yielded a family of strains that enabled successful de novo synthesis from glucose but approached a limit of performance at a yield of 180 mmol mol -1 . Detailed analysis of the producers at the transcriptome, proteome, and metabolome levels revealed that the requirements of the introduced route were largely incompatible with the cellular environment, which could not be overcome after several further rounds of metabolic engineering. Based on the gained knowledge, we based the strain design on l-lysine 6-aminotransferase instead, which enabled a substantially higher in vivo flux toward l-pipecolic acid. The tailormade producer C. glutamicum PIA-7 formed l-pipecolic acid up to a yield of 562 mmol mol -1 , representing 75% of the theoretical maximum. Ultimately, the advanced mutant PIA-10B achieved a titer of 93 g L -1 in a fed-batch process on glucose, outperforming all previous efforts to synthesize this valuable molecule de novo and even approaching the level of biotransformation from l-lysine. Notably, the use of C. glutamicum allows the safe production of GRAS-designated l-pipecolic acid, providing extra benefit toward addressing the high-value pharmaceutical, medical, and cosmetic markets. In summary, our development sets a milestone toward the commercialization of biobased l-pipecolic acid., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a program for the re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavoring ingredients in food. In this publication, tenth in the series, NFCs containing a high percentage of at least one naturally occurring allylalkoxybenzene constituent with a suspected concern for genotoxicity and/or carcinogenicity are evaluated. In a related paper, ninth in the series, NFCs containing anethole and/or eugenol and relatively low percentages of these allylalkoxybenzenes are evaluated. The Panel applies the threshold of toxicological concern (TTC) concept and evaluates relevant toxicology data on the NFCs and their respective constituent congeneric groups. For NFCs containing allylalkoxybenzene constituent(s), the estimated intake of the constituent is compared to the TTC for compounds with structural alerts for genotoxicity and when exceeded, a margin of exposure (MOE) is calculated. BMDL 10 values are derived from benchmark dose analyses using Bayesian model averaging for safrole, estragole and methyl eugenol using EPA's BMDS software version 3.2. BMDL 10 values for myristicin, elemicin and parsley apiole were estimated by read-across using relative potency factors. Margins of safety for each constituent congeneric group and MOEs for each allylalkoxybenzene constituent for each NFC were determined that indicate no safety concern. The scope of the safety evaluation contained herein does not include added use in dietary supplements or any products other than food. Ten NFCs, derived from basil, estragon (tarragon), mace, nutmeg, parsley and Canadian snakeroot were determined or affirmed as generally recognized as safe (GRAS) under their conditions of intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Drs. Cohen, Eisenbrand, Fukushima, Gooderham, Guengerich, Hecht, Rietjens and Rosol are members of the Expert Panel of the Flavor and Extract Manufacturers Association. Authors Davidsen, Harman and Taylor are employed by Verto Solutions which provides scientific and management support services to FEMA. A full description of the conflict of interest protections and procedures used to ensure that the FEMA Expert Panel decisions are fully objective and based solely on the merits of the available information have been published (Marnett et al., 2013) and are available on the FEMA website at https://www.femaflavor.org/gras#conflict., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Poly-γ-glutamic acid (γ-PGA) is a bio-derived water-soluble, edible, hydrating, non-immunogenic polymer. Bacillus subtilis natto is a wild-type γ-PGA producer originally isolated from Japanese fermented natto beans whose activity has been shown to be enhanced through ion-specific activation of Extrachromosomal DNA maintenance mechanisms. Being a GRAS γ-PGA producer, this microorganism has attracted great interest in its use within an industrial context. Here we successfully synthesised amorphous, crystalline and semi-crystalline γ-PGA between 11-27 g/L. In line with circular economy principles, scalable macroalgal biomass has been evaluated as substrate for γ-PGA, displaying great potential in both yields and material composition. In this study whole cell, freeze dried seaweed -namely Laminaria digitata, Saccharina latissima and Alaria esculenta- were pre-treated by means of mechanical methods, sterilised and subsequently inoculated with B. subtilis natto. High shear mixing was found to be the most suitable pre-treatment technique. Supplemented L. digitata (9.1 g/L), S. latissima (10.2 g/L), A. esculenta (13 g/L) displayed γ-PGA yields comparable to those of standard GS media (14.4 g/L). Greatest yields of pure γ-PGA were obtained during the month of June for L. digitata (Avg. 4.76 g/L) comparable to those obtained with GS media (7.0 g/L). Further, pre-treated S. latissima and L. digitata complex media enabled for high molar mass (4,500 kDa) γ-PGA biosynthesis at 8.6 and 8.7 g/L respectively. Compared to standard GS media, algal derived γ-PGA displayed significantly higher molar masses. Further studies will be necessary to further evaluate the impact of varying ash contents upon the stereochemical properties and modify the properties of algal media based γ-PGA with the aid of key nutrients; however, the material synthesised to date can directly displace a number of fossil fuel derived chemicals in drug delivery applications, cosmetics, bioremediation, wastewater treatment, flocculation and as cryoprotectants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Parati, Philip, Mendrek, Townrow, Khalil, Tchuenbou-Magaia, Stanley, Kowalczuk, Adamus and Radecka.)

The bioconversion of jackfruit seed into a valuable product like Single-Cell Protein (SCP) was carried out using Generally Recognized As Safe (GRAS), five filamentous fungi such as Penicillium expansum, Aspergillus niger, Aspergillus oryzae, Rhizopus microsporus, and Candida intermedia by submerged fermentation. Data obtained from the study showed that supplementation of jackfruit seed extracts with inorganic nitrogen sources and glucose as a carbon source enhanced fungal biomass and SCP production. Among the various fungal strains, A. niger gave the highest biomass and protein yield of 4.01 ± 0.07 g/l and 1.82 ± 0.03 g/l, respectively, on the 9th day of fermentation followed by P. expansum (3.65 ± 0.04 g/l and 1.68 ± 0.03 g/l, respectively). C. intermedia growth was not recorded in all the experimental media. The present study revealed that fungal biomass has shown low crude fat, crude fiber, and total genomic DNA content ranged from 1.10 ± 0.20 to 4.95 ± 0.40%. A. niger and P. expansum were the most efficient in the conversion of sugar (55.83 ± 0.3% and 54.71 ± 0.4%, respectively) to yield biomass in sugar supplementation media. P. expansum and A. niger were the most promising fungal strains to produce fungal biomass protein using inexpensive agro-waste materials., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Key Message: GhSCL13-2A, a member of the PAT1 subfamily in the GRAS family, positively regulates cotton resistance to Verticillium dahliae by mediating the jasmonic acid and salicylic acid signaling pathways and accumulation of reactive oxygen species. Verticillium wilt (VW) is a devastating disease of upland cotton (Gossypium hirsutum) that is primarily caused by the soil-borne fungus Verticillium dahliae. Scarecrow-like (SCL) proteins are known to be involved in plant abiotic and biotic stress responses, but their roles in cotton defense responses are still unclear. In this study, a total of 25 GhPAT1 subfamily members in the GRAS family were identified in upland cotton. Gene organization and protein domain analysis showed that GhPAT1 members were highly conserved. GhPAT1 genes were widely expressed in various tissues and at multiple developmental stages, and they were responsive to jasmonic acid (JA), salicylic acid (SA), and ethylene (ET) signals. Furthermore, GhSCL13-2A was induced by V. dahliae infection. V. dahliae resistance was enhanced in Arabidopsis thaliana by ectopic overexpression of GhSCL13-2A, whereas cotton GhSCL13-2A knockdowns showed increased susceptibility. Levels of reactive oxygen species (ROS) and JA were also increased and SA content was decreased in GhSCL13-2A knockdowns. At the gene expression level, PR genes and SA signaling marker genes were down-regulated and JA signaling marker genes were upregulated in GhSCL13-2A knockdowns. GhSCL13-2A was shown to be localized to the cell membrane and the nucleus. Yeast two-hybrid and luciferase complementation assays indicated that GhSCL13-2A interacted with GhERF5. In Arabidopsis, V. dahliae resistance was enhanced by GhERF5 overexpression; in cotton, resistance was reduced in GhERF5 knockdowns. This study revealed a positive role of GhSCL13-2A in V. dahliae resistance, establishing it as a strong candidate gene for future breeding of V. dahliae-resistant cotton cultivars., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) applies its procedure for the safety evaluation of natural flavor complexes (NFCs) to re-evaluate the safety of Asafetida Oil (Ferula assa-foetida L.) FEMA 2108, Garlic Oil (Allium sativum L.) FEMA 2503 and Onion Oil (Allium cepa L.) FEMA 2817 for use as flavoring in food. This safety evaluation is part of a series of evaluations of NFCs for use as flavoring ingredients conducted by the Expert Panel that applies a scientific procedure published in 2005 and updated in 2018. Using a group approach that relies on a complete chemical characterization of the NFC intended for commerce, the constituents of each NFC are organized into well-defined congeneric groups and the estimated intake of each constituent congeneric group is evaluated using the conservative threshold of toxicological concern (TTC) concept. Data on the metabolism, genotoxic potential and toxicology for each constituent congeneric group are reviewed as well as studies on each NFC. Based on the safety evaluation, Asafetida Oil (Ferula assa-foetida L.), Garlic Oil (Allium sativum L.) and Onion Oil (Allium cepa L.) were affirmed as generally recognized as safe (GRASa) under their conditions of intended use as flavor ingredients., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Propionibacterium freudenreichii belongs to the class Actinobacteria (Gram positive with a high GC content). This 'Generally Recognized As Safe' (GRAS) species is traditionally used as (i) a starter for Swiss-type cheeses where it is responsible for holes and aroma production, (ii) a vitamin B12 and propionic acid producer in white biotechnologies, and (iii) a probiotic for use in humans and animals because of its bifidogenic and anti-inflammatory properties. Until now, only strain CIRM-BIA1T had been sequenced, annotated and become publicly available. Strain CIRM-BIA129 (commercially available as ITG P20) has considerable anti-inflammatory potential. Its gene content was compared to that of CIRM-BIA1 T. This strain contains 2384 genes including 1 ribosomal operon, 45 tRNA and 30 pseudogenes. [ABSTRACT FROM AUTHOR]

Background Although the concept of probiotics was known since the era of Elie Metchnikoff for more than one century ago, the attention of probiotic therapy as effective approach to prevent and treat a wide range of diseases and disordersis increased in the recent years. Probiotic therapy is the treatment of diseases by use live micro-organisms known as probiotics and the aim of this therapy is to increase the numbers of microbiota and enhance their activities until such time that microbiota can be rebalanced. In this review; the definition and properties of probiotics, their types and mechanisms of action, as well as their health beneficial effects and safety have been reported. [ABSTRACT FROM AUTHOR]

Myricitrin, a flavonol rhamnoside of myricetin extracted from the Chinese bayberry (Myrica rubra Siebold) plant, has been used in Japan since 1992 as a flavour modifier in snack foods, dairy products, and beverages. It is affirmed as generally recognised as safe (GRAS) by the US Flavour and Extract Manufacturer Association (FEMA) and is considered safe by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) at current estimated dietary exposures. In anticipation of expanded marketing, 97% pure myricitrin was fed to male and female Sprague–Dawley rats at dietary concentrations of 0.5%, 1.5% and 5.0% in a 90-day toxicity study. There was increased food consumption and decreased body weight gain in males exposed to 5% myricitrin. Blood values were within laboratory reference ranges except for mean increases in basophils in low- and high-dose males and serum phosphorus in high-dose males. In the absence of abnormal clinical or histopathological changes, these changes are not considered adverse. Based on the 90-day rat toxicity study, the no observed adverse effect level (NOAEL) is 2926 mg kg–1 day–1 in males and 3197 mg kg–1 day–1 in females. Gavage administration of myricitrin resulted in blood levels of myricitrin within 1 h after single oral doses of 250, 500 or 1000 mg kg–1 body weight, indicating direct absorption of the glycosylated form of this flavonoid. Blood levels of myricetin, a metabolite of myricitrin, were not present in rats dosed orally with 1.6 mg kg–1 myricetin, but were present only at 12 or 24 h in one of five, in three of five, and in four of five rats dosed with 250, 500 and 1000 mg myricitrin kg–1 body weight, respectively, possibly a result of hepatic conversion of myricitrin to myricetin and enterohepatic recirculation of the resulting myricetin. The current studies further support prior safety assessments of myricitrin as a food flavouring. [ABSTRACT FROM AUTHOR]

In the present study, recombinant organophosphorus hydrolase OPHC2 was successfully produced byYarrowia lipolyticaand purified. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blot analyses showed a major polypeptide band of 36 kDa. The purified enzyme was optimally active at 65°C and pH 8.5 and also displayed good thermal and pH stability using methyl parathion (O,O-dimethyl-O-4-p-nitrophenyl phosphorothioate) as a substrate. Moreover, asY. lipolyticais a non-pathogenic, generally regarded as safe (GRAS) yeast, the cell culture supernatant can be used directly on vegetables and fruits that are contaminated by organophosphorus pesticides. [ABSTRACT FROM PUBLISHER]

Background: The clinical course of advanced adrenocortical carcinoma (ACC) is heterogeneous. Our study aimed primarily to refine and make headway in the prognostic stratification of advanced ACC. Patients and methods: Patients with advanced ENSAT ACC (stage III or stage IV) at diagnosis registered between 2000 and 2009 in the ENSAT database were enrolled. The primary end point was overall survival (OS). Parameters of potential prognostic relevance were selected. Univariate and multivariate analyses were carried out: model 1 'before surgery'; model 2 'post-surgery'. Results: Four hundred and forty-four patients with advanced ENSAT ACC (stage III: 210; stage IV: 234) were analyzed. After a median follow-up of 55.2 months, the median OS was 24 months. A modified ENSAT (mENSAT) classification was validated: stage III (invasion of surrounding tissues/organs or the vena renalis/cava) and stage IVa, IVb, IVc (, 3 or >3 metastatic organs, including N, respectively). Two- or 5-year OS was 73%, 46%, 26% and 15% or 50%, 15%, 14% and 2% for stages III, IVa, IVb and IVc, respectively. In the multivariate analysis, mENSAT stages (stages IVa, IVb, or IVc, respectively) were significantly correlated with OS (P < 0.0001), as well as additional parameters: age ≥50 years (P < 0.0001), tumor- or hormone-related symptoms (P = 0.01 and 0.0, respectively) in model 1 but also the R status (P = 0.001) and Grade (Weiss >6 and/or Ki67 ≥20%, P = 0.06) in model 2. Conclusion: The mENSAT classification and GRAS parameters (Grade, R status, Age and Symptoms) were found to best stratify the prognosis of patients with advanced ACC. [ABSTRACT FROM AUTHOR]

Bacteriocin producing Lactobacillus spicheri G2, isolated from Gundruk - a traditional fermented vegetable product of North East India. L. spicheri G2 identified by morphological, biochemical techniques followed by 16S rRNA gene technique. The 16Sr RNA sequence of bacteriocin producer is registered in NCBI under accession no. JX481912. The bacteriocin producing potential of L. spicheri is being reported for the first time in the present investigation. Bacteriocin of L. spicheri G2 showed strong antagonism against food spoiling and pathogenic bacteria viz. Listeria monocytogenes, Staphlococcus aureus, Clostridium perfringens, Streptococcus mutans, Lactobacillus plantarum, Leuconostoc mesenteroides and Bacillus cereus. Bacteriocin production of L. spicheri G2 was enhanced by optimization of production time, pH of medium and incubation temperature by following one variable at a time method. Maximum bacteriocin activity (2000 AU/ml) was recorded in MRS broth at 34 h with an initial pH of 4.0 after incubating at 35 °C. The bacteriocin was purified by single step gel exclusion chromatography. Molecular weight of this novel bacteriocin was determined by SDS PAGE which was found to be 43 kDa. Purified bacteriocin was found resistant to high temperature and varied pH range but sensitive to proteolytic enzymes like trypsin and proteinase k, the characters desirable for food preservation. [ABSTRACT FROM AUTHOR]

Background: Drought is a major abiotic stress factors that reduces agricultural productivity. GRAS transcription factors are plant-specific proteins that play diverse roles in plant development. However, the functions of a number of GRAS genes identified in rice are unknown, especially the GRAS genes related to rice drought resistance have not been characterized. Results: In this study, a novel GRAS transcription factor gene named OsGRAS23, which is located in a drought-resistant QTL interval on chromosome 4 of rice, was isolated. The expression of OsGRAS23 was induced by drought, NaCl, and jasmonic acid treatments. The OsGRAS23-GFP fused protein was localized in the nucleus of tobacco epidermal cells. A trans-activation assay in yeast cells demonstrated that the OsGRAS23 protein possessed a strong transcriptional activation activity. OsGRAS23-overexpressing rice plants showed improved drought resistance and oxidative stress tolerance as well as less H2O2 accumulation compared with the wild-type plants. Furthermore, microarray analysis showed that several anti-oxidation related genes were up-regulated in the OsGRAS23-overexpressing rice plants. The yeast one hybrid test indicated that OsGRAS23 could bind to the promoters of its potential target genes. Conclusions: Our results demonstrate that OsGRAS23 encodes a stress-responsive GRAS transcription factor and positively modulates rice drought tolerance via the induction of a number of stress-responsive genes. [ABSTRACT FROM AUTHOR]

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m-Anisic acid is generally recognized as a safe (GRAS) flavouring substance. Its GRAS status and the functional groups present in the molecule make it an interesting candidate for pharmaceutical co-crystallization studies. The knowledge of m-anisic acid crystalline structure/polymorphic behaviour is important information for its applications. In this work, a crystalline structure of m-anisic acid form I, T = 105 °C, was solved by X-ray diffraction: monoclinic, space group P2/ n, with a = 13.8075(5) Å, b = 5.0221(2) Å, c = 21.4455(8) Å, β = 99.325(3)°, Mr = 331.37, V = 1467.44(10) Å, Z = 8 and R = 0.0395 (CCDC No. 985352). A recent paper by Raffo et al. (J Mol Struct 1070:86-93, ) (CCDC No. 985637) also resolved the structure, and within experimental error, the two structures are equal. The molecular flexibility of m-anisic acid results in the presence of two conformers in the unit cell, a rare case of conformational isomorphism. These two conformers were found by ab initio calculations to differ in energy by 4.9 kJ mol. Solid samples were generated by crystallization from solutions and by melt cooling. Using a multidisciplinary approach involving thermal analysis (DSC, PLTM), infrared spectroscopy and X-ray powder diffraction, a new monotropic solid form II, T = 94 °C, was identified and characterized. Polymorph II slowly transforms into polymorph I at room temperature. [ABSTRACT FROM AUTHOR]

Jumonji-C (JmjC) domain-containing protein JMJ524 plays important roles in tomato growth and development through conferring altered gibberellin responses by transcriptional regulation of a GRAS protein lacking the DELLA domain gene SGLD1. These results broaden our understanding of the JmjC domain-containing proteins, which not only control genic DNA methylation but are also involved in the GA response.Plants integrate responses to independent hormonal and environmental signals to survive adversity. In particular, the phytohormone gibberellin (GA) regulates a variety of developmental processes and stress responses. In this study, the Jumonji-C (JmjC) domain-containing gene JMJ524 was characterized in tomato. JMJ524 responded to circadian rhythms and was upregulated by GA treatment. Knockdown of JMJ524 by RNAi caused a GA-insensitive dwarf phenotype with shrunken leaves and shortened internodes. However, in these transgenic plants, higher levels of endogenous GAs were detected. A genome-wide gene expression analysis by RNA-seq indicated that the expression levels of two DELLA-like genes, SlGLD1 (‘GRAS protein Lacking the DELLA domain’) and SlGLD2, were increased in JMJ524-RNAi transgenic plants. Nevertheless, only the overexpression of SlGLD1 in tomato resulted in a GA-insensitive dwarf phenotype, suggesting that SlGLD1 acts as a repressor of GA signalling. This study proposes that JMJ524 is required for stem elongation by altering GA responses, at least partially by regulating SlGLD1. [ABSTRACT FROM PUBLISHER]

Daidzin and genistin, present in soy, can be transformed by the intestinal microbiota into equol and 5-hydroxy-equol; compounds beneficial to human health, although these compounds are only produced by a fraction of the population and in low concentrations. Hence, there is an interest in the ingestion of these compounds. Engineered lactic acid bacteria strains harbouring daidzein reductase, dihydrodaidzein reductase, tetrahydrodaidzein reductase and the dihydrodaidzein racemase genes from Slackia isoflavoniconverten s DSM22006 were incubated in five commercial soy beverages with strains with a high capacity to transform glycosylated isoflavones into their aglycones. High equol and 5-hydroxy-equol concentrations were produced in the five commercial soy beverages; the soy beverages from VegeDia and Carrefour being the ones that showed the highest concentrations of equol (456.02 ± 14.62 and 366.07 ± 25.81 μmol/L) and 5-hydroxy-equol (126.07 ± 6.23 and 121.00 ± 9.57 μmol/L). Moreover, dihydrodaidzein racemase and Bifidobacterium pseudocatenulatum INIA P815, with a high capacity to transform isoflavone glycosides into their aglycones, were necessary to achieve high equol and 5-hydroxy-equol concentrations. Bacteria generally recognized as safe produced the first fermented soy beverages enriched in equol and 5-hydroxy-equol. The high concentrations of these compounds attained in these beverages would have great potential for consumer health. • Equol and 5-hydroxy-equol are isoflavones with high beneficial effects on the human health. • The first fermented soy beverages enriched in equol and 5-hydroxy-equol were development. • The high equol and 5-hydroxy-equol concentrations would have great potential for consumer health. • Glycosidase and dihydrodaidzein racemase are essential in the production of high equol and 5-hydroxy-equol concentrations. [ABSTRACT FROM AUTHOR]