Genetically modified DP915635 maize is agronomically and compositionally comparable to non-genetically modified maize.

DP915635 maize was genetically modified (GM) to express the IPD079Ea protein for corn rootworm (Diabrotica spp.) control. DP915635 maize also expresses the phosphinothricin acetyltransferase (PAT) protein for tolerance to glufosinate herbicide and the phosphomannose isomerase (PMI) protein that was used as a selectable marker. A field study was conducted at ten sites in the United States and Canada during the 2019 growing season. Of the 11 agronomic endpoints that were evaluated, two of them (early stand count and days to flowering) were statistically significant compared with the control maize based on unadjusted p-values; however, these differences were not significant after FDR-adjustment of p-values. Composition analytes from DP915635 maize grain and forage (proximates, fiber, minerals, amino acids, fatty acids, vitamins, anti-nutrients, and secondary metabolites) were compared to non-GM near-isoline control maize (control maize) and non-GM commercial maize (reference maize). Statistically significant differences were observed for 7 of the 79 compositional analytes (16:1 palmitoleic acid, 18:0 stearic acid, 18:1 oleic acid, 18:2 linoleic acid, 24:0 lignoceric acid, methionine, and α-tocopherol); however, these differences were not significant after FDR-adjustment. Additionally, all of the values for composition analytes fell within the range of natural variation established from the in-study reference range, literature range, and/or tolerance interval. These results demonstrate that DP915635 is agronomically and compositionally comparable to non-GM maize represented by non-GM near-isoline control maize and non-GM commercial maize.

Investigation of Possible Changes Induced by RNA Silencing in Some Leaf Metabolites of Transgenic Sugar Beet Events.

Sugar beet is vulnerable to rhizomania as the most destructive viral disease. Two selected events of transgenic sugar beet carrying cassettes inducing RNA silencing mechanism, 219-T3:S3-13.2 (S3) and 6018-T3:S6-44 (S6), were shown to inhibit propagation of Beet Necrotic Yellow Vein Virus, the causative agent. As a method for signifying the substantial equivalence, we analyzed the levels of some metabolites through LC-MS in order to demonstrate possible unintended changes in the leaves of the transgenic events. There was no significant difference in the concentrations of examined key metabolites but cis-aconitate and fructose-1,6-bisphosphatase which were decreased in S3. Also, ATP was reduced in both genetically modified sugar beets. Among free amino acids, only glycine level in S6 was increased compared to the wild plant, while the production levels of 5 and 12 ones were increased in S3 compared to S6 event and the wild type plants, respectively.

Proposed Standard Test Protocols and Outcome Measures for Quantitative Comparison of Emissions from Electronic Nicotine Delivery Systems.

This study introduces and demonstrates a comprehensive, accurate, unbiased approach to robust quantitative comparison of electronic nicotine delivery systems (ENDS) appropriate for establishing substantial equivalence (or lack thereof) between inhaled nicotine products. The approach is demonstrated across a family of thirteen pen- and pod-style ENDS products. Methods employed consist of formulating a robust emissions surface regression model, quantifying the empirical accuracy of the model as applied to each product, evaluating relationships between product design characteristics and maximum emissions characteristics, and presenting results in formats useful to researchers, regulators, and consumers. Results provide a response surface to characterize emissions (total particulate matter and constituents thereof) from each ENDS appropriate for use in a computer model and for conducting quantitative exposure comparisons between products. Results demonstrate that emissions vary as a function of puff duration, flow rate, e-liquid composition, and device operating power. Further, results indicate that regulating design characteristics of ENDS devices and consumables may not achieve desired public health outcomes; it is more effective to regulate maximum permissible emissions directly. Three emissions outcome measures (yield per puff, mass concentration, and constituent mass ratio) are recommended for adoption as standard quantities for reporting by manufacturers and research laboratories. The approach provides a means of: (a) quantifying and comparing maximal emissions from ENDS products spanning their entire operating envelope, (b) comparative evaluation of ENDS devices and consumable design characteristics, and (c) establishing comparative equivalence of maximal emissions from ENDS. A consumer-oriented product emissions dashboard is proposed for comparative evaluation of ENDS exposure potential. Maximum achievable power dissipated in the coil of ENDS is identified as a potentially effective regulatory parameter.

Comparative proteome analyses of rhizomania resistant transgenic sugar beets based on RNA silencing mechanism.

Rhizomania is an economically important disease of sugar beet, which is caused by Beet necrotic yellow vein virus (BNYVV). As previously shown, RNA silencing mechanism effectively inhibit the viral propagation in transgenic sugar beet plants. To investigate possible proteomic changes induced by gene insertion and/or RNA silencing mechanism, the root protein profiles of wild type sugar beet genotype 9597, as a control, and transgenic events named 6018-T3:S6-44 (S6) and 219-T3:S3-13.2 (S3) were compared by two-dimensional gel electrophoresis. The accumulation levels of 25 and 24 proteins were differentially regulated in S3 and S6 plants, respectively. The accumulation of 15 spots were increased or decreased more than 2-fold. Additionally, 10 spots repressed or induced in both, while seven spots showed variable results in two events. All the differentially expressed spots were analyzed by MALDI-TOF-TOF mass spectrometry. The functional analysis of differentially accumulated proteins showed that most of them are related to the metabolism and defense/stress response. None of these recognized proteins were allergens or toxic proteins except for a spot identified as phenylcoumaran benzylic ether reductase, Pyrc5, which was decreased in the genetically modified S6 plant. These data are in favor of substantial equivalence of the transgenic plants in comparison to their related wild type cultivar since the proteomic profile of sugar beet root was not remarkably affected by gene transfer and activation RNA silencing mechanism.

Comparative compositional analysis of cassava brown streak disease resistant 4046 cassava and its non-transgenic parental cultivar.

Compositional analysis is an important component of an integrated comparative approach to assessing the food and feed safety of new crops developed using biotechnology. As part of the safety assessment of cassava brown streak disease resistant 4046 cassava, a comprehensive assessment of proximates, minerals, amino acids, fatty acids, vitamins, anti-nutrients, and secondary metabolites was performed on leaf and storage root samples of 4046 cassava and its non-transgenic parental control, TME 204, collected from confined field trials in Kenya and Uganda over two successive cropping cycles. Among the 100 compositional components that were assessed in samples of 4046 and control TME 204 cassava roots (47 components) and leaves (53 components), there were no nutritionally relevant differences noted. Although there were statistically significant differences between the transgenic and control samples for some parameters, in most cases the magnitudes of these differences were small ( < 20%), and in every case where comparative literature data were available, the mean values for 4046 and control cassava samples were within the range of normal variation reported for the compositional component in question. Overall, no consistent patterns emerged to suggest that biologically meaningful adverse changes in the composition or nutritive value of the leaves or storage roots occurred as an unintended or unexpected consequence of the genetic modification resulting in 4046 cassava. The data presented here provide convincing evidence of the safety of 4046 cassava with respect to its biochemical composition for food and feed, and it could be considered as safe as its non-transgenic control.

Comparative Evaluation of Nutritional and Mineral Composition Between Transgenic Sugarcane Overexpressing SoSPS 1 Gene and Non-transgenic Counterpart.

BACKGROUND AND OBJECTIVE: The high sucrose yield of transgenic sugarcane has been developed through the overexpression of gene for sucrose-phosphate synthase. Modification of the genome may result in alteration of biochemical profiles. This study was conducted to compare and evaluate the nutritional and mineral compositions between the transgenic and non-transgenic (NT) sugarcane counterpart. MATERIALS AND METHODS: Four of transgenic lines with overexpressing SoSPS 1 gene and NT sugarcane were grown in greenhouse for 11 months. The nutritional and mineral compositions from leaves and stems were analyzed at the harvest. RESULTS: Results revealed no significant differences in moisture, carbohydrates, crude fat and ash content between the transgenic lines and NT sugarcane. Protein and nitrogen contents were found to be significantly greater in steam of transgenic lines SP1 and SP3, including potassium content in both of the leaves and stems of transgenic lines. Although, the nutritional and mineral compositions were varied but their contents still within the range of Organization for Economic Co-operation and Development (OECD) reference values. CONCLUSION: The results indicated that the nutritional and mineral compositions are substantially equivalent between transgenic and NT sugarcane.

Comparing the preclinical risk profile of inhalable candidate and potential candidate modified risk tobacco products: A bridging use case.

Cigarette smoking causes major preventable diseases, morbidity, and mortality worldwide. Smoking cessation and prevention of smoking initiation are the preferred means for reducing these risks. Less harmful tobacco products, termed modified-risk tobacco products (MRTP), are being developed as a potential alternative for current adult smokers who would otherwise continue smoking. According to a regulatory framework issued by the US Food and Drug Administration, a manufacturer must provide comprehensive scientific evidence that the product significantly reduces harm and the risk of tobacco-related diseases, in order to obtain marketing authorization for a new MRTP. For new tobacco products similar to an already approved predicate product, the FDA has foreseen a simplified procedure for assessing "substantial equivalence". In this article, we present a use case that bridges the nonclinical evidence from previous studies demonstrating the relatively reduced harm potential of two heat-not-burn products based on different tobacco heating principles. The nonclinical evidence was collected along a "causal chain of events leading to disease" (CELSD) to systematically follow the consequences of reduced exposure to toxicants (relative to cigarette smoke) through increasing levels of biological complexity up to disease manifestation in animal models of human disease. This approach leverages the principles of systems biology and toxicology as a basis for further extrapolation to human studies. The experimental results demonstrate a similarly reduced impact of both products on apical and molecular endpoints, no novel effects not seen with cigarette smoke exposure, and an effect of switching from cigarettes to either MRTP that is comparable to that of complete smoking cessation. Ideally, a subset of representative assays from the presented sequence along the CELSD could be sufficient for predicting similarity or substantial equivalence in the nonclinical impact of novel products; this would require further validation, for which the present use case could serve as a starting point.

Agronomic and compositional assessment of genetically modified DP23211 maize for corn rootworm control.

DP23211 maize was genetically modified (GM) to express DvSSJ1 double-stranded RNA and the IPD072Aa protein for control of corn rootworm (Diabrotica spp.). DP23211 maize also expresses the phosphinothricin acetyltransferase (PAT) protein for tolerance to glufosinate herbicide, and the phosphomannose isomerase (PMI) protein that was used as a selectable marker. A multi-location field trial was conducted during the 2018 growing season at 12 sites selected to be representative of the major maize-growing regions of the U.S. and Canada. Standard agronomic endpoints as well as compositional analytes from grain and forage (e.g., proximates, fibers, minerals, amino acids, fatty acids, vitamins, anti-nutrients, secondary metabolites) were evaluated and compared to non-GM near-isoline control maize (control maize) and non-GM commercial maize (reference maize). A small number of agronomic endpoints were statistically significant compared to the control maize, but were not considered to be biologically relevant when adjusted using the false discovery rate method (FDR) or when compared to the range of natural variation established from in-study reference maize. A small number of composition analytes were statistically significant compared to the control maize. These analytes were not statistically significant when adjusted using FDR, and all analyte values fell within the range of natural variation established from in-study reference range, literature range or tolerance interval, indicating that the composition of DP23211 maize grain and forage is substantially equivalent to conventional maize represented by non-GM near-isoline control maize and non-GM commercial maize.

Genetically Modified Products, Perspectives and Challenges.

It is a common ground that humans have always modified the genome of both plants and animals. This intrusive process that has existed for thousands of years, many times through mistakes and failures, was initially carried out through the crossing of organisms with desirable features. This was done with the aim of creating and producing new plants and animals that would benefit humans, that is , they would offer better quality food, more opportunities for people to move and transport products, greater returns to work, resistance to diseases, etc. However, creating genetically modified organisms does not proceed without conflicts. One part of the equation concerns objections made by disputants of genetically modified organisms to the manipulation of life, as opposed to defenders who argue that it is essentially an extension of traditional plant cultivation and animal breeding techniques. There are also conflicts regarding the risks to the environment and human health from using genetically modified organisms. Concerns about the risks to the environment and human health from genetically modified products have been the subject of much debate, which has led to the development of regulatory frameworks for the evaluation of genetically modified crops. However, the absence of a globally accepted framework has the effect of slowing down technological development with negative consequences for areas of the world that could benefit from new technologies. So, while genetically modified crops can provide maximum benefits in food safety and in adapting crops to existing climate change, the absence of reforms, as well as the lack of harmonization of the frameworks and regulations about the genetic modifications results in all those expected benefits of using genetically modified crops being suspended. However, it is obvious that the evolution of genetically modified products is not going to stop. For that reason, research on the impact of genetic modification on medical technologies, agricultural production, commodity prices, land use and on the environment in general, should therefore continue.

Transcriptome profiling of the fungus Aspergillus nidulans exposed to a commercial glyphosate-based herbicide under conditions of apparent herbicide tolerance.

Glyphosate-based herbicides, such as Roundup®, are the most widely used non-selective, broad-spectrum herbicides. The release of these compounds in large amounts into the environment is susceptible to affect soil quality and health, especially because of the non-target effects on a large range of organisms including soil microorganisms. The soil filamentous fungus Aspergillus nidulans, a well-characterized experimental model organism that can be used as a bio-indicator for agricultural soil health, has been previously shown to be highly affected by Roundup GT Plus (R450: 450 g/L of glyphosate) at concentrations far below recommended agricultural application rate, including at a dose that does not cause any macroscopic effect. In this study, we determined alterations in the transcriptome of A. nidulans when exposed to R450 at a dose corresponding to the no-observed-adverse-effect level (NOAEL) for macroscopic parameters. A total of 1816 distinct genes had their expression altered. The most affected biological functions were protein synthesis, amino acids and secondary metabolisms, stress response, as well as detoxification pathways through cytochromes P450, glutathione-S-transferases, and ABC transporters. These results partly explain the molecular mechanisms underlying alterations in growth parameters detected at higher concentrations for this ascomycete fungus. In conclusion, our results highlight molecular disturbances in a soil fungus under conditions of apparent tolerance to the herbicide, and thus confirm the need to question the principle of "substantial equivalence" when applied to plants made tolerant to herbicides.

Metabolomics should be deployed in the identification and characterization of gene-edited crops.

Gene-editing techniques are currently revolutionizing biology, allowing far greater precision than previous mutagenic and transgenic approaches. They are becoming applicable to a wide range of plant species and biological processes. Gene editing can rapidly improve a range of crop traits, including disease resistance, abiotic stress tolerance, yield, nutritional quality and additional consumer traits. Unlike transgenic approaches, however, it is not facile to forensically detect gene-editing events at the molecular level, as no foreign DNA exists in the elite line. These limitations in molecular detection approaches are likely to focus more attention on the products generated from the technology than on the process in itself. Rapid advances in sequencing and genome assembly increasingly facilitate genome sequencing as a means of characterizing new varieties generated by gene-editing techniques. Nevertheless, subtle edits such as single base changes or small deletions may be difficult to distinguish from normal variation within a genotype. Given these emerging scenarios, downstream 'omics' technologies reflective of edited affects, such as metabolomics, need to be used in a more prominent manner to fully assess compositional changes in novel foodstuffs. To achieve this goal, metabolomics or 'non-targeted metabolite analysis' needs to make significant advances to deliver greater representation across the metabolome. With the emergence of new edited crop varieties, we advocate: (i) concerted efforts in the advancement of 'omics' technologies, such as metabolomics, and (ii) an effort to redress the use of the technology in the regulatory assessment for metabolically engineered biotech crops.

Composition of forage and grain from genetically modified DP202216 maize is equivalent to non-modified conventional maize (Zea mays L.).

DP202216 maize was genetically modified to increase and extend the expression of the zmm28 gene relative to native zmm28 gene expression, resulting in plants with enhanced grain yield potential. Standard nutritional and compositional parameters for maize grain and forage (e.g., proximates, fiber, minerals, amino acids, fatty acids, vitamins, anti-nutrients, secondary metabolites) from DP202216 maize were compared to grain and forage from non-modified near-isoline maize (control). Three amino acids (glycine, methionine, and serine) and two vitamins (vitamin B1 and vitamin B3) were statistically different between DP202216 and control maize grain but were not statistically different when adjusted using the false discovery rate method. These analyte values also fell within the ranges of natural variation of non-modified commercial maize varieties supporting that statistical differences were not biologically relevant. The composition of grain and forage from DP202216 maize is comparable to grain and forage from non-modified maize with a history of safe use.

Transgênicos e o princípio de equivalência substancial / Transgenics and the principle of substantial equivalence

Objetivamos discutir os principais argumentos que estão envolvidos no debate sobre a cientificidade do Princípio de Equivalência Substancial (PES), que afirma serem os OGM quimicamente equivalentes aos organismos selecionados pelas técnicas tradicionais de melhoramento, não requerendo, portanto, estudos toxicológicos adicionais. Problematizamos a cientificidade do PES, especialmente no que diz respeito à questão propriamente química. De fato, o PES estrutura-se conceitualmente na comparação quantitativa entre alguns componentes químico-biológicos da planta transgênica e os da não transgênica. Nesse sentido, as análises químicas propostas não conseguem relacionar sozinhas os possíveis efeitos bioquímicos, toxicológicos e imunológicos dos alimentos transgênicos, pois o princípio restringe as análises à composição química, molecular e analítica dos transgênicos. Emerge assim o problema do locus da incerteza científica, seja como questão epistemológica, seja como questão normativa e moral.

We aim to discuss the main arguments involved in the debate on the scientificity of the Principle of Substantial Equivalence (PSE), which claims that GMOs are chemically equivalent to organisms selected by traditional breeding techniques and therefore do not require additional toxicological studies. We question the scientific character of the PSE, especially with regard to the chemical question itself. Indeed, the PSE is conceptually structured in the quantitative comparison between some chemical--biological components of the transgenic plant and those of the non-transgenic plant. In this sense, the proposed chemical analyses cannot by themselves assess the possi-ble biochemical, toxicological and immunological effects of transgenic foods, since the principle restricts the analysis to the chemical, molecular and analytical composition of transgenics. This gives rise to the problem of the locus of scientific uncertainty, whether as an epistemological question or as a normative and moral issue.

Comparative compositional analysis of transgenic potato resistant to potato tuber moth (PTM) and its non-transformed counterpart.

In this study, the compositions of transgenic potatoes (TPs) resistant to potato tuber moth (Phthorimaea operculella) were compared with those of its non-transgenic (NTP) counterparts. The light inducible promoter, phosphoenolpyruvate carboxylase led to the expression of Cry1Ab only in the leaves and light-treated tubers of the TPs. No significant differences were found in the moisture, ash, dry weight, total soluble protein, carbohydrate, starch, fiber, ascorbate, cations, anions, fatty acids, and glycoalkaloids contents of TP and NTP. Moreover, light treatment significantly affected the contents of ascorbate, acetate and nitrite anions, palmitic, stearic and linolenic fatty acids, α-haconine and α-solanine glycoalkaloids in TP and NTP tubers. While, significant differences were observed in the amino acid contents in light-treated tubers of TPs than the NTP ones. Although, light treatment in potato tubers resulted in marked metabolic changes, all the variations observed in the metabolites compositions were found to be within the desired reference ranges for potato plants. In conclusion, the results indicated that the TPs were substantially and nutritionally equivalent to the NTP counterparts.

Quantitative risk assessment of tobacco products: A potentially useful component of substantial equivalence evaluations.

Quantitative risk assessment (QRA), a scientific, evidence-based analytical process that combines chemical and biological data to quantify the probability and potential impact of some defined risk, is used by regulatory agencies for decision-making. Thus, in tobacco product regulation, specifically in substantial equivalence (SE) evaluations, QRA can provide a useful, practical, and efficient approach to address questions that might arise regarding human health risk and potential influence on public health. In SE reporting, when differences in product characteristics may necessitate the determination of whether a new product raises different questions of public health, the results from QRA are a valuable metric. An approach for QRA in this context is discussed, which is modeled after the methodology for assessment of constituent mixtures by the US Environmental Protection Agency for environmental Superfund site assessment. Given the intent in both cases is an assessment of the public health impact resulting from the totality of exposure to a mixture of constituents, the application is appropriate. Although some uncertainties in the information incorporated may exist, relying on the most appropriate of the available data increases the confidence and decreases the uncertainty in the risk characterization using this data-driven methodology.

Proteomic analysis of the soil filamentous fungus Aspergillus nidulans exposed to a Roundup formulation at a dose causing no macroscopic effect: a functional study.

Roundup® is a glyphosate-based herbicide (GBH) used worldwide both in agriculture and private gardens. Thus, it constitutes a substantial source of environmental contaminations, especially for water and soil, and may impact a number of non-target organisms essential for ecosystem balance. The soil filamentous fungus Aspergillus nidulans has been shown to be highly affected by a commercial formulation of Roundup® (R450), containing 450 g/L of glyphosate (GLY), at doses far below recommended agricultural application rate. In the present study, we used two-dimensional gel electrophoresis combined to mass spectrometry to analyze proteomic pattern changes in A. nidulans exposed to R450 at a dose corresponding to the no-observed-adverse-effect level (NOAEL) for macroscopic parameters (31.5 mg/L GLY among adjuvants). Comparative analysis revealed a total of 82 differentially expressed proteins between control and R450-treated samples, and 85% of them (70) were unambiguously identified. Their molecular functions were mainly assigned to cell detoxification and stress response (16%), protein synthesis (14%), amino acid metabolism (13%), glycolysis/gluconeogenesis/glycerol metabolism/pentose phosphate pathway (13%) and Krebs TCA cycle/acetyl-CoA synthesis/ATP metabolism (10%). These results bring new insights into the understanding of the toxicity induced by higher doses of this herbicide in the soil model organism A. nidulans. To our knowledge, this study represents the first evidence of protein expression modulation and, thus, possible metabolic disturbance, in response to an herbicide treatment at a dose that does not cause any visible effect. These data are likely to challenge the concept of "substantial equivalence" when applied to herbicide-tolerant plants.

Developing transgenic wheat to encounter rusts and powdery mildew by overexpressing barley chi26 gene for fungal resistance.

BACKGROUND: The main aim of this study was to improve fungal resistance in bread wheat via transgenesis. Transgenic wheat plants harboring barley chitinase (chi26) gene, driven by maize ubi promoter, were obtained using biolistic bombardment, whereas the herbicide resistance gene, bar, driven by the CaMV 35S promoter was used as a selectable marker. RESULTS: Molecular analysis confirmed the integration, copy number, and the level of expression of the chi26 gene in four independent transgenic events. Chitinase enzyme activity was detected using a standard enzymatic assay. The expression levels of chi26 gene in the different transgenic lines, compared to their respective controls, were determined using qRT-PCR. The transgene was silenced in some transgenic families across generations. Gene silencing in the present study seemed to be random and irreversible. The homozygous transgenic plants of T4, T5, T6, T8, and T9 generations were tested in the field for five growing seasons to evaluate their resistance against rusts and powdery mildew. The results indicated high chitinase activity at T0 and high transgene expression levels in few transgenic families. This resulted in high resistance against wheat rusts and powdery mildew under field conditions. It was indicated by proximate and chemical analyses that one of the transgenic families and the non-transgenic line were substantially equivalent. CONCLUSION: Transgenic wheat with barley chi26 was found to be resistant even after five generations under artificial fungal infection conditions. One transgenic line was proved to be substantially equivalent as compared to the non-transgenic control.

FDA, CE mark or something else?-Thinking fast and slow.

There is a robust debate going on among the Medical Device stake-holders whether FDA is better or CE mark or something else. Currently process of obtaining an FDA approval is bogged down by ever-increasing unpredictability, inconsistency, prolonged time, and huge expense but CE mark has its own problems. Historically, the Japanese review process has tended to be the slowest among the big three but recently with the introduction of accelerated review process there has been a significant progress. While the goal of an innovator/manufacturer is to develop, manufacture and market a medical device that addresses an unmet clinical need, the requisite regulatory approval process can be very confusing. Not only there is a whole lot of jargon tossed around by regulatory affair professionals: "substantial equivalence," "PMDA," "CE mark," "Notified body," "510K" and "PMA" but the actual approval process can also be very tardy, inconsistent and expensive.

Substantial Equivalence Standards in Tobacco Governance: Statutory Clarity and Regulatory Precedent for the FSPTCA.

The Family Smoking Prevention and Tobacco Control Act (FSPTCA) of 2009 creates the first national system of premarket regulation of tobacco products in American history. The FDA must now review and give marketing authorization to all new tobacco products, based on a public health standard, before they can be legally marketed. Yet the law also contains an alternative pathway for market entry-the substantial equivalence (SE) clause-by which novel and altered tobacco products can be marketed by demonstrating their substantial equivalence to existing products. Over 99 percent of tobacco product applications sent to the FDA under the new law have used this mechanism, and loose application of the SE mechanism carries the risk of undoing the FDA's gatekeeping power under the law. We review the statutory and regulatory precedent for SE, examining the FSPTCA itself as well as regulatory precedent from drug and device regulation (from which the term substantial equivalence and much of the associated statutory language was derived). Our review of standards and scientific precedent demonstrates that exacting scrutiny under the public health standard should govern all SE reviews and that clinical data incorporating social scientific evidence should be routinely required for SE claims by tobacco product sponsors.

Metabolic Regulation of Carotenoid-Enriched Golden Rice Line.

Vitamin A deficiency (VAD) is the leading cause of blindness among children and is associated with high risk of maternal mortality. In order to enhance the bioavailability of vitamin A, high carotenoid transgenic golden rice has been developed by manipulating enzymes, such as phytoene synthase (psy) and phytoene desaturase (crtI). In this study, proteome and metabolite analyses were carried out to comprehend metabolic regulation and adaptation of transgenic golden rice after the manipulation of endosperm specific carotenoid pathways. The main alteration was observed in carbohydrate metabolism pathways of the transgenic seeds. The 2D based proteomic studies demonstrated that carbohydrate metabolism-related enzymes, such as pullulanase, UDP-glucose pyrophosphorylase, and glucose-1-phosphate adenylyltransferase, were primarily up-regulated in transgenic rice seeds. In addition, the enzyme PPDK was also elevated in transgenic seeds thus enhancing pyruvate biosynthesis, which is the precursor in the carotenoids biosynthetic pathway. GC-MS based metabolite profiling demonstrated an increase in the levels of glyceric acid, fructo-furanose, and galactose, while decrease in galactonic acid and gentiobiose in the transgenic rice compared to WT. It is noteworthy to mention that the carotenoid content, especially ß-carotene level in transgenic rice (4.3 µg/g) was significantly enhanced. The present study highlights the metabolic adaptation process of a transgenic golden rice line (homozygous T4 progeny of SKBR-244) after enhancing carotenoid biosynthesis. The presented information would be helpful in the development of crops enriched in carotenoids by expressing metabolic flux of pyruvate biosynthesis.