Comparison of regulatory pathways for the approval of advanced therapies in the European Union and the United States

Background aimsRegulatory agencies in the European Union (EU) and in the United States of America (USA) have adapted and launched regulatory pathways to accelerate patient access to innovative therapies, such as advanced therapy medicinal products (ATMPs). The aim of this study is to analyze similarities and differences between regulatory pathways followed by the approved ATMPs in both regions.MethodsA retrospective analysis of the ATMPs approved by EU and US regulatory agencies was carried out until May 31, 2020. Data were collected on the features and timing of orphan drug designation (ODD), scientific advice (SA), expedited program designation (EP), marketing authorization application (MAA) and marketing authorization (MA) for both regions.ResultsIn the EU, a total of fifteen ATMPs were approved (eight gene therapies, three somatic cell therapies, three tissue-engineered products and one combined ATMP), whereas in the USA, a total of nine were approved (five gene therapies and four cell therapies); seven of these were authorized in both regions. No statistical differences were found in the mean time between having the ODD or EP granted and the start of the pivotal clinical trial or MAA in the EU and USA, although the USA required less time for MAA assessment than the EU (mean difference, 5.44, P = 0.012). The MAA assessment was shorter for those products with a PRIME or breakthrough designation.. No differences were found in the percentage of ATMPs with expedited MAA assessment between the EU and the USA (33.3% versus 55.5%, respectively, P = 0.285) or in the time required for the MAA expedited review (mean difference 4.41, P = 0.105). Approximately half of the products in both regions required an Advisory Committee during the MAA review, and 60% required an oral explanation in the EU. More than half of the approved ATMPs (67% and 55.55% in the EU and the USA, respectively) were granted an ODD, 70% by submitting preliminary clinical data in the EU. The mean number of SA and protocol assistance per product conducted by the European Medicines Agency was 1.71 and 3.75, respectively, and only 13% included parallel advice with health technology assessment bodies. A total of 53.33% of the products conducted the first SA after the pivotal clinical study had started, reporting more protocol amendments. Finally, of the seven ATMPs authorized in both regions, the type of MA differed for only two ATMPs (28.6%), and four out of eight products non-commercialized in the USA had a non-standard MA in the EU.ConclusionsThe current approved ATMPs mainly target orphan diseases. Although EU and US regulatory procedures may differ, the main regulatory milestones reached by the approved ATMPs are similar in both regions, with the exception of the time for MAA evaluation, the number of authorized products in the regions and the type of authorization for some products. More global regulatory convergence might further simplify and expedite current ATMP development in these regions.     

Use of 'natural' products as alternatives to antibiotic feed additives in ruminant production

The banning in 2006 of the use of antibiotics as animal growth promoters in the European Union has increased demand from producers for alternative feed additives that can be used to improve animal production. This review gives an overview of the most common non-antibiotic feed additives already being used or that could potentially be used in ruminant nutrition. Probiotics, dicarboxylic acids, enzymes and plant-derived products including saponins, tannins and essential oils are presented. The known modes of action and effects of these additives on feed digestion and more especially on rumen fermentations are described. Their utility and limitations in field conditions for modern ruminant production systems and their compliance with the current legislation are also discussed.     

Fermented pig liquid feed: nutritional, safety and regulatory aspects

Fermented liquid feed has been lately much investigated in order to compensate the use of antibiotics in pig production. The fermentation process has been claimed to be the reason of the benefits associated with this type of feeding. However, contradictory results have been obtained in feeding trials due to the variable conditions in each experiment. This review focuses on the different factors that would ensure a proper fermentation with all its beneficial effects. In particular, while fermenting a liquid diet with lactic acid bacteria has been shown to improve the quality of feed and to be beneficial to the health of the animals, spontaneously fermented liquid feed appears to be unsafe for the pigs and eventually affects the consumers' safety. Consequently, the use of specific starters or inoculants to ensure the proper fermentation could be a practical solution. The regulatory status of fermented liquid feed in the EU is still unclear, but the use of specific inoculants could be considered as a special case of microbial feed additives.     

蛹虫草饲料添加剂的研究现状及展望

蛹虫草饲料添加剂包括蛹虫草子实体、蛹虫草培养残基、蛹虫草及其培养残基提取物、蛹虫草菌固液发酵产物、微生物发酵蛹虫草残基等产品.蛹虫草饲料添加剂含有粗蛋白、粗脂肪、氨基酸等营养成分,以及虫草素、腺苷、多糖等活性成分,在畜禽、反刍动物、水产品等动物养殖中的应用均获得较好的效果.对蛹虫草子实体、蛹虫草培养残基、蛹虫草及其培养...     

Tumorigenicity assessment of cell therapy products: The need for global consensus and points to consider

Pluripotent stem cells offer the potential for an unlimited source for cell therapy products. However, there is concern regarding the tumorigenicity of these products in humans, mainly due to the possible unintended contamination of undifferentiated cells or transformed cells. Because of the complex nature of these new therapies and the lack of a globally accepted consensus on the strategy for tumorigenicity evaluation, a case-by-case approach is recommended for the risk assessment of each cell therapy product. In general, therapeutic products need to be qualified using available technologies, which ideally should be fully validated. In such circumstances, the developers of cell therapy products may have conducted various tumorigenicity tests and consulted with regulators in respective countries. Here, we critically review currently available in vivo and in vitro testing methods for tumorigenicity evaluation against expectations in international regulatory guidelines. We discuss the value of those approaches, in particular the limitations of in vivo methods, and comment on challenges and future directions. In addition, we note the need for an internationally harmonized procedure for tumorigenicity assessment of cell therapy products from both regulatory and technological perspectives.     

The EU’s GM crop conundrum: Did the EU policy strategy to convert EFSA GMO guidance into legislation deliver on its promises?

Efforts by the EU to improve its regulatory framework for importing GM food and feed have done nothing to make the process easier and more predictable for applicants. Subject Categories: Biotechnology & Synthetic Biology, Economics, Law & Politics, Plant Biology

The first genetically modified (GM) crops were introduced more than two decades ago and have been planted globally on more than 190 million hectares (ISAAA, 2020), a surface area larger than all the arable land in the EU. Thousands of risk assessments have consistently concluded that they are as safe as conventional crops in regard to human and animal health (Smyth et al, 2021) and many countries have been growing GM crops for years. Despite political commitments to innovation and investments into research (EC, 2010), the EU is still lagging behind in adopting this technology on a wider scale owing to diverging views among its member states, the European Commission (EC) and the European parliament. Various attempts to resolve this tension by legal and regulatory means have created the most cumbersome and byzantine regulatory system for GM crops in the world. The Implementing Regulation (EU) No 503/2013, meant to ease the regulatory process, has made things even more complicated.

Various attempts to resolve this tension by legal and regulatory means have created the most cumbersome and byzantine regulatory system for GM crops in the world.

A major conundrum for the EU is the need to import large quantities of protein‐rich crops such as soybean to supply the continent’s livestock industry with high‐quality feed.In the light of the current Russia–Ukraine situation, which has added a layer of instability to already tense markets, the importance of the global agricultural market to ensure food security is even more pronounced.Given the high adoption rate of GM crops outside the EU, most of these imported commodities inevitably contain GM crops. Under EU law, food and feed products that contain or were produced from GM crops need an import authorisation by the European Commission (EC), which is a lengthy, costly and unpredictable process.In 2002, the EU set up a centralised review system under Regulation (EC) 178/2002 (the General Food Law Regulation) and an independent scientific body to conduct this review: the European Food Safety Authority (EFSA). EFSA is responsible for performing the risk assessment for food and feed regulated products, including GM crops; their advice “opinion” is used by the EC to draft a decision whether or not to authorise import. EU member states then vote whether or not to follow the EC’s draft decision. To date, not a single GM product has received a qualified majority decision for authorisation. The EC then makes the final decision based on EFSA’s risk assessment.There are many reasons why the member states disagree, mostly owing to political and economic agendas. Some members with a large and important agri‐food sector tend to vote in line with EFSA’s opinions, while others consistently vote against authorisation or abstain their vote mainly for political reasons. This ongoing disagreement has made it very difficult to establish an EU‐wide policy for agricultural biotechnology.

…the continuous proliferation, update and reinterpretation of EU requirements means that studies that were conducted in compliance with the guidelines at a particular time may no longer comply with changed requirements…

     

Fermented liquid feed for pigs

Since the announcement of the ban on the use of antibiotics as antimicrobial growth promoters in the feed of pigs in 2006 the investigation towards alternative feed additives has augmented considerably. Although fermented liquid feed is not an additive, but a feeding strategy, the experimental work examining its possible advantages also saw a rise. The use of fermented liquid feed (FLF) has two main advantages, namely that the simultaneous provision of feed and water may result in an alleviation of the transition from the sow milk to solid feed and may also reduce the time spent to find both sources of nutrients, and secondly, that offering FLF with a low pH may strengthen the potential of the stomach as a first line of defence against possible pathogenic infections. Because of these two advantages, FLF is often stated as an ideal feed for weaned piglets. The results obtained so far are rather variable, but in general they show a better body weight gain and worse feed/gain ratio for the piglets. However, for growing-finishing pigs on average a better feed/gain ratio is found compared to pigs fed dry feed. This better performance is mostly associated with less harmful microbiota and better gut morphology. This review provides an overview of the current knowledge of FLF for pigs,dealing with the FLF itself as well as its effect on the gastrointestinal tract and animal performance.     

Comparison of new Brazilian legislation for the approval of advanced therapy medicinal products with existing systems in the USA,European Union and Japan

Background aimsAdvanced therapy medicinal products (ATMPs) are a class of biological products for human use that are based on genes, cells and tissues. The first ATMP received marketing authorization in Europe in 2009, whereas Brazil granted the first authorization in 2020. The objective of this study was to compare the regulatory models adopted by Brazil, the USA, Japan and the European Union, which comprise the member countries of the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use, with regard to the marketing authorization of ATMPs.MethodsThe authors performed a review of the scientific literature and official documents of the regulatory agencies in the aforementioned countries.ResultsThe legislation and regulatory guidelines adopted by the regulatory agencies exhibit similarities and differences. It was not possible to assess whether these differences can be translated into divergent final recommendations by regulatory authorities upon a request for marketing authorization.ConclusionsIn the future, it will be appropriate to start a progressive process of harmonization between these agencies in terms of terminology, legal recommendations and characterization requirements. This is particularly important for emerging countries such as Brazil. In this sense, some measures can be taken to achieve alignment between regulators.     

Patient access to and ethical considerations of the application of the European Union hospital exemption rule for advanced therapy medicinal products

Hospital exemption (HE) is a regulated pathway that allows the use of advanced therapy medicinal products (ATMPs) within the European Union (EU) under restrictive conditions overseen by national medicine agencies. In some EU countries, HE is granted for ATMPs with no demonstrated safety and efficacy; therefore, they are equivalent to investigational drugs. In other countries, HE is granted for ATMPs with demonstrated quality, safety and efficacy and for which centralized marketing authorization has not been requested. The Committee on the Ethics of Cell and Gene Therapy of the International Society for Cell & Gene Therapy reflects here on the ethical issues concerning HE application from the perspective of the patient, including risk–benefit balance, accessibility and transparency, while providing evidence that HE must not be regarded as a conduit for unproven and unethical ATMP-based interventions. Indeed, HE represents a legal instrument under which a patient's need for access to novel ATMPs is reconciled with ethics. Moreover, for some unmet medical needs, HE is the only pathway for accessing innovative ATMPs. Nonetheless, HE harmonization across EU Member States and limitations of ATMP use under the HE rule when similar products have already been granted centralized marketing authorization to avoid a parallel regulatory pathway are controversial issues whose political and economic consequences are beyond the scope of this review. Finally, the institution of an EU registry of HE applications and outcomes represents a priority to improve transparency, reduce patient risks, increase efficiency of health systems, facilitate company awareness of business opportunities and boost progressive entry of ATMPs into the therapeutic repertoire of health systems.     

Fusarium mycotoxins: Effects on reproductive function in domestic animals—A review

On a global scale, cereal grains and animal feed may be contaminated with trichothecenes, such as deoxynivalenol and T-2 toxin, zearalenone (ZEA), and fumonisins, the major mycotoxins of Fusarium fungi. Of these mycotoxins, ZEA is unequivocally implicated in reproductive disorders of swine and other domestic animals. Experiments in vivo and in vitro indicate that ZEA and its metabolites exert estrogenic effects resulting in functional and morphological alterations in reproductive organs. Recently, the potential of trichothecenes and fumonisins to cause reproductive disorders in domestic animals has been investigated. The present review summarizes the toxicological data on the effects of Fusarium mycotoxins on ovarian function, testicular function, placenta and fetus, and puberty/sexual maturity of domestic animals. The results of in vivo animal studies and in vitro tests are reported and discussed.     

Strategic Focus on 3R Principles Reveals Major Reductions in the Use of Animals in Pharmaceutical Toxicity Testing

The principles of the 3Rs, Replacement, Reduction and Refinement, are being increasingly incorporated into legislations, guidelines and practice of animal experiments in order to safeguard animal welfare. In the present study we have studied the systematic application of 3R principles to toxicological research in the pharmaceutical industry, with particular focus on achieving reductions in animal numbers used in regulatory and investigatory in vivo studies. The work also details major factors influencing these reductions including the conception of ideas, cross-departmental working and acceptance into the work process. Data from 36 reduction projects were collected retrospectively from work between 2006 and 2010. Substantial reduction in animal use was achieved by different strategies, including improved study design, method development and project coordination. Major animal savings were shown in both regulatory and investigative safety studies. If a similar (i.e. 53%) reduction had been achieved simultaneously within the twelve largest pharmaceutical companies, the equivalent reduction world-wide would be about 150,000 rats annually. The results point at the importance of a strong 3R culture, with scientific engagement, collaboration and a responsive management being vital components. A strong commitment in leadership for the 3R is recommended to be translated into cross-department and inter-profession involvement in projects for innovation, validation and implementation. Synergies between all the three Rs are observed and conclude that in silico-, in vitro- and in vivo-methods all hold the potential for applying the reduction R and should be consequently coordinated at a strategic level.     

Saccharomyces cerevisiae as a probiotic feed additive to non and pseudo-ruminant feeding: a review

The production of livestock and poultry faces major challenges to meet the global demand for meat and dairy products and eggs due to a steady increase in the world’s population and the ban of antibiotics in animal production. This ban has forced animal nutritionists to seek for natural alternatives to antibiotics. In this context, the yeast Saccharomyces cerevisiae has received considerable attention in the last decade. It has been reported that feed supplementation with live yeast cells improve feed efficiency, enhance feed digestibility, increase animal performance, reduce the number of pathogenic bacteria, improve animal health and reduce the negative environmental impacts of livestock production. The current review sheds light on the effects of the use of live S. cerevisiae cells in the diets of nonruminant and pseudo-ruminant’s animals and the mechanisms by which they exert its effects. This review work revealed that the addition of S. cerevisiae in poultry feed causes a phenomenon called competitive exclusion of pathogenic bacteria capable of causing disease adhere to the yeast surface, and so removing a large amount of harmful micro-organisms and allowing the Animal defend more effectively, the production of antimicrobial agents, the balancing the gut microbiota and stimulation of host adaptive immune system and improving gut morphological structure, thus these benefits are reflected on the overall poultry health. In addition, in the presence of live S. cerevisiae cells, the immunity of rabbits was improved due to the high number of white blood cell. In addition, apparent digestibility of acid and neutral detergent fibre was improved in horses and rabbits. Saccharomyces cerevisiae in pig diets augment mucosal immunity by increasing IgM and IgA activity against pathogens, enhance intestinal development and function, adsorb mycotoxins, modulate gut microbiota and reduce postweaning diarrhoea.     

Significance of Antioxidant Potential of Plants and its Relevance to Therapeutic Applications

Oxidative stress has been identified as the root cause of the development and progression of several diseases. Supplementation of exogenous antioxidants or boosting endogenous antioxidant defenses of the body is a promising way of combating the undesirable effects of reactive oxygen species (ROS) induced oxidative damage. Plants have an innate ability to biosynthesize a wide range of non-enzymatic antioxidants capable of attenuating ROS- induced oxidative damage. Several in vitro methods have been used to screen plants for their antioxidant potential, and in most of these assays they revealed potent antioxidant activity. However, prior to confirming their in vivo therapeutic efficacy, plant antioxidants have to pass through several physiopharmacological processes. Consequently, the findings of in vitro and in vivo antioxidant potential assessment studies are not always the same. Nevertheless, the results of in vitro assays have been irrelevantly extrapolated to the therapeutic application of plant antioxidants without undertaking sufficient in vivo studies. Therefore, we have briefly reviewed the physiology and redox biology of both plants and humans to improve our understanding of plant antioxidants as therapeutic entities. The applications and limitations of antioxidant activity measurement assays were also highlighted to identify the precise path to be followed for future research in the area of plant antioxidants.     

Tumor treating fields: An emerging treatment modality for thoracic and abdominal cavity cancers

Tumor treating fields (TTFields)-an intermediate-frequency, electric field therapy-has emerged as a promising alternative therapy for the treatment of solid cancers. Since the first publication describing the anticancer effects of TTFields in 2004 there have been numerous follow-up studies by other groups, either to confirm the efficacy of TTFields or to study the primary mechanism of interaction. The overwhelming conclusion from these in vitro studies is that TTFields reduce the viability of aggressively replicating cell lines. However, there is still speculation as to the primary mechanism for this effect; moreover, observations both in vitro and in vivo of inhibited migration and metastases have been made, which may be unrelated to the originally proposed hypothesis of replication stress. Adding to this, the in vivo environment is much more complex spatially, structurally, and involves intricate networks of cell signaling, all of which could change the efficacy of TTFields in the same way pharmaceutical interventions often struggle transitioning in vivo. Despite this, TTFields have shown promise in clinical practice on multiple cancer types, which begs the question: has the primary mechanism carried over from in vitro to in vivo or are there new mechanisms at play? The goal of this review is to highlight the current proposed mechanism of action of TTFields based primarily on in vitro experiments and animal models, provide a summary of the clinical efficacy of TTFields, and finally, propose future directions of research to identify all possible mechanisms in vivo utilizing novel tumor-on-a-chip platforms.     

Mould and mycotoxin contamination of pig feed in northwest Croatia

The aim of this study was to investigate the contamination of pig feed with moulds and the occurrence of mycotoxins. A total of 30 feed samples were collected at different animal feed factories in the north-western part of Croatia. Mycological analysis showed that the total number of moulds ranged from 1?×?10³ to 1?×?10⁵?cfu/g with samples contaminated with Aspergillus spp. (63?%), Penicillium spp. (80?%), and Fusarium spp. (77?%). A determination of aflatoxin B1 (AFB₁), ochratoxin A (OTA), zearalenone (ZEA), deoxynivalenol (DON), T-2 toxin (T-2) and fumonisin (FUM) concentration was done using the validated ELISA method. The mean concentrations of AFB₁ (0.5?±?0.6???g/kg), OTA (1.53?±?0.42???g/kg) and FUM (405?±?298???g/kg) were below the maximum levels or recommended values in the EU in all the investigated samples. The observed results indicated an increased contamination of pig feed with Fusarium mycotoxins DON and ZEA with mean concentrations of 817?±?447 and 184?±?214???g/kg, higher than recommended in 40 and 17?% of the analysed samples, respectively.     

Potential applications and emerging trends of species of the genus Dietzia: a review

Interest in attractive biological sources with multicriteria applications has been increasing during recent years. This study scrutinized the applications of Dietzia bacteria for future prospects. Apart from such present and well-established applications—as in therapeutic biotreatments for adult paratuberculosis animals, production of carotenoid pigments, and animal feed additives—their uses in biosurfactants and biodemulsifiers production, the pollutants bioremediation, biodegradation of petroleum hydrocarbons and crude oil and also production of extracellular polymeric substances (EPSs) have been exploited. The use of these bacteria as a biotechnological tool may lead to improve the optimization and quality assurance of food ingredients and products, the capability of degradation and remediation of environmental pollutants, and the efficiency of bioconversion systems for energy recovery and bioprocessing of value-added products.     

Effect of animal mixing as a stressor on biomarkers of autophagy and oxidative stress during pig muscle maturation

The objective of this work was to study the postmortem evolution of potential biomarkers of autophagy (Beclin 1, LC3-II/LC3-I ratio) and oxidative stress (total antioxidant activity, TAA; superoxide dismutase activity, SOD and catalase activity, CAT) in the Longissimus dorsi muscle of entire male ((Large White×Landrace)×Duroc) pigs subjected to different management treatments that may promote stress, such as mixing unfamiliar animals at the farm and/or during transport and lairage before slaughter. During the rearing period at the farm, five animals were never mixed after the initial formation of the experimental groups (unmixed group at the farm, UF), whereas 10 animals were subjected to a common routine of being mixed with unfamiliar animals (mixed group at the farm, MF). Furthermore, two different treatments were used during the transport and lairage before slaughter: 10 pigs were not mixed (unmixed group during transport and lairage, UTL), whereas five pigs were mixed with unfamiliar animals on the lorry and during lairage (mixed group during transport and lairage, MTL). These mixing treatments were then combined into three pre-slaughter treatments – namely, UF-UTL, MF-UTL and MF-MTL. The results show that MF-UTL and MF-MTL increased significantly the muscle antioxidant defense (TAA, SOD and CAT) at short postmortem times (4 and 8 h; P<0.001), followed by an earlier depletion of the antioxidant activity at 24 h postmortem (P<0.05). We also found that mixing unfamiliar animals, both at the farm and during transport and lairage, triggers postmortem muscle autophagy, which showed an earlier activation (higher expression of Beclin 1 and LC3-II/LC3-I ratio at 4 h postmortem followed by a decreasing pattern of this ratio along first 24 h postmortem) in the muscle tissues of animals from the MF-UTL and MF-MTL groups, as an adaptive strategy of the muscle cells for counteracting induced stress. From these results, we propose that monitoring the evolution of the main biomarkers of autophagy (Beclin 1, LC3-II/LC3-I ratio) and muscle antioxidant defense (TAA, SOD, CAT) in the muscle tissue within the first 24 h postmortem may help the detection of animal stress and its potential effect on the postmortem muscle metabolism.     

Ability of a commercial feed additive to modulate expression of innate immunity in sheep immunosuppressed with dexamethasone

In the first study, we tested the ability of a commercial feed additive (OmniGen-AF) to affect markers of innate immunity in immunosuppressed sheep and the ability of a pathogen challenge (mould) to affect the immune response to the additive. Treatments consisted of (1) control, (2) immunosuppressed with dexamethasone (DEX), (3) immunosuppressed plus the feed additive, (4) immunosuppressed plus Aspergillus fumigatus and (5) immunosuppressed, A. fumigatus and the additive. Animal health was monitored and indexes of innate immunity (neutrophil L-selectin and interleukin-1β (IL-1β)) were collected. DEX caused immunosuppression (i.e. reduced abundance of neutrophil L-selectin and IL-1β). This immunosuppressive effect was countered by the provision of the additive in the ration. Provision of mould in the ration increased the ability of the additive to regulate markers of innate immune function. A second study was completed to re-assess the properties of the additive and other feed products. The study consisted of seven treatments: (1) immunosuppressed, (2) immunosuppressed with additive, (3) immunosuppressed with additive in pelleted form (low-temperature pellet) and (4) immunosuppressed with additive in a high-temperature pellet. The remaining three treatments assessed abilities of three other additives to regulate markers of innate immune function. In this study, OmniGen-AF increased expression of neutrophil L-selectin abundance in immunosuppressed animals and this was unaffected by the pelleting temperature. None of the other additives affected markers of innate immunity. In these studies we discovered mechanisms by which a feed product may affect the immune function of ruminant livestock. The product countered DEX-dependent down-regulation of markers of innate immune function and its actions were enhanced by the presence of pathogen (mould) in the ration.     

Biomass of Spirulina maxima enriched by biosorption process as a new feed supplement for swine

This paper deals with the new mineral feed additives with Cu produced in a biosorption process from a semi-technical scale. The natural biomass of edible microalga Spirulina sp. was enriched with Cu(II) and then used as a mineral supplement in feeding experiments on swine to assess its nutrition properties. A total of 24 piglets divided into two groups (control and experimental) were used to determine the bioavailability of a new generation of mineral feed additives based on Spirulina maxima. The control group was feed using traditional inorganic supplements of microelements, while the experimental group was fed with the feed containing the biomass of S. maxima enriched with Cu by biosorption. The apparent absorption was 30 % (P?<?0.05) higher in the experimental group. No effect on the production results (average daily feed intake, average daily gain, feed conversion ratio) was detected. It was found that copper concentration in feces in the experimental group was 60 % (P?<?0.05) lower than in the control group. The new preparation—a dietary supplement with microelements produced by biosorption based on biomass of microalgae S. maxima—is a promising alternative to currently used inorganic salts as the source of nutritionally important microelements.