Toxicology and safety study of L-tryptophan and its impurities for use in broiler feed.

L-tryptophan has been utilized as a feed additive in animal nutrition to improve growth performance, as well as a dietary supplement to alleviate various emotional symptoms in humans. Despite its benefits, concerns regarding its safety arose following the outbreak of eosinophilia-myalgia syndrome (EMS) among individuals who consumed L-tryptophan. The causative material of EMS was determined to be not L-tryptophan itself, but rather L-tryptophan impurities resulting from a specific manufacturing process. To investigate the effect of L-tryptophan and its impurities on humans who consume meat products derived from animals that were fed L-tryptophan and its impurities, an animal study involving broiler chickens was conducted. The animals in test groups were fed diet containing 0.065%-0.073% of L-tryptophan for 27 days. This study aimed to observe the occurrence of toxicological or EMS-related symptoms and analyze the residues of L-tryptophan impurities in meat products. The results indicated that there was no evidence of adverse effects associated with the test substance in the investigated parameters. Furthermore, most of the consumed EMS-causing L-tryptophan impurities did not remain in the meat of broiler chickens. Thus, this study demonstrated the safety of L-tryptophan and some of its impurities as a feed additive.

Toxicology and safety study of L-tryptophan and its impurities for use in swine.

L-tryptophan, an essential amino acid for physiological processes, metabolism, development, and growth of organisms, is widely utilized in animal nutrition and human health as a feed additive and nutritional supplement, respectively. Despite its known benefits, safety concerns have arisen due to an eosinophilia-myalgia syndrome (EMS) outbreak linked to L-tryptophan consumed by humans. Extensive research has established that the EMS outbreak was caused by an L-tryptophan product that contained certain impurities. Therefore, safety validations are imperative to endorse the use of L-tryptophan as a supplement or a feed additive. This study was conducted in tertiary hybrid [(Landrace × Yorkshire) × Duroc] pigs to assess general toxicity and potential risks for EMS-related symptoms associated with L-tryptophan used as a feed additive. Our investigation elucidated the relationship between L-tryptophan and EMS in swine. No mortalities or clinical signs were observed in any animals during the administration period, and the test substance did not induce toxic effects. Hematological analysis and histopathological examination revealed no changes in EMS-related parameters, such as eosinophil counts, lung lesions, skin lesions, or muscle atrophy. Furthermore, no test substance-related changes occurred in other general toxicological parameters. Through analyzing the tissues and organs of swine, most of the L-tryptophan impurities that may cause EMS were not retained. Based on these findings, we concluded that incorporating L-tryptophan and its impurities into the diet does not induce EMS in swine. Consequently, L-tryptophan may be used as a feed additive throughout all growth stages of swine without safety concerns.

Simultaneous determination of L-tryptophan impurities in meat products.

L-tryptophan has been used as a feed additive for swine and poultry and as a nutrient supplement for humans. However, some impurities in L-tryptophan have been reported as causative components of eosinophilia-myalgia syndrome. Therefore, from a safety perspective, it is important to analyze meat samples for these impurities. This study aims to develop an analytical method for the simultaneous detection of L-tryptophan impurities in meat products using LC-MS/MS. Among the various impurities, detection methods for (S)-2-amino-3-(5-hydroxy-1H-indol-3-yl)propanoic acid (5-hydroxytryptophan) (HTP), 1-methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid (MTCA), 3a-hydroxy-1,2,3,3a,8,8a-hexahydropyrrolo-[2,3-b]-indole-2-carboxylic acid (PIC), and 1,1'-ethylidenebistryptophan (EBT) and 2-(3-indoylmethyl)-L-tryptophan (IMT) were developed. The developed method allowed simultaneous determination of these four impurities in 5 min. No interferences from the matrix were observed, and the method showed good sensitivity to each analyte. The method detection limit and limit of quantification in meat matrices were below 11.2 and 35.7 µg/kg, respectively.

[Hypereosinophilic syndrome and other rheumatic diseases with hypereosinophilia]. / Hypereosinophiles Syndrom und weitere rheumatische Erkrankungen mit Hypereosinophilie.

Among the eosinophilic diseases treated by rheumatologists other than eosinophilic granulomatosis with polyangiitis, there are further organ-related and systemic diseases with hypereosinophilia. Only the exact differential diagnostic demarcation of the diseases enables a pathogenetic oriented treatment. This article focuses on the hypereosinophilic syndromes. The potential differential diagnoses of Ig(immunoglobulin)G4-related disease, eosinophilic fasciitis and drug-induced vasculitis as well as eosinophilia-myalgia syndrome and toxic oil syndrome as historic drug-induced inflammatory rheumatic diseases are described and the clinical manifestations and treatment are summarized.

The Importance of Quality Specifications in Safety Assessments of Amino Acids: The Cases of l-Tryptophan and l-Citrulline.

The increasing consumption of amino acids from a wide variety of sources, including dietary supplements, natural health products, medical foods, infant formulas, athletic and work-out products, herbal medicines, and other national and international categories of nutritional and functional food products, increases the exposure to amino acids to amounts far beyond those normally obtained from the diet, thereby necessitating appropriate and robust safety assessments of these ingredients. Safety assessments of amino acids, similar to all food constituents, largely rely on the establishment of an upper limit [Tolerable Upper Intake Level (UL)] considered to be a guide for avoiding high intake, above which adverse or toxic effects might occur. However, reliable ULs have been difficult or impossible to define for amino acids because of inadequate toxicity studies in animals and scarce or missing clinical data, as well as a paucity or absence of adverse event reporting data. This review examines 2 amino acids that have been associated with in-market adverse events to show how quality specifications might have helped prevent the adverse clinical outcomes. We further highlight the importance of various factors that should be incorporated into an overall safety assessment of these and other amino acids. In addition to the traditional reliance on the established UL, well-defined quality specifications, review of synthesis and production strategies, potential interactions with drugs, contraindications with certain disease states, and cautionary use within certain age groups should all be taken into consideration.

Safety concerns regarding impurities in L-Tryptophan associated with eosinophilia myalgia syndrome.

L-tryptophan is one of the essential amino acids in humans and across the animal kingdom. It has been widely used as a feed additive for domestic animals and is also administered through dietary supplements in humans. Safety concerns have been raised however since a disease known as eosinophilia-myalgia syndrome (EMS) was reported to be related to L-tryptophan supplements. EMS is a rare condition characterized by inflammation in various organ systems including the muscles, skin, and lungs. Through several studies, it has been speculated that the six components generated during the process of L-tryptophan synthesis are related to the induction of EMS. In this review, we discuss the history of EMS and its controversial correlation with L-tryptophan use reported in several studies. Many in vitro and in vivo studies have been conducted to assess the putative correlation between impurities in L-tryptophan preparations and EMS, but no clear and convincing conclusions have been drawn so far.

Safety evaluation of dried L-tryptophan fermentation product in Sprague-Dawley rats.

The subchronic toxicity of oral L-tryptophan produced by fermentation with metabolically engineered Corynebacterium glutamicum was evaluated in Sprague-Dawley rats. Doses of 0, 500, 1000, and 2000 mg/kg/day were administered to groups of 10 male and 10 female rats for 90 days. For the groups administered 0 and 2000 mg/kg/day, an additional 5 male and 5 female rats were tested as a recovery group. No adverse effects associated with the test substance were observed in all rats during the 90-day administration of the product, irrespective of dose, and at 4 weeks of recovery at dosages of 0 and 2000 mg/kg/day. Furthermore, histochemical and immunohistochemical analyses for L-tryptophan-associated eosinophilia-myalgia syndrome (EMS) did not reveal significant changes in both sexes of groups administered 0 or 2000 mg/kg/day. Based on these results, it could be concluded that there were no significant adverse effects related to the test substance in all animals; therefore, dried L-tryptophan fermentation product can be used as feed additive material.

Not just eosinophilic fasciitis.

This case report describes a rare case of progressive muscle weakness in a patient treated for eosinophilic fasciitis (EF) for many years before being diagnosed with a second autoimmune disease: dermatomyositis. Our case is a report of a 65-year-old male diagnosed with eosinophilic fasciitis 7 years before being evaluated in our service at Mayo Clinic in Jacksonville, Florida, due to progressive muscle weakness despite the chronic treatment with methotrexate. Contrast-enhanced magnetic resonance imaging of the lower extremity showed enhancement throughout the thigh musculature, which led us to pursue biopsies of the fascia and muscle in order to confirm the diagnosis of EF associated with myopathy. This case illustrates the need to consider the possibility of myopathy in patients diagnosed with EF whenever muscle weakness is more prominent than expected.

Peak AAA fatty acid homolog contaminants present in the dietary supplement l-Tryptophan associated with the onset of eosinophilia-myalgia syndrome.

The eosinophilia-myalgia syndrome (EMS) outbreak that occurred in the USA and elsewhere in 1989 was caused by the ingestion of Showa Denko K.K. (SD) L-tryptophan (L-Trp). "Six compounds" detected in the L-Trp were reported as case-associated contaminants. Recently the final and most statistically significant contaminant, "Peak AAA" was structurally characterized. The "compound" was actually shown to be two structural isomers resulting from condensation reactions of L-Trp with fatty acids derived from the bacterial cell membrane. They were identified as the indole C-2 anteiso (AAA1-343) and linear (AAA2-343) aliphatic chain isomers. Based on those findings, we utilized a combination of on-line HPLC-electrospray ionization mass spectrometry (LC-MS), as well as both precursor and product ion tandem mass spectrometry (MS/MS) to facilitate identification of a homologous family of condensation products related to AAA1-343 and AAA2-343. We structurally characterized eight new AAA1-XXX/AAA2-XXX contaminants, where XXX represents the integer molecular ions of all the related homologs, differing by aliphatic chain length and isomer configuration. The contaminants were derived from the following fatty acids of the bacterial cell membrane, 5-methylheptanoic acid (anteiso-C8:0) for AAA1-315; n-octanoic acid (n-C8:0) for AAA2-315; 6-methyloctanoic acid (anteiso-C9:0) for AAA1-329; n-nonanoic acid (n-C9:0) for AAA2-329; 10-methyldodecanoic acid (anteiso-C13:0) for AAA1-385; n-tridecanoic acid (n-C13:0) for AAA2-385; 11-methyltridecanoic acid (anteiso-C14:0) for AAA1-399; and n-tetradecanoic acid (n-C14:0) for AAA2-399. The concentration levels for these contaminants were estimated to be 0.1-7.9 µg / 500 mg of an individual SD L-Trp tablet or capsule The structural similarity of these homologs to case-related contaminants of Spanish Toxic Oil Syndrome (TOS) is discussed.

Structure determination of disease associated peak AAA from l-Tryptophan implicated in the eosinophilia-myalgia syndrome.

The eosinophilia-myalgia syndrome (EMS) outbreak of 1989 that occurred in the USA and elsewhere was caused by the ingestion of l-Tryptophan (L-Trp) solely manufactured by the Japanese company Showa Denko K.K. (SD). Six compounds present in the SD L-Trp were reported to be case-associated contaminants. However, "one" of these compounds, Peak AAA has remained structurally uncharacterized, despite the fact that it was described as "the only statistically significant (p=0.0014) contaminant". Here, we employ on-line microcapillary-high performance liquid chromatography-electrospray ionization mass spectrometry (LC-MS), and tandem mass spectrometry (MS/MS) to determine that Peak AAA is in fact two structurally related isomers. Peak AAA1 and Peak AAA2 differed in LC retention times, and were determined by accurate mass-LC-MS to both have a protonated molecular ion (MH+) of mass 343.239Da (Da), corresponding to a molecular formula of C21H30N2O2, and possessing eight degrees of unsaturation (DoU) for the non-protonated molecule. By comparing the LC-MS and LC-MS-MS retention times and spectra with authentic synthetic standards, Peak AAA1 was identified as the intermolecular condensation product of L-Trp with anteiso 7-methylnonanoic acid, to afford (S)-2-amino-3-(2-((S,E)-7-methylnon-1-en-1-yl)-1H-indol-3-yl)propanoic acid. Peak AAA2 was determined to be a condensation product of L-Trp with decanoic acid, which produced (S)-2-amino-3-(2-((E)-dec-1-en-1-yl)-1H-indol-3-yl)propanoic acid.

Eosinophilia in Rheumatologic/Vascular Disorders.

Peripheral and tissue eosinophilia can be a prominent feature of several unique rheumatologic and vascular diseases. These diseases span a wide range of clinical features, histologic findings, therapeutic approaches, and outcomes. Despite the rare nature of these entities--which makes large-scale studies challenging--knowledge has continued to grow regarding their epidemiology, pathophysiology, and management. This review compares and contrasts 5 rheumatologic and vascular conditions in which eosinophilia can be seen: eosinophilic granulomatosis with polyangiitis (Churg-Strauss), immunoglobulin G4-related disease, diffuse fasciitis with eosinophilia, eosinophilia-myalgia syndrome, and eosinophilic myositis.

Eosinophilia-myalgia syndrome induced by excessive L-tryptophan intake from cashew nuts.

Eosinophilia is characterized by more than 0.5 × 109 eosinophils per liter in the full blood count. A wide range of conditions, from asthma to parasitic infections, autoimmune diseases, and certain forms of cancer, have been known to trigger abnormally high amount of eosinophils. It is essential to reach the correct diagnosis and treat the underlying disease aggresively. Definition of the eosinophilia-myalgia syndrome was offered in 1980s by Centers for Disease Control and Prevention for surveillance purposes, and criteria were revised in 2001, with high specificity. We report a case of 59-year old female who started a special weight-reducing diet regimen that included excessive cashew nut ingestion. Several months after she has presented with periferal blood eosinophilia and constitutional symptoms. Detailed work-up has not found elements for haematological, systemic autoimmune, neoplastic or infectious disease. She was diagnosed with eosinophilia-myalgia syndrome due to extreme L-tryptophan intake, a compound found in the cashew nut's oil. She responded well to cashew nut withdrawal and steroid therapy. In the follow-up period she remained stable with normal eosinophil count and there was not a need for any specific therapy.

Insuficiência respiratória aguda como manifestação da síndrome de eosinofilia-mialgia associada à ingestão de L-triptofano / Acute respiratory failure as a manifestation of eosinophilia-myalgia syndrome associated with L-tryptophan intake

A síndrome da eosinofilia-mialgia foi descrita em 1989 em pacientes que apresentavam mialgia progressiva e incapacitante e eosinofilia sérica, nos líquidos e secreções. A maioria dos pacientes relatava uso prévio de L-triptofano. Sintomas respiratórios são relatados em até 80 por cento dos casos, eventualmente como manifestação única. O tratamento inclui suspensão da droga e corticoterapia. Relatamos o caso de uma mulher de 61 anos com insuficiência respiratória aguda após uso de L-triptofano, hidroxitriptofano e outras drogas. A paciente apresentava eosinofilia no sangue, lavado broncoalveolar e derrame pleural. Após a suspensão da medicação e corticoterapia, houve melhora clínica e radiológica em poucos dias.

Eosinophilia-myalgia syndrome was described in 1989 in patients who presented progressive and incapacitating myalgia and eosinophilia in blood, fluids and secretions. Most patients report previous L-tryptophan intake. Respiratory manifestations are found in up to 80 percent of the cases, occasionally as the only manifestation. Treatment includes drug discontinuation and administration of corticosteroids. Here, we describe the case of a 61-year-old female admitted with acute respiratory failure after using L-tryptophan, hydroxytryptophan and other drugs. The patient presented eosinophilia, together with elevated eosinophil counts in the bronchoalveolar lavage and pleural effusion. After discontinuation of the drugs previously used, corticosteroids were administered, resulting in clinical and radiological improvement within just a few days.