Toxicological analysis to determine the TDI of melamine in food

[[abstract]]Abstract: Intensified food safety concern over melamine has prompted national authorities to assess its tolerable daily intake (TDI) for protection of general population including young children. TDI is calculated by dividing a no-observed-adverse-effect level (NOAEL) by a safety factor (SF). Based on appropriate choices of values, the U.S. Food and Drug Administration determined two TDI values in the unit of mg per kg body weight per day as first 0.63 and then 0.063, while the World Health Organization, 0.5 and then 0.2, as a result of increasing the SF values in calculation. We used the benchmark dose analysis method on the same set of toxicity data to determine an appropriate NOAEL, and then used an SF of 1000 to obtain a TDI of 0.0081. Arguments in support of this considerably lower TDI value were provided to alert the international communit

Toxicological analysis points to a lower tolerable daily intake of melamine in food

[[abstract]]Intensified food safety concern over melamine has prompted national authorities to assess its tolerable daily intake (TDI) for protection of general population including young children. TDI is calculated by dividing a no-observed-adverse-effect level (NOAEL) by a safety factor (SF). Based on appropriate choices of values, the US Food and Drug Administration determined two TDI values in the unit of mg per kg body weight per day as first 0.63 and then 0.063, while the World Health Organization, 0.5 and then 0.2, as a result of increasing the SF values in calculation. We used a similar procedure, with judicious selection of pertinent values, to obtain a TDI of 0.0081. Arguments in support of this lower TDI value were provided to alert the international community

Dynamics of C2 toxin and chlorophyll-a formation in the dinoflagellate Alexandrium tamarense during large scale cultivation

The production of paralytic shellfish toxins (PSTs) by the dinoflagellate Alexandrium tamarense ATCI01, a toxigenic strain isolated from South China coastal waters, was studied in batch cultures in relatively large volumes (201). Under nutrient-replete conditions, this strain produced C2 toxin (C2T) as a predominant PST. In a 15-day production culture, phosphate was depleted by day 4, the stationary phase began at day 6, and the toxin productivity peaked at day 10, in which the cell content of C2T reached 76 fmol per cell. Much of the toxin was produced after the depletion of phosphate in the medium suggesting that C2T is a secondary metabolite. Aeration with small bubbles was useful in increasing cell mass and toxin yield. Chlorophyll-a (Ch1-a) was formed in algal cells until the culture entered the stationary phase, after which Chl-a began to disappear rapidly from the culture while the C2T content continued to rise. These results suggest a metabolic relationship between Chl-a and C2T. (C) 2001 Elsevier Science Ltd. All rights reserved

Effects of nitrate and phosphate on growth and C2 toxin productivity of Alexandrium tamarense CI01 in culture

Growth and C2 toxin productivity of a marine dinoflagellate, Alexandrium tamarense C101 (ATC101) which predominantly produces C2 toxin, were studied in unialgal batch cultures to optimize the concentrations of nitrate and phosphate for a maximal toxin yield. A range of start concentrations of the two major nutrients was determined in which algal growth was proportional to the nutrient concentrations used. The highest concentrations of nitrate and phosphate in this growth-enhancing range were 264 and 20 muM, respectively. In this concentration range, the C2 toxin yield (mug/l) and cellular toxin content (Qt, fmole per cell) reached a maximum at the lowest end of phosphate (5 muM) and the highest end of nitrate (264 muM). Further increase in the supply of nitrate continued to enhance the toxin yield. Our results indicated that the growth and toxin productivity of this algal strain in batch cultures had distinctly different optimal ranges of nitrate and phosphate concentrations. For a maximum toxin yield, a judicious use of phosphate under a nitrate-replete condition is called for. (C) 2002 Published by Elsevier Science Ltd

Growth and toxin production in batch cultures of a marine dinoflagellate Alexandrium tamarense HK9301 isolated from the South China Sea

Nutritional and environmental conditions were characterized for a batch culture of the marine dinoflagellate Alexandrium tamarense HK9301 isolated from the South China Sea for its growth (cells ml(-1)), cellular toxin content (Qt in fmol cell(-1)) and toxin composition (mol\%). Under a nutrient replete condition, Qt increased with cell growth and peaked at the late stationary phase. Toxin content increased with the nitrate concentration in the culture while it reached a maximum at 5 muM phosphate. When nitrate was replaced with ammonia, Qt decreased by 4.5-fold. Salinity and light intensity were important factors affecting Qt. The latter increased two-fold over the range of salinity from 15 to 30parts per thousand, while decreased 38\% as light intensity increased from 80 to 220 muE m(-2) s(-1). Toxin composition varied with growth phase and culture conditions. In nutrient replete cultures, toxin composition varied greatly in the early growth phase (first 3 days) and then C1/C2, C3/C4 and GTX1 remained relatively constant while GTX4 increased from 32 to 46\% and GTX5 decreased from 28 to 15\%. In general, the composition of GTXs was affected in a much greater extent than C toxins by changes in nutrient conditions, salinity and light intensity. This is especially true with GTX4 and GTX5. These data indicate that the cellular toxin content and toxin composition of A. tamarense HK9301 are not constant, but that they vary with growth phase and culture conditions. Use of toxin composition to identify a toxigenic marine dinoflagellate is not always valid. The data also reveal that high salinity and low light intensity, together with high nitrate and low phosphate concentrations, would favor toxin production by this species. (C) 2004 Elsevier B.V. All rights reserved

Interactions of paralytic shellfish toxins with xenobiotic-metabolizing and antioxidant enzymes in rodents

Paralytic shellfish toxins (PSTs) are neurotoxins known to block voltage-gated sodium channels in intoxicated animals and humans. Their metabolism in mammalian systems and their effects on other receptors are not as well understood. In this study, we investigated the in vitro metabolism of two classes of PSTs, gonyautoxin 2/3 (GTX2/3) and C1/2 toxins (C1/2), using rat and mouse liver enzyme preparations. We also analyzed the effects of these toxins on several antioxidant and xenobiotic-metabolizing enzymes in mice. These toxins were selected for their prevalence in the coastal waters of Southern China. When the toxins were incubated with liver preparations containing Phase I and Phase II xenobiotic metabolizing enzymes and appropriate co-factors, no transformation of the toxins was detectable. When mice were given sub-lethal doses of GTX2/3, a loss of activity was observed in hepatic ethoxyresorufin-O-deethylase, penthoxyresorufin-O-deethylase, glutathione peroxidase and superoxide dismutase, but not in glutathione S-transferase, catalase and glutathione reductase. Exposure to the same mouse units of C1/2 caused only a slight reduction in the activity of penthoxyresorufin-O-deethylase and glutathione peroxidase. Our results indicated that these toxins may not be metabolized readily in mammals and that they may cause adverse effects other than sodium channel blocking. (C) 2003 Elsevier Ltd. All rights reserved

Effects of toxic dinoflagellate Alexandrium tamarense on the energy budgets and growth of two marine bivalves

Harmful algal blooms (HAB) may impose a serious threat to aquatic lives and human health. We determined the effects of a toxic dinoflagellate Alexandrium tamarense (clone ATC101, isolated from Hong Kong coastal waters) on the energy budget, quantified as scope for growth (SFG), and the growth rate of the manila clam Ruditapes philippinarum and the green-lipped mussel Perna viridis. To quantify the SFG, clams and mussels were dosed with different amounts of toxic A. tamarense for 6 days, resulting in different toxin burdens in the tissues. Clearance rate, absorption efficiency, and respiration rate were subsequently measured in these toxin containing bivalves. Clearance rate significantly declined at the highest toxin burden in the clams only, while there was no significant difference in the clearance rate among different treatments for the mussels. The respiration rate of either bivalve was not significantly affected by toxin accumulation in the tissues. In contrast, the absorption efficiency of both clams and mussels decreased, in a concentration-dependent manner for mussels but not for clams, when the tissue accumulated the toxins. With an increase in paralytic shellfish poisoning (PSP) toxin burden, SFG in both clams and mussels was significantly reduced, primarily because of the decrease of absorption efficiency. The growth rate of juvenile clam R. philippinarum, measured as an increment in tissue dry weight over a 15 d exposure period, was significantly lower during their feeding on toxic dinoflagellate than the growth rate of clams feeding on the diatom Thalassiosira pseudonana. The juvenile mussel P. viridis, however, exhibited similar growth rates after feeding on the toxic dinoflagellates and the nontoxic diatom. This study showed that SFG can provide a sensitive and integrated measure of the effect of HAB on the physiology of bivalves. Clam R. philippinarum may be more sensitive, in terms of their energy budget, to PSP toxin accumulation than the mussel P. viridis. (C) 2001 Published by Elsevier Science Ltd. All rights reserved

In vitro aflatoxin B1-induced p53 mutations

The tumor suppressor gene p53 is commonly mutated with high frequencies at certain hot spots in human cancers. In liver cancers there is an especially high frequency of mutations at codon 249. To study the impact of carcinogen targeting and the role of cytosine methylation on the mutation spectrum, a common liver cancer carcinogen aflatoxin B1 (AFB1), was studied using the p53 cDNA template to examine mutation induction. Subsequent mutations were detected with a yeast p53 functional assay and identified by DNA sequencing. The results indicated that cytosine methylation enhances AFB1-induced guanine mutations at CpG sites. However, no mutations were detected at codon 249. (C) 2003 Elsevier Ireland Ltd. All rights reserved

Toxin composition variations in cultures of Alexandrium species isolated from the coastal waters of southern China

The composition of the paralytic shellfish toxins (PSTs) of five Alexandrium tamarense strains isolated from the coastal waters of southern China and one Alexandrium minutum strain from Taiwan Island were investigated. A. tamarense CI01 and A. tamarense Dapeng predominantly produced C2 toxin (over 90\%) with trace amounts of C1 toxin (C1), gonyautoxin-2 (GTX2) and GTX3 two strains of A. tamarense HK9301 maintained in different locations produced C1-4 toxins and GTX1, 4, 5 and 6: no PSTs were found in A. tamarense NEW, while A. minutum TW produced only GTX1-4. The toxin compositions of cultured A. tamarense strains did not vary as much during different growth phases as did the toxin composition of A. minutum TW. The toxin compositions of A. tamarense HK9301-1 did not change significantly under different salinity, light intensity, and nitrate and phosphate levels in the culture medium, although the toxin productivity varied expectably. Another strain HK9301-2 maintained in a different location produced much less toxins with a considerably different toxin composition. Under similar culture maintenance conditions for 3 years, the toxin profiles of A. tamarense HK9301-1 did not change as much as did A. tamarense CI01. Our results indicate that toxin compositions of the dinoflagellate strains are strain-specific and are subject to influence by nutritional and environmental conditions but not as much by the growth phase. Use of toxin composition in identifying a toxigenic strain requires special caution. (C) 2004 Elsevier B.V. All rights reserved