Response to Intravenous N-Acetylcysteine Supplementation in Critically Ill Patients with COVID-19.

Administering N-acetylcysteine (NAC) could counteract the effect of free radicals, improving the clinical evolution of patients admitted to the Intensive Care Unit (ICU). This study aimed to investigate the clinical and biochemical effects of administering NAC to critically ill patients with COVID-19. A randomized controlled clinical trial was conducted on ICU patients (n = 140) with COVID-19 and divided into two groups: patients treated with NAC (NAC-treated group) and patients without NAC treatment (control group). NAC was administered as a continuous infusion with a loading dose and a maintenance dose during the study period (from admission until the third day of ICU stay). NAC-treated patients showed higher PaO2/FiO2 (p ≤ 0.014) after 3 days in ICU than their control group counterparts. Moreover, C-reactive protein (p ≤ 0.001), D-dimer (p ≤ 0.042), and lactate dehydrogenase (p ≤ 0.001) levels decreased on the third day in NAC-treated patients. Glutathione concentrations decreased in both NAC-treated (p ≤ 0.004) and control (p ≤ 0.047) groups after 3 days in ICU; whereas glutathione peroxidase did not change during the ICU stay. The administration of NAC manages to improve the clinical and analytical response of seriously ill patients with COVID-19 compared to the control group. NAC is able to stop the decrease in glutathione concentrations.

Erythrocyte Zn concentration and antioxidant response after supplementation with Zn in a postmenopausal population. A double-blind randomized trial.

BACKGROUND & AIMS: Menopausal hormonal changes increase the risk of deficiencies of minerals such as zinc (Zn), which could further worsen the decreased antioxidant defense of postmenopausal women. This study assesses the effect of 8 weeks of Zn supplementation upon the antioxidant status and clinical nutritional parameters of a postmenopausal population. METHODS: Fifty-one postmenopausal women were divided into two groups: placebo (PbG) and zinc supplementation (ZnG). Mineral status was determined by Flame Atomic Absorption Spectrophotometry (FAAS). Total Antioxidant Capacity (TAC) and Superoxide Dismutase (SOD) were analyzed by kinetic colorimetric methods. Glutathione Peroxidase (GPx) was assessed by an enzymatic immunological method. RESULTS: Poor Zn status was initially observed in erythrocyte samples. Total antioxidant capacity showed a significant correlation (r = 0.730; p < 0.05) to erythrocyte Zn after the intervention (ZnG: r = 0.96; p < 0.001). Moreover, erythrocyte Zn concentration in ZnG was positively correlated to GPx activity after the intervention (r = 0.61; p < 0.01). CONCLUSIONS: The postmenopausal women initially presented Zn deficiency, and the status of this mineral improved after the intervention. Zinc supplementation may be an effective approach for correcting the observed deficiencies, enhancing antioxidant defense in this risk population. CLINICAL TRIAL REGISTRATION: The present study is registered at the US National Institutes of Health (ClinicalTrials.gov), NCT03672513.

Folic acid and vitamin B12 as biomarkers of morbidity and mortality in patients with septic shock.

Introduction: Introduction and objective: a study was made of the folic acid (Fol) and vitamin B12 (B12) serum concentrations in critical patients with septic shock upon admission and after three days of stay in the Intensive Care Unit (ICU), with an analysis of their association to inflammatory parameters and patient morbidity-mortality. Methods: a prospective analytical study was made of 30 critically ill patients with septic shock. Demographic data, comorbidities, clinical information and severity scores were recorded. Data collected included serum Fol and B12 levels using the DxI® Autoanalyzer (Beckman Coulter) based on a competitive electrochemoluminescence immunoassay. Results: mean serum Fol was within the reference range stipulated by the laboratory on the first day. Nevertheless, a total of 21.4 % of the patients had high Fol levels, with 14.2 % being Fol deficient. An association was observed between Fol (p ˂ 0.012) status and 28-day mortality, and the number of days of mechanical ventilation, fraction of inspired oxygen (FiO2) and fibrinogen increased in patients with higher Fol levels (p ˂ 0.05). In addition, 85.7 % of cases had B12 levels above the reference values, with a correlation being observed between B12 and Fol. Conclusions: this study proposes Fol as a novel morbidity-mortality biomarker in critical septic patients, and reinforces the usefulness of B12 as a morbidity biomarker. It is thus suggested that the measurement of Fol upon admission and over the first 72 hours of hospital stay could provide prognostic information about the clinical course and outcome of septic shock patients.

Introducción: Introducción y objetivo: se realizó un estudio de las concentraciones séricas de ácido fólico (Fol) y vitamina B12 (B12) en pacientes críticos con shock séptico al ingreso y después de tres días de estancia en la Unidad de Cuidados Intensivos (UCI), con un análisis de su asociación con los parámetros inflamatorios y la morbimortalidad de los pacientes. Método: se realizó un estudio analítico prospectivo de 30 pacientes críticos con shock séptico. Se registraron datos demográficos, comorbilidades, información clínica y puntuaciones de gravedad. Los datos recopilados incluyeron los niveles séricos de Fol y B12 utilizando el autoanalizador DxI® (Beckman Coulter) basado en un inmunoensayo de electroquimioluminiscencia competitivo. Resultados: la media de Fol sérico estuvo dentro del rango de referencia estipulado por el laboratorio el primer día. Sin embargo, el 21,4 % de los pacientes presentaban niveles altos de Fol y el 14,2 % presentaban deficiencia de Fol. Se observó una asociación entre el estado de Fol (p ˂ 0,012) con la mortalidad a los 28 días, con el número de días de ventilación mecánica, con la fracción de oxígeno inspirado (FiO2) y con el fibrinógeno, que aumentaron en los pacientes con niveles de Fol más altos (p ˂ 0,05). Además, el 85,7 % de los casos tenían niveles de B12 por encima de los valores de referencia, observándose una correlación entre B12 y Fol. Conclusiones: este estudio propone al Fol como nuevo biomarcador de morbimortalidad en los pacientes críticos con sepsis y refuerza la utilidad de la B12 como biomarcador de morbilidad. Por tanto, se sugiere que la medición de Fol al ingreso y durante las primeras 72 horas de estancia hospitalaria podría proporcionar información pronóstica sobre el curso clínico y el resultado de los pacientes con shock séptico.

Bad Prognosis in Critical Ill Patients with COVID-19 during Short-Term ICU Stay regarding Vitamin D Levels.

BACKGROUND AND AIMS: Vitamin D inadequacy may be involved in the mechanisms of SARS-CoV-2 infection and in potential risk factors for disease propagation or control of coronavirus disease 2019 (COVID-19). This study assessed a short-term evolution of vitamin D status and its influence upon different clinical parameters in critically ill patients with COVID-19. METHODS: A prospective analytical study in which 37 critically ill volunteers between 41 and 71 years of age with COVID-19 were evaluated at baseline and three days of intensive care unit (ICU) stay. 25-OH-D3 and 25-OH-D2 were analyzed by liquid chromatography-tandem mass spectrometry and total 25-OH-D levels were calculated as the sum of both. RESULTS: All patients presented low 25-OH-D levels at baseline, decreasing total 25-OH-D (p = 0.011) mainly through 25-OH-D2 (p = 0.006) levels during ICU stay. 25-OH-D2 levels decreased a mean of 41.6% ± 89.6% versus 7.0% ± 23.4% for the 25-OH-D3 form during the ICU stay. Patients who did not need invasive mechanical ventilation presented higher levels of 25-OH-D2 at baseline and follow-up. Lower 25-OH-D and 25-OH-D3 levels were associated with higher D-dimer at baseline (p = 0.003; p = 0.001) and at follow up (p = 0.029), higher procalcitonin levels (p = 0.002; p = 0.018) at follow up, and lower percentage lymphocyte counts (p = 0.044; p = 0.040) during ICU stay. CONCLUSIONS: Deficient vitamin D status in critical patients was established at the admission and further worsened after three days of stay. Lower vitamin D levels were related to key altered clinical and biochemical parameters on patients with SARS-CoV-2 infection. Given the different response of the 25-OH-D3 and 25-OH-D2 forms, it would be useful to monitor them on the evolution of the critically ill patient.

Current trends in the analytical determination of vitamin D.

INTRODUCTION: Vitamin D is a micronutrient that plays a large role in bone disease, and researchers are now discovering that it also does so in non-skeletal disease, thus making high-quality analytical determination necessary. To make this determination, a series of immunochemical and physical methods are used. These methods present a series of different ways of handling samples as well as different methodologies that bring a series of advantages and limitations based on the scope of work in which the vitamin D analysis methodology is applied. Although the Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) is the gold standard method of analytical vitamin D determination, and is the only one to offer a more complete and accurate view of all metabolites of this vitamin, it is necessary to standardize all the analysis methodologies that allow accurate, reliable and quality analytical determination, since it is essential to obtain results that can reliably be extrapolated to the population, and that can be decisive in assessing a large number of pathologies.

INTRODUCCIÓN: La vitamina D es un micronutriente que ejerce un gran papel en enfermedades óseas y actualmente se está descubriendo que también lo hace en enfermedades no óseas, por lo que una determinación analítica de calidad es necesaria. Para realizar esta determinación se emplean una serie de métodos inmunoquímicos y físicos, los cuales van a presentar una serie de tratamientos diferentes de las muestras, así como diferentes metodologías que van a traer una serie de ventajas y limitaciones conforme al ámbito de trabajo en que se aplique la metodología de análisis de la vitamina D. A pesar de que la cromatografía líquida-espectrometría de masas en tándem (LC-MS/MS) es el método gold standard de determinación analítica de la vitamina D, y de que es el que ofrece una visión más completa y precisa de todos los metabolitos de esta vitamina, es necesaria una estandarización de todas las metodologías de análisis que permitan una determinación analítica precisa, fiable y de calidad, ya que es imprescindible obtener unos resultados que sean extrapolables con fiabilidad a la población y que puedan ser determinantes para valorar un gran número de patologías.

Pyroglutamic acidosis by glutathione regeneration blockage in critical patients with septic shock.

AIM: The aim of this study was to evaluate oxidative stress from glutathione depletion in critically ill patients with a septic shock through the abnormal presence of pyroglutamic acid (PyroGlu) in the urine (indirectly) and through its serum level (directly). METHODS: This was a prospective analytical study of 28 critically ill patients with a septic shock who were monitored from admission (initial) to 3 days of stay (final) in the intensive care unit (ICU). Data collected included PyroGlu and glutamic acid (Glu) using liquid chromatography/mass spectrometry, and glutathione peroxidase (GPX) activity with a colorimetric assay. The differences in Glu, PyroGlu, and GPX activity between the septic shock group and healthy control group serving as reference values were evaluated using the Mann-Whitney test. The correlations between Glu, PyroGlu, and GPX activity and clinical outcomes were determined using Spearman's correlation coefficient. RESULTS: In patients with septic shock, serum and urine PyroGlu levels were higher, erythrocyte GPX activity/gr Hb was lower, and urine Glu levels were lower compared to healthy control reference values, for both initial and final values. Initial serum Glu levels were also lower. Serum PyroGlu levels had a correlation with both initial and final serum Glu levels; levels also correlated in the urine. Initial serum Glu correlated with the days of mechanical ventilation (P = 0.016) and the days of ICU stay (P = 0.05). Urine Glu/mg creatinine correlated with APACHE II (P = 0.030). This positive correlation observed for serum Glu was not observed for PyroGlu. CONCLUSIONS: The current study found that septic patients have higher levels of PyroGlu, lower levels of Glu, and lower erythrocyte GPX activity, suggesting that these biomarkers could be used as an indicator of glutathione depletion. In addition, Glu is related to severity parameters. This study can guide future studies on the importance of monitoring the levels of pyroglutamic acidosis in critical patients with septic shock in order to preserve the oxidative status and its evolution during the stay in the ICU.

Effect of Melatonin Supplementation on Antioxidant Status and DNA Damage in High Intensity Trained Athletes.

The aim of the study was to evaluate the effect of melatonin supplementation on antioxidant capacity and DNA damage in high intensity interval training (HIIT) athletes. A 2-week randomised, double-blinded, placebo-controlled trial with two groups was conducted. Placebo (PG) and melatonin (MG) (20 mg/d) athletes were monitored over a two-week period of HIIT and strength training. The total antioxidant capacity (TAC) and the glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities were analysed in blood samples. DNA damage was measured in isolated lymphocytes by comet assay prior to and immediately after exercise. The supplementation increased plasma melatonin levels in the melatonin-treated group (p<0.05) after two weeks of intervention. Analysis of antioxidant status indicated higher (p<0.05) TAC and GPx in MG than PG post-intervention. No differences were found in SOD enzyme activity. DNA damage was diminished in MG (p<0.05) compared to PG in post-training conditions. Antioxidant status was associated with DNA damage (r=-0.679; p=0.047) in the melatonin-treated athletes. The present study suggest that melatonin supplementation improves antioxidant status and may prove to have beneficial effects preventing DNA damage induced by high intensity training.

Loss of Bone Mineral Density Associated with Age in Male Rats Fed on Sunflower Oil Is Avoided by Virgin Olive Oil Intake or Coenzyme Q Supplementation.

The role of dietary fat unsaturation and the supplementation of coenzyme Q have been evaluated in relation to bone health. Male Wistar rats were maintained for 6 or 24 months on two diets varying in the fat source, namely virgin olive oil, rich in monounsaturated fatty acids, or sunflower oil, rich in n-6 polyunsaturated fatty acids. Both dietary fats were supplemented or not with coenzyme Q10 (CoQ10). Bone mineral density (BMD) was evaluated in the femur. Serum levels of osteocalcin, osteopontin, receptor activator of nuclear factor κB ligand (RANKL), osteoprotegerin (OPG), adrenocorticotropin (ACTH) and parathyroid hormone (PTH), as well as urinary F2-isoprostanes were measured. Aged animals fed on virgin olive oil showed higher BMD than those fed on sunflower oil. In addition, CoQ10 prevented the age-related decline in BMD in animals fed on sunflower oil. Urinary F2-isoprostanes analysis showed that sunflower oil led to the highest oxidative status in old animals, which was avoided by supplementation with CoQ10. In conclusion, lifelong feeding on virgin olive oil or the supplementation of sunflower oil on CoQ10 prevented, at least in part mediated by a low oxidative stress status, the age-related decrease in BMD found in sunflower oil fed animals.

Age-Related Loss in Bone Mineral Density of Rats Fed Lifelong on a Fish Oil-Based Diet Is Avoided by Coenzyme Q10 Addition.

During aging, bone mass declines increasing osteoporosis and fracture risks. Oxidative stress has been related to this bone loss, making dietary compounds with antioxidant properties a promising weapon. Male Wistar rats were maintained for 6 or 24 months on diets with fish oil as unique fat source, supplemented or not with coenzyme Q10 (CoQ10), to evaluate the potential of adding this molecule to the n-3 polyunsaturated fatty acid (n-3 PUFA)-based diet for bone mineral density (BMD) preservation. BMD was evaluated in the femur. Serum osteocalcin, osteopontin, receptor activator of nuclear factor-κB ligand, ostroprotegerin, parathyroid hormone, urinary F2-isoprostanes, and lymphocytes DNA strand breaks were also measured. BMD was lower in aged rats fed a diet without CoQ10 respect than their younger counterparts, whereas older animals receiving CoQ10 showed the highest BMD. F2-isoprostanes and DNA strand breaks showed that oxidative stress was higher during aging. Supplementation with CoQ10 prevented oxidative damage to lipid and DNA, in young and old animals, respectively. Reduced oxidative stress associated to CoQ10 supplementation of this n-3 PUFA-rich diet might explain the higher BMD found in aged rats in this group of animals.

Implementation of a nutrition education program in a handball team; consequences on nutritional status.

OBJECTIVE: To evaluate nutritional status and dietary habits after implementation of a nutritional education program in professional handball players. RESEARCH METHODS AND PROCEDURES: Longitudinal study of 14 handball players evaluated with 72-h recall, a questionnaire on food consumption and anthropometric measures during 4 months. The intervention consisted of a nutrition education program. RESULTS: Energy intake was consistently below the recommended allowances. Macronutrient intakes as a percentage of total energy intake were below the recommended allowances for carbohydrates, and above recommended allowances for fats. Nutritional education was followed by a significant increase (p < 0.01) in total energy and macronutrient intakes, with no significant changes in macronutrient or micronutrient intakes after adjustment for energy intake. DISCUSSION: The imbalance in nutrient intake in handball players suggests that detailed re-analysis is needed to determine specific recommendations for this population. Nutritional education with continuous follow-up to monitor athletes' dietary habits may lead them to adopt appropriate nutritional habits to optimize dietary intakes. The lack of specific recommendations for micronutrient intakes in athletes leads to confusion regarding appropriate intakes; biochemical tests that yield normal values (albeit approaching cut-off values for deficiency) may disguise deficient status for some nutrients when strenuous exercise is involved. CONCLUSION: In-depth studies with nutrition education programs that include long-term follow-up are advisable to avoid deficiencies that can lead to irreversible damage in competitive athletes.

Objetivos: Evaluar el estado nutricional y los hábitos dietéticos en respuesta a la aplicación de un programa de educación nutricional en jugadores profesionales de balonmano. Sujetos y metodología: Estudio longitudinal realizado a una muestra de 14 jugadores pertenecientes a un equipo de balonmano de alto rendimiento, a los que se les evaluó mediante recordatorio de 72 horas, un cuestionario de frecuencia de consumo, medidas antropométricas a lo largo de 4 meses, y a los que se les aplicó un programa de educación nutricional al inicio del estudio. Los valores de ingesta y de frecuencia de consumo fueron comparados con las recomendaciones de macronutrientes existentes para deportistas y micronutrientes para población sana, respectivamente, y con la pirámide de alimentos para población sana española. Resultados: La ingesta de energía de los deportistas se situó por debajo de las recomendaciones a lo largo de todo el estudio. La ingesta de macronutrientes respecto a la energía ingerida, se situó por debajo de las recomendaciones para la ingesta de carbohidratos y por encima de las recomendaciones para la ingesta de grasa, mostrada en los resultados obtenidos de frecuencia de consumo de alimentos. La educación nutricional produjo un incremento significativo (p < 0,01) en la ingesta de energía y macronutrientes tras su aplicación. A pesar de ello, no se produjeron cambios significativos en la ingesta de macronutrientes y micronutrientes al ajustar por energía ingerida. Los niveles bioquímicos se encontraron dentro de los rangos de normalidad durante todo el estudio. Discusión: El desequilibrio en la ingesta de nutrientes presente en los jugadores de balonmano hace necesario realizar un ajuste nutricional completo para poder establecer recomendaciones específicas para este tipo de población. La aplicación de un programa de educación nutricional monitorizada de manera continuada mediante seguimiento en los deportistas, puede tener como consecuencia la instauración de hábitos nutricionales adecuados que lleve a una optimización en la ingesta. La ausencia de recomendaciones específicas de micronutrientes en el deporte, provoca una cierta confusión a la hora de establecer una ingesta adecuada de micronutrientes, ya que en muchos casos demuestran normalidad en los niveles bioquímicos, aunque muy cercanos a la deficiencia, pudiendo comprometer el estatus de algún nutriente en situaciones de ejercicio extremo. Conclusión: Sería aconsejable realizar estudios exhaustivos de valoración del estatus nutricional que planteen la instauración de programas de educación nutricional a largo plazo, con el fin de evitar carencias que deriven en daños irreversibles en el deportista de competición.

Characterization of L-cysteine capped CdTe quantum dots and application to test Cu(II) deficiency in biological samples from critically ill patients.

The catalytic activity of copper ion gives, from the physiological point of view, a central role in many biological processes. Variations in the composition and location of cellular copper have been addressed given their physiological and pathological consequences. In this paper L-cysteine capped CdTe quantum dots is used for the fluorimetric determination of Cu(II) in biological samples from healthy individuals and patients admitted to the Intensive Care Units (ICU). An acceptable homogeneity in the CdTe QDs size has been obtained with an average value of 3 nm. No significant alterations in the spectral properties were observed for 2 months when stored in vacutainers at 6°C and a concentration of approximately 2 µM. Data from oxidative stress markers such superoxide dismutase, total antioxidant capacity and DNA damage can be correlated with a Cu(II) deficiency for the ICU patients as measured by flame-atomic absorption spectroscopy (FAAS) and inductively coupled plasma source mass spectrometry (ICP-MS). Aqueous solutions 0.3 µM of L-cysteine capped CdTe QDs in MOPS buffer (6 mM, pH 7.4) used at 21°C in the range 15-60 min after preparation of the sample for the measurements of fluorescence gives contents in Cu(II) for erythrocytes in good agreement with those obtained in FAAS and ICP-MS but the comparative ease of use makes the fluorimetric technique more suitable than the other two techniques for routine analysis.

Assessment of iron and zinc intake and related biochemical parameters in an adult Mediterranean population from southern Spain: influence of lifestyle factors.

This study assessed factors influencing iron and zinc intake and associated biochemical parameters in an adult population from southern Spain to identify patterns of intake and groups at risk for deficiency. A cross-sectional survey was done in Andalusia (southern Spain) to study nutrient intakes in a random sample of 3421 subjects (1747 men, 1674 women, age between 25 and 60 years). Blood samples were obtained for biochemical assays in a random subsample of 354 subjects (170 men, 184 women). Food consumption was assessed by 48-h recall. In blood samples, we measured red blood cells, haemoglobin (Hb), haematocrit, total iron binding capacity (TIBC) and plasma concentration of Fe and Zn. Information about educational level, smoking habit, alcohol consumption and physical exercise was collected with a structured questionnaire. Intakes were below two thirds of the recommended dietary allowances (RDA) in 22.45% of the sample for Fe and in 56.45% for Zn. Iron deficiency [two or more abnormal values for plasma Fe, TIBC, transferrin saturation and mean cell volume (MCV)] was found in 12.7% of the sample, and iron-deficient anaemia (low values for Hb, MCV, mean cell Hb and mean corpuscular Hb concentration) was found in 2.1%. In smokers, plasma levels of Fe were higher, and MCV was lower than in nonsmokers. Plasma Zn concentrations were below the reference value in 17.8% of the persons. Age and body mass index correlated inversely with plasma Zn (P<.01). Gender, age, obesity, smoking, alcohol consumption and physical activity were associated with differences in nutrient intakes. Logistic regression analysis showed that female gender and older age were associated with the risk of low intakes of Fe and Zn.

Factors influencing the intake and plasma levels of calcium, phosphorus and magnesium in southern Spain.

OBJECTIVE: The aim of the study was to assess factors influencing the intake and plasma levels of calcium (Ca), phosphorus (P) and magnesium (Mg) in an adult population from southern Spain, in order to identify patterns of intake and groups at risk for deficiency. METHODS: A cross-sectional survey was carried out in Andalusia, a western Mediterranean region in southern Spain. Nutrient intakes were studied in a random sample of 3,421 subjects (1,747 men, 1,674 women) between 25 and 60 years of age. Blood samples were obtained for biochemical assays in a random subsample of 354 subjects (170 men, 184 women). Food consumption was assessed by a 48-h recall. Concentrations of Ca, P and Mg were measured in plasma. Information about level of education, smoking habit, alcohol consumption and physical exercise was collected with a structured questionnaire. RESULTS: Intakes were below two-thirds of the RDA in 39.52% of the sample for Ca, and in 32.31% for Mg. Gender, age, educational level, obesity, smoking, alcohol use and physical activity were associated with differences in nutrient intakes. Plasma concentrations were below the reference value in 14.81% of the sample for Ca, and in 8.82% for Mg. Obese persons [body mass index (BMI) >/= 30 kg/m(2)] consumed less Ca, P and Mg, and had lower plasma Ca concentrations (P < 0.05) than non-obese persons. BMI showed an inverse correlation with plasma concentrations of Ca (r = -0.21, P < 0.05). CONCLUSION: Our results provide an estimate of the intake of Ca, P and Mg in the adult population of southern Spain. Of the factors that affected the intake of these nutrients, logistic regression analysis showed that only female gender and older age were associated with the risk of low plasma Ca concentrations.

Tumor-promoting activity of p-hydroxybenzenediazonium is accelerated in Mg-deficient rats.

Tumor-promoting activity caused by the short-term administration of p-hydroxybenzenediazonium (PDQ) has been assayed in rats fed on a Mg-deficient diet as a reference model versus rats fed on a standard diet as controls. For 5 weeks groups of 20 rats, fed either on the Mg-deficient or standard diet, were treated simultaneously with PDQ. A group of 10 Mg-deficient rats remained untreated. Topical application of PDQ was followed by the appearance of macroscopic alterations in the skin, which were more evident in the Mg-deficient rats. No deaths occurred during the treatment. After 5 weeks' PDQ treatment the rats were killed and histological analyses were made. Tissues from the skin, liver, heart, kidney, lung and thymus were screened by conventional staining methods. Both the PDQ-treated Mg-deficient and PDQ-treated control rats presented tissue lesions, although to a different extent. The untreated Mg-deficient rats showed no such lesions. Mg-deficient rats treated with PDQ developed significant incipient fibrosarcomas (p<0.05) and extended hyperplasia (p<0.001), particularly in the skin, accompanied by a significant increase in the thickness of collagen (mean value: 445.4+/-47.2microm, p<0.05) compared to the control PDQ-treated group (mean values: 258.7+/-36.4microm). The overall results provide objective proof of tumor-promoting activity after 5 weeks' treatment with PDQ. Such a fast response is interpreted as being linked to the increased vulnerability of the membrane caused by Mg deficiency, which would more readily facilitate the toxic activity of p-hydroxybenzenediazonium ions.