ALA Dehydrase Research Articles

5-Aminolevulinic acid promotes anthocyanin accumulation in Fuji apples

5-Aminolevulinic acid (ALA) is an essential precursor of all tetrapyrrole compounds such as chlorophylls and heme in plants. It has also been suggested widely for applications to crops to enhance growth and production as a plant growth regulator. However, how successful ALA can be used in fruit production was rarely reported. We conducted a field experiment at eight locations in four provinces across eastern China; and the results showed that application of ALA solutions to ‘Fuji’ apple (Malus × domestica Borkh.) fruits 20 days prior to harvest significantly increased the anthocyanin content in the fruit skin. Also, ALA treatment increased the anthocyanin content of the detached apple skin in a growth chamber. Results from the semi-quantitive RT-PCR analysis showed that ALA induced gene expressions related to anthocyanin biosynthesis, including the structural genes Pal, Chs and Ufgt, and regulatory genes Myb, bHLH and Wd40. When levulinic acid (LA), an inhibitor of ALA dehydrase, was added, ALA promotion of anthocyanin accumulation and up-regulation of gene expressions were inhibited. Taken together, these results suggest that ALA promotion of anthocyanin accumulation in apples was facilitated by the up-regulation of gene expression, which might be related to the conversion of ALA to porphyrins.

Promotion by 5-Aminolevulinic Acid of Germination of Pakchoi (Brassica campestris ssp. chinensis var. communis Tsen et Lee) Seeds Under Salt Stress

: The seed germination and seedling growth of pakchoi (Brassica campestris ssp. chinensis var. communis Tsen et Lee cv. Hanxiao) were not significantly inhibited until the concentration of NaCl was increased to150 mmol/L. Treatment of pakchoi seeds with exogenous 5-aminolevulinic acid (ALA), at concentrations ranging from 0.01 to 10.00 mg/L, promoted seed germination when seeds were stressed by salinity, whereas levulinic acid (LA), an inhibitor of ALA dehydrase, significantly inhibited seed germination and seedling growth, suggesting that metabolism of ALA into porphyrin compounds was necessary for seed germination and seedling growth. Determination of respiratory rate during seed germination showed that ALA increased seed respiration under both normal conditions and salt stress. Furthermore, salt stress decreased levels of endogenous ALA, as well as heme, in etiolated seedlings. More salt-tolerant cultivars of pakchoi contained higher relative levels of endogenous ALA and heme under conditions of salt stress. These results indicate that salt stress may inhibit the biosynthesis of endogenous ALA and then heme, which is necessary for seed germination, and treatment of seeds with exogenous ALA prior to germination may be associated with the biosynthesis of heme. (Managing editor: Ping HE)

Porphyrias: Clinical Evaluation and Interpretation of Laboratory Tests

Porphyria is a hereditary enzyme disorder that affects the heme biosynthetic pathway and results in excess accumulation and excretion of porphyrin or porphyrin precursors (Fig. 1). Heme is a functional component of hemoglobin and heme-requiring enzymes, such as the cytochrome P-450 system. The structural constituents of heme are protoporphyrin IX (a tetrapyrrole ring) and iron. Heme production is especially active in developing erythrocytes and in hepatic cells. Clinical Spectrum of Porphyrias.—An enzyme defect in any of the heme biosynthetic steps (Fig. 1) causes excess accumulation of the related substrate porphyrinogen or porphyrinogen-precursor molecule. Disease manifestation depends on the type of the excess porphyrinogen intermediate and related porphyrin. If the excess is of the early precursor molecules (9-aminolevulinic acid [ALA], porphobilinogen [PBG], or both), the manifestations are neuropsychiatrie. These consist of recurrent acute attacks of autonomie dysfunction expressed as abdominal pain, vomiting, constipation, tachycardia, and hypertension in association with psychiatric symptoms, fever, leukocytosis, proximal paresis, paresthesias, and occasionally the syndrome of inappropriate antidiuretic hormone. If the excess is of the distal intermediates (uroporphyrins, coproporphyrins, and protoporphyrins), the manifestations are cutaneous and consist of photosensitivity, blister formation, facial hypertrichosis, and hyperpigmentation. Finally, if the excess is of both the early and the distal porphyrin intermediates, the manifestations are both neuropsychiatrie and cutaneous. Accordingly, the porphyrias are clinically classified by their potential to involve the nervous system or the skin (or both). Three groups have been identified: neural, neurocutaneous, and cutaneous porphyrias. The first two are also known as acute hepatic porphyrias, an indication of the principal site of overproduction of porphyrin. All porphyrias are inherited as autosomal dominant traits except plumboporphyria (PP) and congenital erythropoietic porphyria (CEP), which are inherited as autosomal recessive traits. Neuroporphyrias.—In acute intermittent porphyria (AIP) and PP, the defective enzymes are PBG deaminase and ALA dehydrase, respectively; thus, PBG or ALA (or both) accumulates. Induction of hemoprotein synthesis for the production of catalase and cytochromes, and induction of ALA synthase by fasting, drugs, hormones, or alcohol may result in precipitation of acute attacks. Accordingly, the best treatment is avoidance of these factors. During attacks, a loading dose of carbohydrates may help because glucose prevents induction of ALA synthase by both exogenous and endogenous factors. Two recent randomized studies failed to demonstrate the clinical value of hematin or tin protoporphyrin therapy, even though biochemical remissions were verified.1·2

Hematin therapy for the neurologic crisis of tyrosinemia

Hereditary tyrosinemia (tyrosinemia type I) is an autosomal recessive disorder caused by a defect of the final enzyme in tyrosine metabolism. The disease is associated with renal tubular defects, acute and chronic liver failure, and hepatocellular carcinoma. An elevation in serum 6-aminolevulinic acid concentration is common and is due to competitive inhibition of ALA dehydrase by succinylacetone, a metabolite of tyrosine degradation. 1 A syndrome of acute neurologic crises, which has striking similarity to the acute porphyrias, is seen in as many as 40% of the children with this disorder. 2 As in the acute porphyrias, the episodes of acute neuropathy associated with tyrosinemia are best correlated with ALA excretion. 2 Hematin suppresses ALA synthase, the first committed step in heme synthesis. 3 Clinical studies in patients with porphyria have established the efficacy of hematin in decreasing ALA excretion and the improvement of clinical symptoms after intravenous administration. 4 The drug has few side effects and has been used without major complications in the treatment of numerous patients with acute porphyria. 4 We describe a case of severe neurologic crises associated with extremely high serum ALA levels in a child with tyrosinemia I. We elected to treat this patient with hematin, followed by liver transplantation. We believe this to be a rational course given the marked similarities of this disorder to the acute porphyrias.

Assignment of the human gene for delta aminolevulinate dehydrase to chromosome 9 by somatic cell hybridization and specific enzyme immunoassay.

A non-competitive enzyme immunoassay specific for delta aminolevulinate dehydrase has been devised and applied to rodent-human hybrid cell lines. Two different conditions have been used, one specific for the human enzyme and the other indicative of both rodent and human enzymes. The ratio of the values obtained under the two conditions was used to discriminate between positive and negative clones. By this method the gene for ALA dehydrase has been assigned to chromosome 9.

Iron and the liver: acute effects of iron-loading on hepatic heme synthesis of rats. Role of decreased activity of 5-aminolevulinate dehydrase.

Acute iron loading of rats, by intraperitoneal administration of iron-dextran (500 mg Fe/kg body wt 18-20 h before killing) decreased by 30% the rate of conversion of 5-amino-[14C]levulinate ([14C]ALA) into heme as measured with a recently described procedure for liver homogenates (1981. Biochem. J. 198: 595-604). The decrease in conversion of labeled ALA into heme caused by iron loading was shown to be due to a 70-80% decrease in activity of ALA dehydrase. The decrease in activity of ALA dehydrase caused by iron loading was not associated with a decrease in hepatic concentrations of GSH, nor could it be reversed by addition of dithiothreitol, Zn2+ or chelators of Fe2+ and Fe3+. Addition of FeSO4, ferric citrate, or ferritin to homogenates of control liver had no effect of activity of ALA dehydrase. The decrease in activity of ALA dehydrase, caused by iron-dextran, was mirrored by a reciprocal increase in ALA synthase. Iron-dextran potentiated the induction of ALA synthase by allylisopropylacetamide. However, this potentiation could be dissociated from the decrease in ALA dehydrase caused by iron loading.

Erythroid colony development as a function of age: the role of marrow cellular heme.

The activities of the heme biosynthetic enzymes ALA synthase (ALAS) and ALA dehydrase (ALAD) and the heme degradative enzyme heme oxygenase were analyzed from bone marrow cells obtained from young, middle-aged, and senescent rats. There was age-related reduced activity of bone marrow ALAS but no age-related difference in the activity of ALAD. In contrast, heme oxygenase activity was 50% greater in the senescent marrow cells. Incorporation of 14C-glycine into heme was 45% less in senescent rat marrow cells, whereas incorporation of 14C-delta-aminolevulinic acid was not related to age. Senescent bone marrow cells demonstrated a marked reduction in 14C-leucine and 3H-uridine incorporated into protein and nucleic acid synthesis, respectively. In vitro erythroid colony (CFUE) growth by senescent bone marrow cells was as much as 40% less compared with young bone marrow cells. The decreased ability to form CFUE by the senescent bone marrow cells may be related to reduced ALAS activity and increased heme oxygenase activity. Thus, part of the aging process appears to involve fluctuations in the enzyme activities and protein synthesis involved with metabolism of heme.

Immunosuppressive activity of succinylacetone.

SA, an inhibitor of ALA dehydrase, the second enzyme of the heme biosynthetic pathway, has been shown to exert immunosuppressive activity in several systems. When the Walker 256 tumor was grown subcutaneously in outbred Sprague-Dawley rats, SA prevented rejection of the tumor by the host. Human peripheral blood lymphocyte transformation in response to mitogens and antigens in vitro was markedly impaired by addition of SA to the medium. Addition of hematin to the medium did not reverse the SA-mediated inhibition of transformation. This finding suggests that SA may exert immunosuppressive activity by a mechanism separate from inhibition of heme biosynthesis. Continuous administration of SA to Fisher 344 rats profoundly suppressed hemolytic antibody production after immunization with SRBC. Administration of large doses (equivalent to the highest dose of Sa given to tumor-bearing animals) for a month produced a 20% decrease of hemoglobin concentration, a 12% decrease in hematocrit, and no significant effect on leukocyte or erythrocyte concentration. There were no significant changes in tissue histology, including marrow cellularity.

内毒素投与のマウスの肝，ひδ‐アミノレブリン酸（ＡＬＡ）シンテターゼ，ＡＬＡデヒドラーゼ活性に及ぼす影響

内毒素投与によるマウス肝,脾臓のヘム合成に関与するδ-aminolevulinate (ALA) synthetase, ALA dehydrase (ALAD)活性の変化を測定した。肝ALA synthetase, ALAD活性は6∼16時間後に著明な減少が見られるが以後回復した。脾臓ではALA synthetase, ALAD活性は6∼16時間後に活性の減少を見るが,以後回復し,さらに活性上昇を示し,約2∼3週の後に正常レベルに復帰した。また血中のALADは内毒素量に応じて減少を示し,のち回復し,3日後からやや増大の傾向を示した。血清鉄は対照に比較して6∼16時間で著しく減少を示し,のち漸次回復の傾向を示した。脾臓での赤芽球の増量はFruhmanの観察と一致した。一方肝薬物代謝系酵素のp-450系の活性を示すマウスのpentobarbital睡眠時間は内毒素の投与により延長した。

A rapid and selective action of 6-benzylaminopurine on 5-aminolevulinate production in excised sunflower cotyledons

When treated with exogenous 6-benzylaminopurine (BAP) cotyledons excised from 6-day-old etiolated sunflower seedlings ( Helianthus annuus L., cv. Tall single) exhibit stimulated chlorophyll (Chl) accumulation on subsequent illumination with white light. The influence of BAP on various enzymes of Chl biosynthesis was investigated. Cotyledons treated with BAP for 1.5 h before illumination show a greatly enhanced capacity for 5-aminolevulinate (ALA) production. The activities of ALA dehydrase and the uroporphyrinogen synthetase/uroporphyrinogen III cosynthetase complex in light are unaffected by a 24 h dark pretreatment with BAP. Under the conditions investigated, BAP-stimulated ALA production is reflected in a similar increase in Chl accumulation. Thus BAP stimulated Chl accumulation appears to be mediated through a rapid and selective enhancement of the capacity for ALA production.

5-aminolevulinic acid stimulation of porphyrin and hemoglobin synthesis by uninduced friend erythroleukemic cells

Porphyrin synthesis and iron accumulation was stimulated by exogenous 5-aminolevulinic acid (ALA) in uninduced Friend erythroleukemic cells (FELC). Uroporphyrin and protoporphyrin were the major intermediate precursors produced. All porphyrin types were conjugated to protein insoluble cellular components and could be extracted only by methanol sulfuric acid esterification. Heme content of the uninduced FELC was increased 6-fold in the presence of 5 × 10 −4 M ALA. As a consequence, the synthesis of the minor murine hemoglobin component was preferentially induced, an effect similar to that expressed by exogenous hemin. Addition of exogenous ALA to 0.5% DMSO-induced cells increased total hemoglobin synthesis with a higher efficiency of the minor hemoglobin. The endogenous synthesis of porphyrin from exogenous ALA was markedly reduced by hemin. Uroporphyrin, coproporphyrin, protoporphyrin and heme were equally repressed, indicating an inhibitory effect of hemin on ALA dehydrase and urosynthetase activities. In addition, hemin repressed [ 3H]leucine incorporation into protein by uninduced cells. Incubation of uninduced cells in culture medium without serum in the presence of hemin blocked their protein synthesis activity, whereas addition of serum exerted a protective effect on living FELC.

Inhibition of δ-aminolevulinic acid synthetase and δ-aminolevulinic acid dehydrase by L-2-amino-4-methoxy- [formula omitted]-3-butenoic acid in the rat

The rate limiting enzyme of heme biosynthesis, δ-aminolevulinic acid synthetase (ALA synthetase), and the second enzyme in the heme biosynthetic pathway, δ-aminolevulinic acid dehydrase (ALA dehydrase), were inhibited by the olefinic amino acid L-2-amino-4-methoxy - trans -3-butenoic acid (AMTB). Administration of AMTB (20 mg/kg; i.p.) to rats inhibited ALA synthetase and ALA dehydrase in control animals and in animals with markedly elevated activity of ALA synthetase which resulted from the administration of 3,5-dicarbethoxy-1,4-dimethyl-collidine (DDC, 200 mg/kg, i.p.) or allylisopropylacetamide (200 mg/kg, s.c.). AMTB also blocked the synthesis of rat hepatic porphyrins and inhibited the increase in the urinary excretion of δ-aminolevulinic acid and porphobilinogen following DDC (150 mg/kg, p.o.) administration. Preincubation of AMTB with liver mitochondria or a soluble fraction of liver decreased the activity of mitochondrial ALA synthetase and soluble ALA dehydrase, respectively.

582 Δ-AMINOLEVULINIC ACID (ALA) SYNTHETASE DEFECT IN A FEMALE WITH CONGENITAL SIDEROBLASTIC ANEMIA

Since birth a 3-year old black girl has had severe refractory hypochromic microcytic anemia. Her usual red blood cell (RBC) values are Hgb 6.0 gm/dl, Hct 20%, Retic. 0.5%, MCV 52 fl, MCH 15 pg. Iron deficiency and clinically significant thalassemia were excluded by: lack of response to oral or parenteral iron, serum Fe/TIBC 231/276 μg/dl and serum ferritin 407 ng/ml; normal hemoglobin electrophoresis, absence of erythroid inclusions in the peripheral blood and bone marrow (BM), α/β globin synthetic ratio of 0.85 in peripheral blood reticulocytes and negative family studies. Intensive erythroid hyperplasia and numerous ring sideroblasts were present in the BM aspirate, and electron microscopy confirmed intramitochondrial deposits of iron. Free RBC protoporphyrin, urine porphyrins and porphyrin precursors, and BM ferrochelatase activity were normal. RBC ALA dehydrase, uroporphyrinogen synthetase and pyridoxal kinase activities were increased. Activity of BM ALA synthetase was markedly reduced to 5.5 pmoles ALA/106 erythroblasts/30 min (normal 127±29) but was enhanced 5-fold by pyridoxal phosphate (normal 0-25% increase). Therapy with oral pyridoxine has thus are not noticeably increased effective RBC production. The sideroblastic anemia in this patient appears to be related to an inherited defect in the initial step of heme biosynthesis.

The biosynthesis of δ-aminolaevulinic acid in plants

In animal and bacterial cells the first enzymic step unique to the tetrapyrrole biosynthetic pathway is the condensation of succinyl-CoA and glycine to yield δ-aminolaevulinic acid (ALA). The enzyme catalysing this reaction could not be detected in extracts from higher plants, or green or blue-green algae. Through the use of laevulinic acid, which competitively inhibits ALA dehydrase, and causes the accumulation of ALA in vivo , the ability of a number of specifically labelled 14 C radioactive compounds to label this ALA has been studied. Glycine and succinate are poor label donors, whereas α-ketoglutarate, glutamate and glutamine are able to donate 14 C to the ALA. Chemical degradation of the [ 14 C]ALA indicates that C 5 arises from C 1 of glutamate and the remaining four carbon atoms of the ALA arise from the remaining four carbon atoms of glutamate. This labelling pattern is incompatible with the succinyl-CoA-glycine condensation reaction and indicates a new pathway for ALA biosynthesis from the intact carbon skeleton of glutamate in greening plant tissues.

航空機用燃料タンク清掃作業において発生した四エチル鉛中毒

Two cases of tetraethyl lead (TEL) poisoning are described. Both subjects had been exposed to TEL in the process of scaling using high pressure water stream during the cleaning work inside the aviation fuel tank. The aviation fuel contains TEL in a concentration of 1. 12 g lead per liter. The affected men failed to wear respirators during the cleaning work because the explosimeter indicated a negative reading for petrol. After one hour of tank cleaning work they suffered from lacrimation, running rhinorrhea and vomiting.Case 1. A 54-year-old man was admitted to a general hospital 3 days after the exposure to TEL and complained of hand tremors, amnesia and disorientation. He was restless, violent and confused in the night. On 12th day after the exposure to TEL, the condition bacame worse with marked agitation, delirium, convulsion, fever and coma. He died on 18th day after the exposure to TEL. During the admission, urinary coproporphyrin and basophilic stippling cells were normal ; no blood and urinary lead determination were done.Case 2. A 48-year-old man, on 2nd day after the exposure to TEL complained of chills, tremors, marked nausea and vomiting which persisted all night. Next morning he was admitted to another hospital. He had generalized tremors, ataxia, disorientation and at night he was suspicious, restless and violent. On 9th day after the exposure to TEL, his insomnia and restlessness gradually improved and he was discharged two months later.In this case, urinary lead determinations were done serially from 20 days to 196 days after the accident and blood lead determination was done once a week. On 20th day after the exposure, blood lead level was 52.3μg/100g, urinary lead concentration 586 μg/l and erythrocyte ALA dehydrase (ALA-D) activity was markedly reduced to 0.11μ mole PBG/ml RBC/hr. On 196th day after the exposure to TEL, his condition was both physically and mentally normal but his blood lead level was slightly elevated to 26. 1 μg/100 g and the urinary lead concentration was still at 37.0μg/l (81μg/24hr). Blood triethyl lead levels were found to be 5.8μg Pb/100g after 56 days, steadily decreasing thereafter to 1.3μg Pb/100 g up to 196 days.In this case, the reactivation of erythrocyte ALA-D and the fall of blood lead levels occurred simultanously in a manner similar to that observed in men exposed to inorganic lead. The regression line for erythrocyte logarithmic ALA-D activities and blood lead levels in this case is identical to that obtained from workers exposed to inorganic lead and the control group occupationally unexposed. These results suggest that the reduced erythrocyte ALA-D activities found in the TEL poisoning was due to inorganic lead resulting from the decomposition of TEL.Workmen handling antiknock additives were investigated regarding potential hazardous effects of tetraalkyl lead (TAL). The subjects consisted of workmen who engaged in mixing TAL into petrol, transportation of TAL by trucks or barges and storage tank cleaing. There were no abnormal values of blood lead levels, erythrocyte ALA-D activities or excreted urinary lead in those workmen.

Biological response and subjective symptoms in low level lead exposure.

Workers occupationally exposed to lead in the air of up to 60μg/cu m were examined for blood lead (Pb-B), urinary δ-aminolevulinic acid (ALA), erythrocyte ALA dehydrase (ALAD), hemoglobin, hematocrit, red blood cell count, and subjective symptoms. The Pb-B and ALAD showed consistent increases and decreases, respectively, corresponding with the levels of Pb exposure, while other tests revealed no association with Pb. The range of Pb exposure as expressed by Pb-B was up to 50μg/100 gm. Subjective symptoms and urinary ALA are not likely to be increased at the Pb-B level of about 50μg/100 gm and less.

Measurement of delta-aminolevulinic acid synthetase activity in human erythroblasts.

A new, specific, and simple method for the determination of delta-aminolevulinic acid (ALA) synthetase activity in human bone marrow cells has been developed. ALA synthetase of erythroblasts was partially purified so as to permit the use of [(14)C]succinyl-CoA as a substrate for this enzyme. In this enzyme preparation there were negligible activities of succinyl-CoA hydrolase, alpha-ketoglutarate dehydrogenase, and succinyl-CoA synthetase and there was no activity of ALA dehydrase. The ALA formed from [(14)C]succinyl-CoA has been isolated by column chromatography. Radioactivity in the eluate from the column has been proved by paper chromatography to be exclusively that of [(14)C]ALA. The entire assay can be completed within 4 h, and [(14)C]succinyl-CoA was incorporated into [(14)C]ALA on the order of several percent. Moderate to marked decreases of ALA synthetase activity have been demonstrated in the erythroblasts of all cases of sideroblastic anemia. In the cases of iron deficiency anemia, on the other hand, normal or slightly elevated activity has been obtained.

Heme synthetase activity of human bone marrow cells--examination of 105 cases of various diseases.

Elucidation of the changes in the process of heme synthesis is considered to be most important for the understanding of the mechanism of anemia in various diseases. For this reason, the activiies of ALA synthetase and ALA dehydrase have been studied by many authors. In the present study, the activity of heme synthetase, an iron-protoporphyrin chelating enzyme which is another important enzyme in heme synthesis, in the bone marrow cells was investigated in various diseases. The paste of nuclei-mitochondria obtained from lysates of bone marrow cells was used as the enzyme source. A considerable increase in the enzyme activity was seen in iron deficiency anemia and in cancer without cachexia. No significant difference from the controls was seen in the hepatic and renal diseases. A decreased activity was seen in some of panmyelo-pathy, cancer with cachexia and hyperthyroidism. Based on these findings, the factors regulating the enzyme activity and their relationship to the development of anemia were discussed.

THE DIAGNOSIS OF DIFFERENT DEGREES OF LEAD ABSORPTION; IN SPECIAL REFERENCES TO CHOICE AND EVALUATION OF VARIOUS PARAMETERS INDICATIVE OF AN INCREASED LEAD ABSORPTION

A series of 44 workers occupationally exposed to lead and 79 habitants naturally exposed to lead was investigated in order to define more clearly the clinical and biochemical criteria of lead exposure after grouping them into five according to the degree of lead exposure and also to the clinical evidence.The parameters used were hemoglobin (Hb), blood lead (Pb-B), urinary lead (Pb- U), urinary coproporphyrin (Copro), urinary δ-aminolevulinic acid (ALA), and ALA- dehydrase (ALA-D) activity in peripheral red blood cells (RBC) estimations.A statistically significant difference was found between mean values respectively for Pb-B and ALA-D of all five groups and also for Pb-U, ALA, Copro and Hb of three occupationally exposed groups. Furthermore, Pb-B correlated negatively with ALA-D in all five groups, and positively with Pb-U and ALA in three occupational groups. Negative correlations were also observed between ALA-D and each of Pb-U and ALA in three occupational groups.As for naturally exposed groups, a significant difference was found between the urban habitants and the rural habitants in regard to the values for Pb-B and ALA- D and, moreover, a close negative correlation was observed between Pb-B and ALA- D at these levels of Pb-B.Increased ALA excretions were observed when ALA-D activity was decreased to approximately 40% of normal levels.As for choice of a parameter, Pb-B and ALA were concluded to be of value for assessment of a heavy exposure and also for the clinical assessment, ALA-D for a minimum to slight exposure, and all the parameters including Hb and Copro for the comparison of the different lead exposure as a group.Pb-B above 65 μg/100 g and ALA above 17 mg/1 were almost always associated with symptoms and signs of lead poisoning and were therefore considered to be the upper limits of safe ranges. These levels corresponded to ALA-D activity of 0.250 μmole PBG/1 hr/1 ml RBC.

Effect of lead on blood regeneration following acute hemorrhage in dogs.

Thirty-six beagle dogs were divided into three hematologically similar groups of six males and six females each. The dogs of one group acted as controls and were fed only the basic diet. The dogs of the other two groups had lead acetate added to their diets, those of one group receiving 100 ppm of lead by weight, those of the other group, 500 ppm for 30 weeks followed by 1000 ppm for 16 weeks. By the end of 46 weeks, both lead-treated groups had accumulated burdens of lead as demonstrated by increases in blood and urine lead concentrations, increased urine δ-aminolevulinic acid excretion, and decreased red cell ALA dehydrase activity. At this time a two-step withdrawal of one-half the estimated blood volume from each dog resulted in a 30 to 40% reduction in hemoglobin concentration, red cell count, and hematocrit ratio. The recoveiy curves of the hemoglobin concentration, red cell count, and hematocrit ratio were not affected by the presence of the lead.