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PubMed2004

Structural basis of citrate-dependent and heparan sulfate-mediated cell surface retention of cobra cardiotoxin A3.

Anionic citrate is a major component of venom, but the role of venom citrate in toxicity other than its inhibitory effect on the cation-dependent action of venom toxins is poorly understood. By immobilizing Chinese hamster ovary cells in microcapillary tubes and heparin on sensor chips, we demonstrated that heparan sulfate-mediated cell retention of the major cardiotoxin (CTX) from the Taiwan cobra, CTX A3, near membrane surfaces is citrate-dependent. X-ray determination of a CTX A3-heparin hexasaccharide complex structure at 2.4 A resolution revealed a molecular mechanism for toxin retention in which heparin-induced conformational changes of CTX A3 lead to citrate-mediated dimerization. A citrate ion bound to Lys-23 and Lys-31 near the tip of loop II stabilizes hydrophobic contact of the CTX A3 homodimer at the functionally important loop I and II regions. Additionally, the heparin hexasaccharide interacts with five CTX A3 molecules in the crystal structure, providing another mechanism whereby the toxin establishes a complex network of interactions that result in a strong interaction with cell surfaces presenting heparan sulfate. Our results suggest a novel role for venom citrate in biological activity and reveal a structural model that explains cell retention of cobra CTX A3 through heparan sulfate-CTX interactions.

Amino Acid Sequence↗
PubMed1987

In vitro estimation of iron availability from a range of plant foods: influence of phytate, ascorbate and citrate.

Plant foods were digested in vitro and the proportion of iron which diffused across a semi-permeable membrane was used as an index of Fe availability. The mean (with SEM) Fe diffusibility from a group of eighteen cereals, legumes and nuts was very low, 2.1 (0.25)%, whereas from a group of sixteen fruits and vegetables it was high, 31.7 (1.09)%. The difference between the two food groups was highly significant (P less than 0.001). The results for Fe diffusibility correlated well with literature values for the in vivo absorption of Fe from similar foods (r0.84, P less than 0.01). When phytate, citrate and ascorbate were added to selected foods in amounts corresponding to endogenous levels, only phytate and citrate gave the expected effects on Fe diffusibility. Ascorbate only enhanced Fe diffusibility to the expected extent when it was added in much larger amounts, not normally found in foods. When added to cereal foods, orange juice was found to enhance greatly Fe diffusibility even when its content of ascorbate was completely destroyed by boiling followed by prolonged storage. When citrate and ascorbate were added to cereal foods in amounts equivalent to those found in fresh orange juice, both enhanced Fe diffusibility but citrate was far more effective. It is concluded that phytate is a major inhibitor of Fe diffusibility in cereals, legumes and nuts. However, citrate rather than ascorbate would appear to be the major enhancer of Fe diffusibility from many fruits and vegetables. The implications of the present results are discussed in relation to the relative influence of phytate, citrate and ascorbate on dietary Fe availability.

Ascorbic Acid↗
PubMed2026

Efficacy and safety of ferric citrate tablets in Chinese patients with hyperphosphatemia undergoing maintenance hemodialysis: a multicenter, randomized, open-label, active-controlled, phase III trial.

BACKGROUND: This study aimed to evaluate the efficacy and safety of ferric citrate tablets in Chinese patients with hyperphosphatemia undergoing maintenance hemodialysis (MHD). METHODS: In this phase III, multicenter, randomized, open-label, non-inferiority trial, patients with hyperphosphatemia on maintenance hemodialysis were randomly assigned to receive either ferric citrate or sevelamer carbonate tablets for 12 weeks. The primary endpoint was the change in serum phosphorus levels from baseline to week 12, with a non-inferiority margin of 0.32 mmol/L. Secondary endpoints included changes in serum calcium, intact parathyroid hormone, and safety assessments. RESULTS: A total of 239 patients were randomized to the ferric citrate group (n = 119) or the sevelamer carbonate group (n = 120). The mean change in serum phosphorus levels was -0.70 ± 0.50 mmol/L in the ferric citrate group and -0.61 ± 0.59 mmol/L in the sevelamer carbonate group (least squares mean difference, -0.09 mmol/L; 95% CI, -0.24 to 0.05 mmol/L; non-inferiority margin, 0.32 mmol/L). No significant inter-group differences were found in the percentage of patients achieving target phosphorus levels (49.09% vs. 48.28%, p = 0.902). Ferric citrate significantly improved iron-related parameters and hemoglobin levels. Most treatment-emergent adverse events were mild, with gastrointestinal disorders being the most common. CONCLUSIONS: Ferric citrate tablets were non-inferior to sevelamer carbonate in reducing serum phosphorus levels in hyperphosphatemia patients on maintenance hemodialysis, with the added benefit of improving iron-related anemia and a favorable safety profile.

Adult↗
PubMed2006

Effect of potassium citrate therapy on stone recurrence and regrowth after extracorporeal shockwave lithotripsy in children.

BACKGROUND AND PURPOSE: Stone disease in children may cause functional and morphologic changes in the urinary tract during longer-term follow-up. Prevention of stone recurrence will be possible only with careful metabolic evaluation and appropriate management. The possible preventive effects of potassium citrate therapy on true stone recurrence, as well as regrowth rates after shockwave lithotripsy (SWL), were evaluated in children treated for renal stones. PATIENTS AND METHODS: Following assessment of the efficacy of SWL, 96 children (72 male, 53 female aged 4 to 14 years with a mean of 6.6 years) were evaluated for the effects of potassium citrate on residual fragments as well as true new stone formation during long-term follow-up. All children had calcium-containing stones with normal renal morphology and function without any urinary-tract infection. Depending on the outcome of treatment, patients with and without residual stones were allocated independently to Group I (N = 52) and Group II (N = 44), which were matched for sex and age. Group I was given oral potassium citrate 1 mEq/kg daily for 12 months; the remaining children served as controls. Follow-up ranged from 12 to 36.6 months, with a mean of 24.4 months. Follow-up results for at least a year with respect to the stone recurrence or stone growth rates were recorded in both groups. RESULTS: Stone-free children undergoing no specific therapy had significant new stone formation compared with the group receiving potassium citrate on a regular basis (34.6% and 7.6%, respectively). Children with residual fragments receiving no specific preventive measure also showed significant new stone formation, along with enlargement of the fragments. CONCLUSION: In addition to stone removal, treatment of pediatric urolithiasis requires a thorough metabolic evaluation. Depending on the abnormalities, each patient should be advised on adequate drinking to increase the urine volume in accordance with body size. Although children with hypocitraturia may well benefit from therapeutic agents that raise the urine citrate concentration, our results did clearly show that all children bearing residual fragments should be counseled on adequate fluid intake along potassium citrate treatment to prevent stone regrowth or formation during long-term follow-up.

Adolescent↗
PubMed2003

Modelling citrate metabolism in fruits: responses to growth and temperature.

Citrate production and degradation during the last stage of fruit development were modelled by representing the fluxes through the enzymes of the citrate cycle and the malic enzyme, the transport of metabolites between the cytosol and the mitochondria, and the stoichiometry equations that relate these reactions. After solving the corresponding system of equations, the rate of citrate synthesis (or degradation) was expressed as a simple function of temperature, mesocarp weight, and respiration. The model was applied to peach fruit, and its parameters were estimated from the data of a 2-year field experiment. The predictions of the model were in agreement with experimental data. Simulations were made to analyse the responses to variations of temperature and fruit growth. Increasing fruit growth before stone hardening stimulated citrate production, while increasing fruit growth after stone hardening reduced it. Delaying the date at which the maximum growth rate was reached enhanced citrate production during most of the period. In the last weeks before harvest, increasing temperature depressed citrate production, while, at the beginning of the period studied, it enhanced it.

Citrates↗
PubMed2003

Alteration of citrate metabolism in cluster roots of white lupin.

Organic acid excretion plays a key role in the superior P(i)-acquisition of barely soluble inorganic P sources from soils. Seedlings of white lupin (Lupinus albus L.) grown for 37 d in -P nutrient solution showed typical -P symptoms, such as low P content, increased root/shoot ratio and the development of cluster roots which released large amounts of citrate. Citrate concentration in the cluster roots was 21.5 micro mol (g FW)(-1), which corresponded to a 4.3- and 2.6-fold increase of +P and -P root apexes, respectively. Cluster roots possessed higher phosphoenolpyruvate carboxylase and phosphoenolpyruvate phosphatase activity than those in +P root apexes, which could result in increasing the supply of substrate for citrate synthase. On the other hand, the cytosolic pathway which converts citrate to 2-oxoglutarate consists of aconitase and NADP-specific isocitrate dehydrogenase activity that was lower in the cluster roots than in +P root apexes, and may contribute to citrate accumulation. Thus, metabolic balance with these alterations would play an important role in increasing citrate concentration in the cluster roots. The molecular characterization of NADP-specific isocitrate dehydrogenase indicated that the cytosolic isoenzyme functions as a hetero-dimer, and that the activity would be regulated by the transcript levels for both isoforms.

Base Sequence↗
PubMed2003

Assessment of blood coagulation in severe liver disease using thromboelastography: use of citrate storage versus native blood.

Thromboelastography evaluates the viscoelastic properties of blood coagulation. Using native blood, measurement must start soon after sampling. With normal coagulation, native and citrated blood values correlate well. No data exists from cirrhotic patients. We compared native and citrate thromboelastography parameters in 30 cirrhotic patients (20 Child-Pugh C class, two liver failure). Thromboelastography was performed within 4 min using native blood and after recalcification within 1-2 h of citrate storage. Thromboelastography variables (, alpha, ) were compared using the Mann-Whitney test, correlation investigated with the Pearson method and the degree of agreement with the Bland-Altman method. There was no significant difference between citrated and native blood for all variables. Median values for native and citrated were, respectively, 16.4 (range 2.3-22.5) and 15.1 (range 9.8-29.9); 6.3 (range 3.5-11.3) and 6.2 (range 2.8-10.9); 48.3 (range 30.7-62.9) and 46.2 (range 30.4-60.4); angle alpha 30.8 (range 18.7-46.8) and 33.2 (range 19.9-55.8). Correlation for each variable was significant ( 0.01). There was a good degree of agreement for all but two patients (both bleeding) for all variables. Citrated blood can substitute native blood using thromboelastography in cirrhotic patients, allowing more time between sampling and the thromboelastography measurement.

Adult↗
PubMed1989

Neuroblastoma: imaging evaluation by sequential Tc-99m MDP, I-131 MIBG, and Ga-67 citrate studies.

Fourteen children with histopathologically confirmed neuroblastoma underwent sequential correlative imaging studies using I-131 MIBG, Tc-99m MDP, and Ga-67 citrate during various stages of the disease. Of the patients 86% showed I-131 MIBG accumulation in the primary tumoral site, whereas 71% showed Tc-99m MDP and 79% Ga-67 citrate uptake. In 86% at least one of the two latter radiopharmaceuticals concentrated in the primary tumor. The use of all three radiopharmaceuticals raised the detection rate to 93%. Of the osseous or extraosseous metastases 100% were detected by Tc-99m MDP studies. The I-131 MIBG studies were positive in 71% of the osseous metastases and in 70% of the extraosseous metastases. No Ga-67 citrate uptake was demonstrated in osseous metastases, although one extraosseous lung metastasis concentrated this radiopharmaceutical. Tc-99m MDP bone imaging was the best method for diagnosing metastatic spread of the disease and for monitoring the results of treatment. Primary tumor uptake was best indicated by I-131 MIBG. Both Ga-67 citrate and I-131 MIBG were superior to Tc-99m MDP with regard to accurately demonstrating the extent of primary tumors. Only Tc-99m MDP indicated the relationship of these tumors to the kidneys and neighboring osseous structures, providing early screening of kidney compression. Ga-67 citrate study was mainly indicated in tumors with catecholamine depletion, which failed to concentrate the other two radiopharmaceuticals. I-131 MIBG proved especially useful in detecting neuroblastoma with negative Tc-99m MDP and Ga-67 citrate studies and also proved to be helpful with those cases in which I-131 MIBG was planned for therapy. The following strategy is suggested for evaluating neuroblastoma.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Iodobenzylguanidine↗
PubMed1992

Lactic acidosis treated with continuous hemodiafiltration and regional citrate anticoagulation.

OBJECTIVE: To evaluate the effectiveness of continuous arteriovenous hemodiafiltration (CAVHD) using citrate as the anticoagulant for the treatment of lactic acidosis in patients with renal failure. DESIGN: Case series with careful monitoring of the clinical course of patients being treated in a medical or surgical ICU. SETTING: University hospital ICU. PATIENTS: Two patients with lactic acidosis are described, along with our experience using CAVHD and citrate in other clinical settings. INTERVENTIONS: CAVHD was used to manage renal failure, while a continuous infusion of citrate was administered to maintain patency of the extracorporeal circuit. MEASUREMENTS: Total and ionized serum calcium concentrations and citrate concentrations were monitored. MAIN RESULTS: CAVHD with citrate as the anticoagulant proved to be a convenient means of managing vascular volume, serum electrolyte concentrations, acid-base balance, and replacement renal function requirements in the setting of severe lactic acidosis, oliguric renal failure, and hemorrhagic diathesis. CONCLUSIONS: CAVHD with citrate as the anticoagulant can be recommended as effective therapy for selected patients, but careful monitoring is needed to avoid serious complications.

Acidosis, Lactic↗
PubMed1991

Combined 67Ga citrate and 99Tcm-human immunoglobulin imaging in human immunodeficiency virus-positive patients with fever of undetermined origin.

Patients infected with the human immunodeficiency virus (HIV) suffer from serious and life-threatening infections. These patients often present with pyrexia but without localizing signs. Despite its high sensitivity in identifying focal infection 67Ga citrate scintigraphy lacks specificity; lymphoma and solid tumours may also be imaged. This presents particular problems in HIV-positive patients with pyrexia where the differential diagnosis is often between infection and lymphoma. In an attempt to improve the specificity of radionuclide imaging in these patients a combination of 67Ga citrate and the new agent 99Tcm-labelled human immunoglobulin (99Tcm-HIG) was used in 25 patients who were sequentially imaged with the two agents. Fourteen patients had 29 sites of microbiologically confirmed infection; 67Ga citrate identified 27 sites and 99Tcm-HIG identified 16 sites. Seven of the nine sites visualized with 99Tcm-HIG, but positive with 67Ga citrate, were intrathoracic. Abnormal concentration of 67Ga citrate, not due to infection, occurred at eight sites; five lymphoma, one gout, one recent fracture and one patient with prominent bone marrow islands. 99Tcm-HIG showed increased concentration of tracer in only one of the patients with lymphoma. A combination of 67Ga citrate and 99Tcm-HIG imaging in HIV-positive patients with pyrexia of unknown origin enables a differentiation between infection and lymphoma to be made more readily. The poor sensitivity of 99Tcm-HIG in the chest will limit its sole use in this patient group.

Adult↗
PubMed1996

A kit for the instant preparation of 99Tcm-citrate for tumour and inflammation scintigraphy.

A lyophilized kit has been developed to produce 99Tcm-citrate (99%) instantly when mixed with 99TcmO4-. The ingredients of the kit, citrate and stannous chloride, are in a ratio of 100:1 (w/w) and are bound to 700 MBq 99Tcm, sufficient for use in a single patient. The shelf life of the kit when kept refrigerated is at least 3 months. The blood clearance and organ distribution of 99Tcm-citrate have been studied in rabbits and mice respectively, showing its fast clearance (55% at 5 min) and low uptake by the reticulo-endothelial system and other organs. The main route of clearance is via the kidneys. Scintigraphic studies in experimental mice have demonstrated high levels of accumulation of 99Tcm-citrate in turpentine-induced abscess (target-to-non-target ratio 4.2:1) and EAT implanted tumour in mice (T/NT 5:1) 1 h after its administration. In keeping with these animal studies, low uptake of 99Tcm-citrate by normal organs and fast blood clearance have been demonstrated in human patients with infection and tumour as early as 5 min post-administration. This study documents a new kit developed for the preparation of 99Tcm-citrate suitable for the scintigraphic assessment of abscess, infection and tumour.

Abscess↗
PubMed1987

Hypothermic preservation of the rat liver assessed by orthotopic transplantation. II. Evaluation of citrate solutions.

Rat livers were flushed with isotonic citrate solution (IC), hypertonic citrate solution (HC), or Collins' solution (C2), and were stored at 0 degree C for 8-16 hr. Following 8-hr preservation, the number of animals surviving for one month was greatest with IC grafts (6/8) and least with C2 grafts (2/7). There was no significant difference between the two citrate groups. Following 12-hr preservation, the IC group was superior to the HC group (3/6 vs. 1/6). Following 16-hr preservation, all four animals in each citrate group died within 24 hr. The ability of livers to produce bile was greater in the citrate groups, compared with the C2 group. A bile flow rate less than 0.3 microliter/min/g liver, 15 min after implantation, appeared to predict subsequent poor survival. There were no significant histological differences between biopsies taken from each group at the end of the transplant operation. Biochemical measures of liver function were best in animals that had received IC grafts, and functional damage after preservation was consistently greater in the C2 group than the citrate groups. The best results were obtained with grafts preserved with IC solution.

Animals↗
PubMed1980

Studies on the induction and turnover of citrate-oxidizing capacity in Klebsiella aerogenes using chemostat culture.

Steady-state chemostat cultures of Klebsiella aerogenes growing on a carbon-limited medium were challenged with pulse of carbon sources, and the resultant changes in dissolved oxygen tension were shown to reflect changes in respiration rate. These changes were used to study the kinetics of induction of citrate-oxidizing capacity, which is most probably limited by citrate permease, and the return to the preinduced state. Previously unexposed cells showed a lag phase, the duration of which decreased with increasing growth rate, with a minimum of 10 min, followed by an induction phase of linear increase of citrate oxidation rate which continued as long as citrate was present. The rate of increase in activity, which can be equated to the rate of induction of citrate permease, was independent of citrate concentration but increased with growth rate. Previously exposed cells showed no lag and some residual activity before further induction. The kinetics of return to the preinduced state were unusual in that activity was short-lived with a half-life of 16 to 23 min while the lag took over 8 h to re-establish. The rate of decay of activity decreased with increasing growth rate.

Citrates↗
PubMed1979

Citrate in plasma and urine during total fasting.

Plasma citrate was determined in 12 obese subjects who underwent total fasting for 10 days. Mean plasma citrate concentration rose significantly from 128 before to 205 micro mol/1 on the 10th day of fasting. Plasma citrate rose continuously during fasting in seven subjects in whom daily determinations were carried out. The 24-hour urinary citrate excretion was followed in six subjects. A significant decrease was found from 2.91 mmol/24 h in the prefasting state to 0.25 mmol/24 h at the end of the fast. Intravenous glucose tolerance test were performed before and on the 10th day of fasting. Kivgtt decreased significantly and was inversely related to plasma citrate concentration on the 10th day of fasting. The results agree well with the concept that an increased citrate level of tissues is of regulatory importance for the decreased glucose utilization during fasting in man.

Adolescent↗
PubMed1977

Purification and properties of citrate lyase ligase from Streptococcus diacetilactis.

Citrate lyase ligase (acetate: SH--[acyl-carrier protein] enzyme ligase (AMP) from Streptococcus diacetilactis was purified 920-fold with a yield of 6.3%. The molecular weight of the enzyme was estimated to be 41000; the ligase consisted of one polypeptide chain. The acetylation of 1 mol of deacetyl-citrate lyase to enzymatically active citrate lyase required 6 mol ATP. The formation of AMP and pyrophosphate in the acetylation reaction was demonstrated. Citrate lyase ligase was specific for the lyase from S. diacetilacitis and did not acetylate lyases from Rhodopseudomonas gelatinosa and Enterobacter aerogenes. The substract acetate and ATP could be replaced by propionate and dATP, repectively. The reaction rates for ATP, acetate and deacetyl-citrate lyase followed Michaelis-Menten kinetics (Km values: 26 micron for ATP, 25 mM for acetate and 38 nM for deacetyl-citrate lyase).

Citrates↗
PubMed1978

On the significance of the prosthetic group composition of citrate lyase.

1. Klebsiella aerogenes contains two different acyl carrier proteins, one specific for citrate lyase, the other for fatty acid synthetase. 2. The acyl carrier protein of fatty acid synthetase from K. aerogenes was isolated and compared with the corresponding protein from Escherichia coli and with the acyl carrier protein of citrate lyase from K. aerogenes. 3. As judged from prosthetic group compositions as well as amino acid and fingerprint analyses, the acyl carrier proteins of the two fatty acid synthetases are nearly identical but different from that of citrate lyase from K. aerogenes. 4. Therefore, the different prosthetic groups alone cannot be responsible for the different specificities of the acyl carrier proteins of fatty acid synthetase and citrate lyase in K. aerogenes. 5. The prosthetic group of citrate lyase, phosphoribosyl dephospho-CoA, apparently represents no incidental, phosphopantetheine-replacing aberration. The requirement of citrate lyase for the CoA-like prosthetic group may arise from the substrate requirement of both subunit enzymes of the enzyme complex.

Amino Acids↗
PubMed1983

Regulation of citrate efflux from mitochondria of oleaginous and non-oleaginous yeasts by long-chain fatty acyl-CoA esters.

1. Citrate efflux from a wide range of yeast mitochondria was inhibited by long-chain fatty-acyl-CoA esters. 2. Fatty-acyl-CoA esters with chain lengths of between 14 and 18 carbons were the most potent inhibitors of citrate efflux which was unaffected by the fatty acids themselves. 3. 50% inhibition of citrate transport was observed using palmitoyl-CoA and oleoyl-CoA at approx. 4-5 microM when the tricarboxylate carrier was saturated with L-malate as counter-anion. 4. The inhibition with palmitoyl-CoA and oleoyl-CoA was competitive with L-malate. 5. The possibility that the fatty-acyl-CoA esters were exerting their effect by acting as detergent was eliminated because of the low concentrations used and appropriate comparisons being made with non-specific detergents. Although detergents inhibited citrate efflux they also released citrate by causing membrane damage. 6. The effect of fatty-acyl-CoA esters on citrate efflux could be decreased by using higher mitochondrial protein levels and by adding bovine serum albumin. 7. The possibility is discussed that this inhibition represents a genuine feedback inhibition which could regulate the amount of lipid being synthesized by an oleaginous yeast.

Acyl Coenzyme A↗
PubMed2004

The use of recalcified citrated whole blood -- a pragmatic approach for thromboelastography in children.

BACKGROUND: Thromboelastography (TEG) is an established way of monitoring the coagulation status of children and adults requiring blood products during surgery. Serial measurements are performed using a nearside machine and blood product prescription may be titrated against changes in TEG. There may also be useful applications when the patient is remote from the TEG machine but these are limited because TEG is usually performed on fresh native whole blood within 6 min of venepuncture. Citrated whole blood can be used for TEG if transport time is more than 6 min. We wished to establish whether TEG parameters for citrated whole blood were comparable with those of native whole blood in healthy children. METHODS: Blood was obtained from 14 healthy children undergoing minor surgical procedures, at the time of intravenous cannula insertion for anaesthesia. Each sample was divided: TEG was performed on part of the sample in its fresh native state at 6 min and second portion of the sample was citrated, kept at room temperature and TEG was performed at 30 min after recalcification. RESULTS: There was a significant difference in TEG parameters (r, k, alpha, MA and LY30) for fresh native whole blood and recalcified citrated whole blood (paired t-test). CONCLUSIONS: The normal range for fresh native whole blood TEG parameters is well established, which is routinely used in practice. There was a significant difference between TEG parameters for fresh native whole blood and citrated whole blood. We recommend that a specific normal range be established for citrated whole blood to enable it to be used in clinical practice.

Adolescent↗