Diagnóstico molecular da talassemia ALFA em indivíduos com microcitose e/ou hipocromia

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Caracterização molecular e laboratorial da talassemia beta e da interação hemoglobina s/talassemia beta

Beta thalassemia arises as a consequence of the reduction (β+, β++, βsilent) or absence (β0) of beta globin chain synthesis and results from a number of mechanisms that lead to genetic defects. The inheritance of beta thalassemia is characterized by the existence of heterozygous individuals, compound heterozygotes, homozygotes and those with coinheritance of beta thalassemia allele and other thalassemias and/or hemoglobin variants. The aim of this study was to perform molecular and laboratory characterization of beta thalassemia in heterozygous and homozygous individuals and in those with coinheritance of S beta thalassemia. A total of 48 individuals were included (35 heterozygotes, 4 homozygotes and 9 S beta thalessemia carriers) referred to the Integrated Laboratory of Clinical Analyses of the Federal University of Rio Grande do Norte (UFRN) and the Hematology Ambulatory Facility of the Dalton Barbosa Cunha Hemocenter (Hemonorte Natal, Brazil). Peripheral blood samples form each patient underwent the following laboratory examinations: erythrogram, hemoglobin electrophoresis at alkaline pH, measurements of Hb A2, Fetal Hb and serum ferritin. DNA was extracted using the illustra blood genomicPrep Mini Spin Kit and molecular characterization was performed by the PCR/RFLP technique, which involves digestion with specific restriction enzymes for IVS-1 nt 1 (G®A), IVS-1 nt 6 (T®C) and codon 39 (CAG®TAG) mutations. Of the 35 heterozygotes, 37.1% showed IVS-1 nt 6 mutation, 42.9% IVS-1 nt 1 and 20% were carriers of other mutations not identified by the technique used. The four homozygous patients presented with the IVS-1 nt 6 mutation, while 66.7% of the individuals with S beta thalassemia had the IVS-1 nt 1 mutation. Codon 39 was not detected in any of the patients investigated. Of the thallasemic alleles found, 40.4% were IVS- 1 nt 1, 40.4% IVS-1 nt 6 and 19.2% were not identified. Laboratory data showed that the heterozygotes exhibited microcytosis and hypochromia, evidenced by MCV ranging from 57 to 75fL and MCH from 15.9 to 23.6 pg. Hemoglobin A2 varied between 3.7 and 7.2%. The homogygotes also showed reduced MCV and MCH and elevated HbA2.. Comparison of laboratory data between heterozygous individuals with IVS-1 nt 1 and IVS-1 nt 6 mutations showed that heterozygotes for the IVS1-1 mutation had significantly lower mean MCV and MCH (p = 0.023 and 0.007, respectively) and significantly higher hemoglobin A2 (p < 0.001) when compared to heterozygotes for the IVS-1 nt 6 mutation. PCR/RFLP was useful in identifying the presence or absence of IVS-1 nt 6, IVS-1 nt 1 and codon 39 mutations in most of the patients investigated here. This is the first study conducted in the state of Rio Grande do Norte, Brazil aimed at identifying beta thalassemia mutations and represents an important contribution to the knowledge regarding the molecular profile of beta thalassemia in our country

Análise da frequência alélica de 15 LOCI STR na população do Rio Grande do Norte

Human population have a significant number of polymorphic loci, whose use and applications range from construction of linkage maps, to study the evolution of populations, through the determination of paternity, forensic medicine and migration. Currently, STRs (Short Tanden Repeats) markers are considered the major markers for human identification, mainly due to its abundance and high variability because of the fact that they are easily amplifiable by PCR (Polymerase Chain Reaction), work with low amounts of DNA and be capable of automation processes involving fluorescence detection. The creation of regional databases containing allele frequencies of population provide subsidies to increase the reliability of the results of determining the genetic link. This paper aims to obtain a database of allele frequencies of 15 polymorphic molecular loci (D8S1179, D21S11, D7S820, CSF1PO, D19S433, vWA, TPOX, D18S51, D3S1358, TH01, D13S317, D16S539, D2S1338, D5S818 e FGA) in a population classifies as born in the State of Rio Grande do Norte, Brazil, totaling 1100 unrelated individuals. To evaluate the frequency, DNA samples were submitted to PCR amplification, followed by capilarry electrophoresis genetic sequencer. The frequencies identified in this study were compared with brazilian population in general and other states in Brazil. Except for the loci D21S11, D19S433 and D2D1338, the genotypes found were in Hardy-Weinberg equilibrium and no significant differences among the frequencies were found in the populations studied. The most informative loci was D2S1338 and D18S51, and the less informative is the locus TPOX

Produção e avaliação de nanoemulsão catiônica como possível vetor não-viral para pIRES2-EGFP

Gene therapy is based on the transfer of exogenous genetic material into cells or tissues in order to correct, supplement or silencing a particular gene. To achieve this goal, efficient vehicles, viral or non-viral, should be developed. The aim of this work was to produce and evaluate a nanoemulsion system as a possible carrier for no-viral gene therapy able to load a plasmid model (pIRES2-EGFP). The nanoemulsion was produced by the sonication method, after been choose in a pseudo-ternary phase diagram build with 5 % of Captex 355®, 1.2 % of Tween 80®, 0.8 % of Span 80®, 0.16% of stearylamine and water (to 100 %). Measurements of droplet size, polydispersity index (PI), zeta potential, pH and conductivity, were performed to characterize the system. Results showed droplets smaller than 200 nm (PI < 0.2) and zeta potential > 30 mV. The formulation pH was near to 7.0 and conductivity was that expected to oil in water systems (70 to 90 μS/s) A scale up study, the stability of the system and the best sterilization method were also evaluated. We found that the system may be scaled up considering the time of sonication according to the volume produced, filtration was the best sterilization process and nanoemulsions were stable by 180 days at 4 ºC. Once developed, the complexation efficiency of the plasmid (pDNA) by the system was tested by agarose gel electrophoresis retardation assay.. The complexation efficiency increases when stearylamine was incorporated into aqueous phase (from 46 to 115 ng/μL); regarding a contact period (nanoemulsion / pDNA) of at least 2 hours in an ice bath, for complete lipoplex formation. The nanoemulsion showed low toxicity in MRC-5 cells at the usual transfection concentration, 81.49 % of survival was found. So, it can be concluded that a nanoemulsion in which a plasmid model was loaded was achieved. However, further studies concerning transfectation efficiency should be performed to confirm the system as non-viral gene carrier

Utilização de um secador solar para desintoxicação da torta de mamona

It´s been motivating motivated by the current rulers as an energy solution the use of the biodiesel as source of energy, that doesn't damage the environment and promotes the development of the areas where the base plant are grown. The process of production of the biodiesel, starting from the castor oil plant and its derivers, generates a known product as castor oil plant pie that has been used as fertilizer. Once disintoxicated, it can serve as animal ration, because it contains high content of proteins, presenting larger joined value. The disintoxication process, can be obtained through the process of drying the pie with the construction and use of models of dryers capable to elevate the temperature of the castor oil plant pie to approximately 60ºC.In this temperature the product tends to eliminate the ricina, with the aid of an aqueous solution of hydroxide of calcium, or still reach higher temperatures to make the disintoxication without a chemical treatment. It was made a bibliographical research about known processes of disintoxication of the castor oil plant pie from the autoclave use to the drying direct exposure to the sun. Starting from the state of the art and identification of the object, it was chosen solar dryers to eliminate the ricina. It was applied two types of solar dryers: the one of solar direct exposure, and the one with concentrador. The castor oil plant pie was separated in samples, with measurement of its initial mass, codified and placed in the dryers. The results were presented in graphs and tables forms, with the values of temperatures noticed. It was noticed the variations of temperature and the relationships analyzed related with the ricina content eliminated from the pie. The analysis of the ricina content was accomplished by Embrapa - Campina Grande, by eletroforese method. The analysis of the result of considering the content of ricina of the samples. It was observed that, we obtained lower rate of ricina in samples that had larger drying time and average value temperature above 60ºC. Comparing with the ones submitted to higher temperatures and in a shorter period of time. It was possible to evaluate the efficiency of the dryers in the desintoxication process of the samples, as well as the type of more appropriate dryer for the drying pie process. Finally, it was concluded that the solar dryer with concentrador presents higher values temperatures than the direct exposure one. So, it´s being more opportune applied in the castor oil plant pie drying process. However, more than one hour for drying time is needed

Avaliação de dois clones de Escherichia coli recombinante quanto ao crescimento e expressão de antígenos de Leishmania chagasi (kmp11 e P36)

Visceral leishmaniosis caused by Leishmania chagasi, also known as calazar, presented, in the period from 1990 to 2005, tax of incidence in Brazil varying between 1 and 3 cases for 100 000 inhabitants. The Northeast region that up to the year of 2000 contributed with almost 90% of the registered cases is reducing his participation in the current decade, reaching 56% in 2005. Conventional leishmaniasis treatment is costly and it shows high toxicity, demanding more research for alternative treatments, with special interest in development of vaccines and diagnosis kits which include production of recombinant antigens by host cells. Escherichia coli has been the microorganism most studied and used as a host for recombinant protein production. Therefore, the aim of this work was to study the influence of induction on cellular growth and to verify the type of Leishmania chagasi antigens expression (intra or extracellular) during two recombinant E. coli clones (kmp11 and P36) cultivation in rotary incubator (shaker) using three different media (2xTY, TB, FASS+EL). For that, tests were carried out using conditions established in the literature for E. coli (37°C and 200 rpm) and media supplemented with antibiotics to guarantee that only competent cells grows. First, tests were carried out without induction in order to verify the two microorganisms kinetic behavior (growth and substrate consumption) in different media. Next, the induction was carried out through the addition of IPTG (1mM as final concentration), at the first hour of cultivation. It was observed that protein expression were intracellular for all clones and media tested, however the highest level of expression was clearly observed by the electrophoresis band density (intensity) for 2xTY medium and kmp11 protein. Although it contains the lowest substrate concentration, consequently, a reduced cellular concentration when compared to other media, it appeared that this medium and clone combination is the most indicated for recombinant protein production. Therefore, the objective of this work was achieved, since the interested proteins were produced. Consequently, this result motivates new studies for production optimization using different cultivation strategies

Análise molecular dos éxons 8 a 11 do gene da p53 em amostras de câncer de colo de útero no Rio Grande do Norte

Mutations on TP53 gene are common in human cancer but not in cervical cancer where they are rarely found and the inactivation and degradation of p53 protein are attributed to the action of E6 viral oncogene from high risk human papillomavirus (HPV). Analysis of cervical cancer cell lines suggests that HPV negative samples shows mutation on TP53, but clinical approaches didn t confirmed this hypothesis. However, in most TP53 mutations studies on cervical cancer, only the exons 5 to 8 were analyzed. Approximately 90% of mutations described are on this region. Recent studies on several cancer suggests that mutation frequency in the other exons must be considered. The aim of this work was to verify whether mutations on coding and non-coding regions occur in cancer tissue from cervical cancer in patients from Rio Grande do Norte using Denaturing Gradient Gel Electrophoresis (DGGE) as screening tool. Exons 8 to 11 were analyzed including some introns from 80 tumor samples and 8 peripheral blood samples from healthy women. DNA were submitted to PCR using primers with GC clamp on the end of one of them. The results were observed for each region after DGGE and silver staining. It was observed no amplified fragment with different migration profile from those obtained from DNA of peripheral blood. These results agree with those from literature where TP53 mutations in cervical cancer have been described in a very low frequency

Desenvolvimento de sistemas farmacêuticos emulsionados para veiculação gênica

Broadly speaking, the concept of gene therapy involves the transfer of a genetic material into a cell, tissue, or organ in order to cure a disease or at least improve the clinical status of a patient. Making it simple, gene therapy consists in the insertion of functional genes into cells containing defective genes by substituting, complementing or inhibiting them. The achievement of a foreigner DNA expression into a population of cells requires its transfer to the target. Therefore, it is a key issue to create systems able to transfer and protect the DNA until it reaches the target, the vectors. The disadvantages related to the use of viral vectors have encouraged efforts to develop emulsions as non-viral vectors. In fact, they are easily produced, present controllable stability and enable transfection. The aim of this work was to develop an emulsion for gene therapy and evaluate its ability to compact nucleic acids by the development of a complex with the plasmid pIRES2-EGFP. The first step was to determine the Hydrophilic Lipophilic Balance (HLB) of the Captex® 355 (oily internal phase of the emulsion) through long and short term stability assays. Based on the results, emulsions composed of Captex® 355, Tween 20® and Span 60® with 10.7 HLB were produced by three different methods: phase inversion, spontaneous emulsification and sonication. The results showed that the lowest diameter and best stability of the emulsions were achieved by the sonication method. The cationic emulsions were made by adding DOTAP to the basic emulsion. Its association with pIRES2-EGFP was evaluated by electrophoresis. Several rates of emulsion and DNA were evaluated and the results showed that 100% of the complex was formed when the rate DOTAP/DNA(nmol/µg) was 130. In conclusion, the overall results show the ability of the proposed emulsion to compact pIRES2-EGFP, which is a requirement to a successful transfection. Therefore, such formulation may be considered a promising candidate for gene therapy

Estudo integrado do potencial fitorremediador da Eichhornia crassipes em ambientes naturais e sua utilização para obtenção de extratos proteicos

This study aimed to analyze the phytoremediation potential of Eichhornia crassipes in natural environments, optimize the extraction process of crude protein from plant tissue and, obtain and characterize this process in order to determine its viability of use instead of the protein sources of animal and/or human feed. For this, it has been determined in Apodi/Mossoró river water the concentration of ammonium ions, nitrite, nitrate, calcium, magnesium, potassium, iron, copper, manganese, zinc, nickel, cobalt, sodium, aluminum, cádmium, lead, and total chromium; It was determined in plant tissue of aquatic macrophytes of Eichhornia crassipes species present in Apodi/Mossoró River the moisture content, ash, calcium, magnesium, potassium, iron, copper, manganese, zinc, nickel, cobalt, sodium, aluminum, cadmium, lead, total chromium, total nitrogen and crude protein. It was also determined the translocation factor and bioaccumulation of all the quantified elements; It was developed and optimized the extraction procedure of crude protein based on the isoelectric method and a factorial design 24 with repetition; It was extracted and characterized the extract obtained by determining the moisture content, ash, magnesium, potassium, iron, copper, manganese, zinc, nickel, cobalt, sodium, cadmium, total nitrogen and crude protein. And finally, it was also characterized the protein extract using Thermogravimetric Analysis (TG), Derived Thermogravimetric (DTG), Differential Scanning Calorimetry (DSC), Infrared Spectroscopy (FT-IR) and jelly-like electrophoresis of polyacrylamide (SDS -PAGE) to assess the their molecular weights/mass. Thus, from the results obtained for the translocation and bioaccumulation factors was found that the same can be used as phytoremediation agent in natural environments of all quantified elements. It was also found that the developed method of extraction and protein precipitation was satisfactory for the purpose of the work, which gave the best conditions of extraction and precipitation of proteins as: pH extraction equal to 13.0, extraction temperature equals 60 ° C, reaction time equals to 30 minutes, and pH precipitation equals to 4.0. As for the extract obtained, the total nitrogen and crude protein were quantified higher than those found in the plant, increasing the crude protein content approximately 116.88% in relation to the quantified contente in the vegetal tissue of macrophyte. The levels of nickel and cadmium were the unique that were found below the detection limit of used the equipment. The electrophoretic analysis allowed us to observe that the protein extract obtained is composed of low polypeptide chains by the molecular and phytochelatins, with 6 and 15 kDa bands. Analysis of TG, DTG, DSC and FT-IR showed similarities in protein content of the obtained extracts based on different collection points and 9 parts of the plant under study, as well as commercial soy protein and casein. Finally, based on all these findings, it was concluded that the obtained extract in this work can be used instead of the protein sources of animal feed should, before that, test its digestibility. As human supplementation, it is necessary to conduct more tests associated with the optimization process in the sense of removing undesirable components and constant monitoring of the water body and the raw material used

Determinação de haplótipos do gene βs em portadores de anemia falcifome

Haplotypes linked to the βS gene represent patterns of DNA polymorphisms along chromosome 11 of individuals bearing the βS gene. Analysis of haplotypes, in addition to serving as an important source for anthropological studies about the ethnic origin of a population, contributes to a better understanding of the variations in clinical severity of sickle cell anemia. The aim of the present study was to determine βS gene haplotypes in a group of patients with sickle cell anemia treated at the Dalton Barbosa Cunha Hematology Center (Hemonorte) in Natal, Brazil and the Oncology and Hematology Center in Mossoró, Brazil. Blood samples were obtained from 53 non-related patients (27 males and 26 females), aged between 3 months and 61 years (mean age: 16.9 ± 12.1 years). Laboratory analyses consisted of the following: erythrogram, reticulocyte count, hemoglobin electrophoresis at alkaline pH, measurement of hemoglobin A2 and Fetal hemoglobin, solubility test and molecular analysis to determine βS gene haplotypes. DNA samples were extracted by illustra blood genomicPrep Mini Spin kit and βS gene haplotypes were determined by PCR-RFLP, using Xmn I, Hind III, Hinc II and Hinf I restriction enzymes for analysis of six polymorphic restriction sites in the beta cluster. Of 106 βS chromosomes studied, 75.5% were Central African Republic (CAR) haplotype, 11.3% Benin (BEN) and 6.6% Cameroon (CAM). The atypical haplotypes had a frequency of 6.6%. More than half the patients (58.5%) were identified as CAR/CAR genotype carriers, 16.9% heterozygous CAR/BEN, 13.2% CAR/CAM and 1.9% BEN/BEN. Patients with atypical haplotype in one or two chromosomes accounted for 9.5% (CAR/Atp, BEN/Atp and Atp/Atp). The genotype groups showed no statistically significant difference (p < 0.05) in their laboratory parameters. This is the first study related to βS haplotypes conducted in state of Rio Grande do Norte and the higher frequency of Cameroon halotype found, compared to other Brazilian states, suggests the existence of a peculiarity of African origin

Otimização da extração de polissacarídeos sulfatados da alga vermelha Gracilaria birdiae e análise da atividade anticoagulante e antioxidante

The sulfated polysaccharides (SP) from the edible red seaweed Gracilaria birdiae were obtained using five different condition extraction (GB1: Water; GB1p: Water/proteolysis; GB1s: Water/sonication; GB1sp: Water/sonication/proteolysis; GB2s: NaOH/sonication; GB2sp: NaOH/sonication/proteolysis. The yield (g) increased in the following order GB2sp>GB1sp>GB1p>GB2s>GB1s>GB1. However, the amount of SP extracted increased in different way GB2sp>GB1p>GB1>GB1sp>GB1s>GB2s. Infrared and electrophoresis analysis showed that all conditions extracted the same SP. In addition, monosaccharide composition showed that ultrasound promotes the extraction of other polysaccharides than SP. In the prothrombin time (PT) test, which evaluates the extrinsic coagulation pathway, none of the samples showed anticoagulant activity. While in the activated partial thromboplastin time (aPTT) test, which evaluates the intrinsic coagulation pathway, all samples showed anticoagulant activity, except GB2s. The aPTT activity decreased in the order of GB1sp>GB2sp>GB1p>GB1>GB1s>GB2s. Total capacity antioxidant (TCA) of the SP was also affected by condition extraction, since GB2s and GB1 showed lower activity in comparison to the other conditions. In conclusion, the conditions of SP extraction influence their biological activities and chemical composition. The data showed NaOH/sonication/proteolysis was the best condition to extract anticoagulant and antioxidant SPs from Gracilaria birdiae.

Análise estrutural e avaliação do efeito condroitim sulfato extraído de tilápia (Oreochromis niloticus) em modelo de peritonite aguda

Chondroitin sulfate (CS) is a naturally glycosaminoglycan found in the extracellular matrix of connective tissues and it may be extracted and purified those tissues. CS is involved in various biological functions, which may be related to the having structural variability, despite the simplicity of the linear chain structure from this molecule. Researches in biotechnology and pharmaceutical field with wastes from aquaculture has been developed in Brazil. In recent decades, tilapia (Oreochromis niloticus), native fish from Africa, has been one of the most cultivated species in various regions of the world, including Brazil. The tilapia farming is a cost-effective activity, however, it generates large amount of wastes that are discarded by producers. It is understood that waste from tilapia can be used in research as a source of molecules with important biotechnological applications, which also helps in reducing environmental impacts and promote the development of an ecofriendly activity. Thus, nile tilapia viscera were subjected to proteolysis, then the glycosaminoglycans were complexed with ion exchange resin (Lewatit), it was fractionated with increasing volumes of acetone and purified by ion exchange chromatography DEAE-Sephacel. Further, the fraction was analyzed by agarose gel electrophoresis and nuclear magnetic resonance (NMR). The electrophoretic profile of the compound together the analysis of 1H NMR spectra and the HSQC correlation allow to affirm that the compound corresponds to a molecule like chondroitin sulfate. MTT assay was used to assess cell viability in the presence of CS tilapia isolated and showed that the compound is not cytotoxic to normal cells such as cells from the mouse embryo fibroblast (3T3). Then, this compound was tested for the ability to reduce the influx of leukocytes in model of acute peritonitis (in vivo) induced by sodium thioglycolate. In this context, it was done total and differential leukocytes counting in the blood and peritoneal fluid collected respectively from vena cava and the peritoneal cavity of the animals subjected to the experiment. The chondroitin sulfate for the first time isolated from tilapia (CST ) was able to reduce the migration of leukocytes to the peritoneal cavity of inflamed mice until 80.4 per cent at a dose 10µg/kg. The results also show that there was a significant reduction (p<0.001) of the population of polymorphonuclear leukocytes from peritoneal cavity in the three tested doses (0.1µg/kg; 1µg/kg and 10µg/kg) when it was compared to the positive control (just thioglycolate). Therefore, since the CST structure and mechanism of action has been completely elucidated, this compound may have potential for therapeutic use in inflammatory diseases

Nanopartículas catiônicas de poli (ácido lático) para liberação modificada de peptídios da peçonha do escorpião Tityus serrulatus

Reported accidents involving the poisoning scorpions are still frequent in Brazil, mainly caused by Tityus serrulatus, known as yellow scorpion. Although antivenom sera are produced routinely by various government laboratories, the effectiveness of its use depends on how quickly treatment is initiated and efficiency in the production of antibodies by the immunized animals. In this study, the development of cationic polymeric nanoparticles of poly(lactic acid) aimed to create a modified delivery system for peptides and proteins of T. serrulatus venom, able to enhance the production of serum antibodies against the scorpion toxins. The cationic nanoparticles were obtained by a low energy nanoprecipitation, after study of the parameters’ variations effects over the physicochemical properties of the particles. The surface functionalization of the nanoparticles with the hyperbranched polyethyleneimine was proved by zeta potential analysis and enabled the adsorption by electrostatic interaction of different types of proteins. The protein loading efficiency of 40-80 % to bovine serum albumin (BSA) and 100 % to scorpion venom peptides evaluated by spectrophotometry and polyacrylamide gel electrophoresis confirmed the success of the selected parameters established for obtainment of nanoparticles, produced with size between 100 to 250 nm. The atomic force microscopy analysis and in vitro release showed that the spherical nanoparticles provided a sustained release profile of proteins by diffusion mechanism, demonstrating the potential for application of the nanoparticles in vivo.

Nanopartículas catiônicas de poli (ácido lático) para liberação modificada de peptídios da peçonha do escorpião Tityus serrulatus

Reported accidents involving the poisoning scorpions are still frequent in Brazil, mainly caused by Tityus serrulatus, known as yellow scorpion. Although antivenom sera are produced routinely by various government laboratories, the effectiveness of its use depends on how quickly treatment is initiated and efficiency in the production of antibodies by the immunized animals. In this study, the development of cationic polymeric nanoparticles of poly(lactic acid) aimed to create a modified delivery system for peptides and proteins of T. serrulatus venom, able to enhance the production of serum antibodies against the scorpion toxins. The cationic nanoparticles were obtained by a low energy nanoprecipitation, after study of the parameters’ variations effects over the physicochemical properties of the particles. The surface functionalization of the nanoparticles with the hyperbranched polyethyleneimine was proved by zeta potential analysis and enabled the adsorption by electrostatic interaction of different types of proteins. The protein loading efficiency of 40-80 % to bovine serum albumin (BSA) and 100 % to scorpion venom peptides evaluated by spectrophotometry and polyacrylamide gel electrophoresis confirmed the success of the selected parameters established for obtainment of nanoparticles, produced with size between 100 to 250 nm. The atomic force microscopy analysis and in vitro release showed that the spherical nanoparticles provided a sustained release profile of proteins by diffusion mechanism, demonstrating the potential for application of the nanoparticles in vivo.

Homogalactanas sulfatadas da alga Codium isthmocladum com atividade anticoagulante

Since the first description of sulfated polysaccharides from seaweeds, the biological activities of these compounds have been evaluated under different aspects and experimental procedures. Among the broad biological activities presented by seaweed polysaccharides, anticoagulant action appears as a promising function. In this present study we have obtained sulfated polysaccharides from the green seaweed Codium isthmocladium by proteolytic digestion, followed by separation into five fractions (0.3, 0.5, 0.7, 0.9 and 1.2) by sequential acetone precipitation. The chemical analyses have demonstrated that all fractions are composed mainly by sulfated polysaccharides. The anticoagulant activity of these fractions was determined by activated partial thromboplastin time (aPTT) and prothrombin time test (PT) using citrate normal human plasma. None fraction has shown anticoagulant activity by PT test. Furthermore, all of them have shown anticoagulant activity by aPTT test. These results indicated that the molecular targets of these sulfated polysaccharides are mainly in the intrinsic via of the coagulation cascade. Agarose gel electrophoresis in 1,3-diaminopropane acetate buffer, pH 9.0, stained with 0.1% toluidine blue showed the presence of two or three bands in several fractions while the fraction 0.9 showed a single spot. By anion exchange chromatography, the acid polysaccharides from the 0.9 acetone fraction were separated into two new fractions eluted respectively with 2.0 and 3.0 M NaCl. These compounds showed a molecular weight of 6.4 and 7.4 kDa respectively. Chemical analyses and infrared spectroscopy showed that Gal 1 and Gal 2 are sulfated homogalactans and differ one from the other in degree and localization of sulfate groups. aPPT test demonstrated that fractions 2,0 and 3,0M (Gal1 and Gal 2, respectively) have anticoagulant activity. This is the first time that anticoagulant sulfated homogalatans have been isolated from green algae. To prolong the coagulation time to double the baseline value in the aPTT, the required amount of sulfated galactan 1 (6,3mg) was similar to low molecular heparin Clexane®, whereas only 0,7mg of sulfated galactan 2 was needed to obtain the same effect. Sulfated galactan 2 in high doses (250mg) induces platelet aggregation. These results suggest that these galactans from C. isthmocladum have a potential application as an anticoagulant drug