Role of Synadenium grantii latex proteases in nematicidal activity on Meloidogyne incognita and Panagrellus redivivus / Papel das proteases do látex de Synadenium grantii na atividade nematicida sobre Meloidogyne incognita e Panagrellus redivivus

Abstract Synadenium grantii is a Euphorbiaceae plant commonly found in Brazil, known as Janaúba or Leitosinha, whose latex is traditionally used for several purposes. However, it is not known whether the nematicidal action of this plant latex occurs due to the action of proteases. The present work aims to evaluate the nematicidal activity of proteases from Synadenium grantii latex on Meloidogyne incognita and Panagrellus redivivus. S. grantii latex used in the present study was collected from specimens found in Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. The drained latex was collected in Eppendorf microtubes and immediately stored on ice at 4 °C. After this extraction, the latex was frozen (-20 °C) during 2 hours, thawed at room temperature (25 °C) and centrifuged at 10,000 g at 4 °C for 30 minutes to remove larger particles and concentrate the proteases. After the centrifugation, assays of enzymatic activity were performed in order to know in which of the phases the enzymes were found. S. grantii latex presented protease, but no chitinase activity. The results show that there was a significant difference (p <0.01) between the treated and control groups, with 100% mortality of Meloidogyne incognita and 72% average mortality of Panagrellus redivivus. In addition, it was demonstrated that the nematicidal action occurred due to the action of the proteases, since the control was only differentiated from the treatment by the presence of the enzymes with biological activity.

Resumo Synadenium grantii é uma planta Euphorbiaceae comumente encontrada no Brasil, conhecida como Janaúba ou Leitosinha, e tem seu látex usado tradicionalmente para diferentes propósitos. Entretanto, não se conhece se a atividade nematicida da planta ocorre devido à ação de proteases. O presente trabalho tem como objetivo avaliar a atividade nematicida das proteases do látex de Synadenium grantii sobre Meloidogyne incognita e Panagrellus redivivus. O látex de S. grantii utilizado no presente trabalho foi coletado a partir de espécimes encontradas na Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. O látex foi coletado em microtubos Eppendorf e imediatamente armazenado em gelo a 4 °C. Após esta extração, o látex foi congelado (-20 °C) durante 2 horas, descongelado à temperatura ambiente (25 °C) e centrifugado a 10000 g a 4 °C durante 30 minutos para a remoção de partículas e concentração das proteases. Após a centrifugação, foram realizados ensaios de atividade enzimática para saber em qual das fases as enzimas foram encontradas. O látex de S. grantii apresentou atividade de protease, mas nenhuma atividade de quitinase. Os resultados mostram que houve diferença significativa (p <0,01) entre os grupos tratados e controle, com 100% de mortalidade de Meloidogyne incognita e 72% de mortalidade média de Panagrellus redivivus. Além disso, foi demonstrado que a ação nematicida ocorreu devido à ação das proteases, uma vez que o grupo controle só foi diferenciado do tratamento pela presença das enzimas com atividade biológica.

Role of Synadenium grantii latex proteases in nematicidal activity on Meloidogyne incognita and Panagrellus redivivus

Abstract Synadenium grantii is a Euphorbiaceae plant commonly found in Brazil, known as Janaúba or Leitosinha, whose latex is traditionally used for several purposes. However, it is not known whether the nematicidal action of this plant latex occurs due to the action of proteases. The present work aims to evaluate the nematicidal activity of proteases from Synadenium grantii latex on Meloidogyne incognita and Panagrellus redivivus. S. grantii latex used in the present study was collected from specimens found in Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. The drained latex was collected in Eppendorf microtubes and immediately stored on ice at 4 °C. After this extraction, the latex was frozen (-20 °C) during 2 hours, thawed at room temperature (25 °C) and centrifuged at 10,000 g at 4 °C for 30 minutes to remove larger particles and concentrate the proteases. After the centrifugation, assays of enzymatic activity were performed in order to know in which of the phases the enzymes were found. S. grantii latex presented protease, but no chitinase activity. The results show that there was a significant difference (p 0.01) between the treated and control groups, with 100% mortality of Meloidogyne incognita and 72% average mortality of Panagrellus redivivus. In addition, it was demonstrated that the nematicidal action occurred due to the action of the proteases, since the control was only differentiated from the treatment by the presence of the enzymes with biological activity.

Resumo Synadenium grantii é uma planta Euphorbiaceae comumente encontrada no Brasil, conhecida como Janaúba ou Leitosinha, e tem seu látex usado tradicionalmente para diferentes propósitos. Entretanto, não se conhece se a atividade nematicida da planta ocorre devido à ação de proteases. O presente trabalho tem como objetivo avaliar a atividade nematicida das proteases do látex de Synadenium grantii sobre Meloidogyne incognita e Panagrellus redivivus. O látex de S. grantii utilizado no presente trabalho foi coletado a partir de espécimes encontradas na Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. O látex foi coletado em microtubos Eppendorf e imediatamente armazenado em gelo a 4 °C. Após esta extração, o látex foi congelado (-20 °C) durante 2 horas, descongelado à temperatura ambiente (25 °C) e centrifugado a 10000 g a 4 °C durante 30 minutos para a remoção de partículas e concentração das proteases. Após a centrifugação, foram realizados ensaios de atividade enzimática para saber em qual das fases as enzimas foram encontradas. O látex de S. grantii apresentou atividade de protease, mas nenhuma atividade de quitinase. Os resultados mostram que houve diferença significativa (p 0,01) entre os grupos tratados e controle, com 100% de mortalidade de Meloidogyne incognita e 72% de mortalidade média de Panagrellus redivivus. Além disso, foi demonstrado que a ação nematicida ocorreu devido à ação das proteases, uma vez que o grupo controle só foi diferenciado do tratamento pela presença das enzimas com atividade biológica.

Cytotoxicity of latex and pharmacobotanical study of leaves and stem of Euphorbia umbellata (Janaúba)

AbstractIn southern Brazil, the bottled latex of Synadenium grantii Hook f., Euphorbiaceae, is popularly used as a treatment of all types of cancer. Similarly, Synadenium umbellatum Pax. is used in the central western region of Brazil for the same purpose and in the same manner of use. Both plants are popularly known as janaúba or leitosinha. The objectives of this study were to use pharmacobotanical analysis to verify whether these two species, which are considered to be distinct, are actually the same to determine anatomical markers; to assist in the identification and differentiation of other Euphorbia; and to evaluate the cytotoxic activity of the latex in relation to HeLa and HRT-18 cells. Leaves and stems of the species were collected in Goiânia and Ponta Grossa and were investigated using scanning electron microscopy and optical microscopy techniques. The latex was also collected and analyzed in relation to its cytotoxic effect by employing MTT and NR techniques. The pharmacobotanical study of the specimens in both localities showed that they were the same species, namely Euphorbia umbellata (Pax) Bruyns, which is the scientific nomenclature accepted and confirmed by an expert taxonomist who specializes in Euphorbia. The pharmacobotanical characteristics highlighted in this study can assist in the identification of the taxon and contribute to the control of the quality of this plant drug. The evaluation of the latex in relation to HRT-18 cells demonstrated action after 48 h of experiment. In contrast, in relation to HeLa cells its induced cytotoxicity in all times and a dose-dependent manner. The IC50 values (72 h) observed were 252.58 ± 18.51 µg/ml and 263.42 ± 15.92 µg/ml to MTT experiment and 250.18 ± 19.48 µg/ml and 430.56 ± 19.71 µg/ml to NR experiment for the HeLa and HRT-18 cells, respectively.

Anti-ulcer activity of Synadenium grantii latex

Synadenium grantii Hook f., Euphorbiaceae, is popularly known as leitosinha or janaúba. The diluted latex (18 drops/L of water) is commonly used in the south of Brazil to treat gastric disturbances. This study evaluated phytochemical screening and toxicity using Artemia salina Leach of crude bark extract and also latex. The toxicity and the anti-ulcer activity of S. grantii latex were also tested in rats. Phytochemical results showed presence of tannins, terpenes, unsaponificable substances, coumarins and anthraquinones in the crude bark extract and terpenes in the latex. Gas chromatography-mass spectrometry (GC-MS) analysis demonstrated the presence of diterpene tigliane esters in the latex, identified as 12-deoxyphorbol-13-(2-metilpropionate) and phorbol 12,13,20-triacetate. The toxicity results using A. salina presented CL50 26.58μg/mL and CL50 778.66μg/mL, for the latex and the crude bark extract respectively. The toxicological hepatic parameters of the diluted latex were not different to the control group (p<0.05). The eosinophils cells showed an increase in both the diluted and pure latex groups. The pure latex showed gastric protection of 90% (p<0.05) and the diluted latex showed 6% compared to the negative control. Therefore, our data indicate that S. grantii latex, under research conditions presented gastric protection. Pure latex showed more toxicity than the diluted latex.

Synadenium umbellatum Pax. promotes cell cycle arrest and induces apoptosis in K-562 leukemia cells

Chronic myeloid leukemia (CML) is a clonal myeloproliferative disease that shows apoptosis resistance. The introduction of imatinib mesylate has revolutionized the treatment of CML, but imatinib resistance may develop at any time and inevitably leads to disease progression. Synadenium umbellatum Pax. belongs to the Euphorbiaceae family and is popularly used in Brazil for the treatment of cancer. The cytotoxicity of Euphorbiaceae is associated with the ability of these plants and their bioactive compounds to induce apoptotic tumor cell death. Therefore, we aimed to investigate the cytotoxicity and the mechanisms of death induced by S. umbellatum extract in leukemic cells. S. umbellatum cytotoxicity was evaluated by trypan blue exclusion assay and flow cytometric analysis of the cell cycle; the mechanisms involved in K-562 cell death were investigated by light microscopy and flow cytometry. The results demonstrate that S. umbellatum is cytotoxic to leukemic cells in a concentration-dependent manner. Morphological analysis revealed that S. umbellatum treatment induced K-562 cell death by an apoptotic pathway. Furthermore, data indicate ROS overproduction, alterations in mitochondrial membrane potential, phosphatidylserine externalization and activation of caspase 9. Taken together, the results demonstrate that S. umbellatum extract arrested the cell cycle and triggered apoptosis at several levels in K-562 cells.

A leucemia mielóide crônica (LMC) é uma doença mieloproliferativa clonal, que apresenta resistência à apoptose. A introdução do mesilato de imatinibe revolucionou o tratamento da LMC, porém a resistência ao imatinibe pode ser desenvolvida em qualquer tempo e, inevitavelmente, leva à progressão da doença. Synadenium umbellatum Pax. pertence à família Euphorbiaceae e é usado popularmente no Brasil para o tratamento do câncer. A citotoxicidade das Euphorbiaceae está associada com a capacidade dessas plantas e seus compostos bioativos em induzir apoptose em células tumorais. Portanto, este trabalho teve como objetivo investigar a citotoxicidade e os mecanismos de morte induzidos por S. umbellatum em células leucêmicas. A citotoxicidade de S. umbellatum foi avaliada pelo ensaio de exclusão do azul de tripano e a análise do ciclo celular foi feita por citometria de fluxo. Os mecanismos envolvidos na morte celular das células K-562 foram investigados por microscopia óptica e por citometria de fluxo. Os resultados demonstraram que S. umbellatum é citotóxico para células leucêmicas de uma maneira dependente da concentração. A análise morfológica revelou que o tratamento com S. umbellatum induziu as célula K-562 à morte por via apoptótica. Além disso, os dados indicam aumento de ERO S, alterações no potencial de membrana mitocondrial, externalização da fosfatidilserina e ativação de caspase 9. Em conjunto, os resultados demonstram que S. umbellatum promoveu retenção do ciclo celular das células K-562 e induziu estas células à morte por apoptose.

Letalidade do extrato de synadenium grantii hook. F. (Euphorbiaceae) frente a caramujos biomphalaria glabrata say, 1818 (gastropoda, planorbidae) / Letalidad del extracto de synadenium grantii hook. F. (Euphorbiaceae) contra caracoles biomphalaria glabrata say, 1818 (gastropoda, planorbidae) / Lethality of synadenium grantii hook. F. (Euphorbiaceae) extract to snails biomphalaria glabrata say, 1818 (gastropoda, planorbidae)

No Brasil, o caramujo Biomphalaria glabrata Say, 1818 (Gastropoda, Planorbidae) atua como principal vetor hospedeiro intermediário do helminto trematódeo Schistosoma mansoni (Schistosomatidae), agente etiológico da esquistossomose. O combate deste hospedeiro com substâncias moluscicidas é considerado uma das formas de redução do risco de transmissão da esquistossomose. Na busca por novos produtos naturais para controlar este vetor, o presente trabalho testou os extratos de algumas espécies vegetais exóticas consideradas tóxicas: Allamanda cathartica L. (Apocynaceae), Breynia nivosa (W. Bull) Small (Phyllanthaceae), Croton floribundus Spreng. (Euphorbiaceae), Dieffenbachia picta Schott (Araceae), Euphorbia milii des Mol. var. breonii (Euphorbiaceae), Jatropha curcas L. (Euphorbiaceae), Monstera deliciosa Liebm. (Araceae) e Synadenium grantii Hook. f. (Euphorbiaceae). Apenas a espécie S. grantii apresentou atividade moluscicida (CL50 = 40,0 µg mL-1) promissora, sendo este o primeiro relato desta atividade para espécies do gênero Synadenium.

In Brazil, Biomphalaria glabrata Say, 1818 (Gastropoda, Planorbidae) snail serves as the principal intermediate vector host of Schistosoma mansoni (Schistosomatidae) trematode helminth, the etiologic agent of schistosomiasis. A way of reducing transmission risk of schistosomiasis is the combat of this host with molluscicide substances. In the search for new natural products to control this vector, the present study tested the extracts of some exotic plant species considered toxic: Allamanda cathartica L. (Apocynaceae), Breynia nivosa (W. Bull) Small (Phyllanthaceae), Croton floribundus Spreng. (Euphorbiaceae), Dieffenbachia picta Schott (Araceae), Euphorbia milii des Mol. var. breonii (Euphorbiaceae), Jatropha curcas L. (Euphorbiaceae), Monstera deliciosa Liebm. (Araceae) and Synadenium grantii Hook. f. (Euphorbiaceae). Only the S. grantii species showed molluscicide activity promising (LC50 = 40.0 mg mL-1), and this is the first report of this activity to the genus Synadenium (Euphorbiaceae).

En Brasil, el caracol Biomphalaria glabrata Say, 1818 (Gastropoda, Planorbidae) actúa como principal vector hospedero intermediario del helmintos trematodo Schistosoma mansoni (Schistosomatidae), agente etiológico de la esquistosomiasis. El combate de este hospedero con sustancias molusquicidas es considerado una de las formas de reducción del riesgo de transmisión de la esquistosomiasis. En la búsqueda por nuevos productos naturales para el control de este vector, el presente estudio examinó los extractos de algunas especies exóticas de plantas consideradas tóxicas: Allamanda cathartica L. (Apocynaceae), Breynia nivosa (W. Bull) Small (Phyllanthaceae), Croton floribundus Spreng. (Euphorbiaceae), Dieffenbachia picta Schott (Araceae), Euphorbia milii des Mol. var. breonii (Euphorbiaceae), Jatropha curcas L. (Euphorbiaceae), Monstera deliciosa Liebm. (Araceae) y Synadenium grantii Hook. f. (Euphorbiaceae). Sólo la especie S. grantii presentó actividad molusquicida prometedores (CL50 = 40,0 ug mL-1) promisora, siendo éste el primer relato de esta actividad para especies del género Synadenium.

Eritrodermia secundaria a planta productora de látex (synadenium grantii) / Erythroderma secondary to latex-producing plants (synadenium grantii)

Las euforbiáceas son una familia de plantas que tienen en común la producción de látex. Existen más de 8000 especies.Tienen amplia distribución mundial, y adoptan la forma de árboles, arbustos o hierbas. Pertenecen a esta familia numerosas especies de importancia y, en nuestro país, se emplean con fines industriales y medicinales. También existen plantas ornamentales, cuya atractiva apariencia estimula la curiosidad de los niños, quienes pueden ingerirlas o manipularlas.En el Centro Nacional de Intoxicaciones, las consultas por plantas representan un escaso número (0,2 por ciento) dentro del total de accidentes en pediatría; sin embargo, pueden revestir gravedad.Presentamos el caso de una niña de 4 años, quien manipuló e ingirió parte de una planta de la familia Euforbiáceas(Synadenium grantii).

The Euphorbiaceae family are plants that have in common latex production. There are over 8000 species. They have worldwide distribution. We can find trees, shrubs or herbs. Many importantspecies belong to this family and they are used in our country for industrial and medical purposes. There are alsoornamental plants with attractive appearence that stimulate children curiosity, intake or manipulation.In the National Poison Center consultations for plants represent a small number (0.2%) of the total accidents in children, however they can be serious. We report the case of a 4 years old girl who manipulated andswallowed part of an Euphorbiaceae family plant (Synadenium grantii).

Angiogenic activity of Synadenium umbellatum Pax latex / Atividade angiogênica do látex de Synadenium umbellatum Pax

Synadenium umbellatum Pax, popularly known as "cola-nota", is a medicinal plant that grows in tropical regions. Latex of this plant is used to treat various diseases such as diabetes mellitus, Hansen´s disease, tripanosomiases, leukemia and several malignant tumors. In the present study, the angiogenic activity of S. umbellatum latex was evaluated using the chick embryo chorioallantoic membrane (CAM) assay. Results showed significant increase of the vascular net (p < 0.05) compared to the negative control (H2O). The histological analysis was in accordance with the results obtained. In conclusion, our data indicate that S. umbellatum latex, under the conditions of this research, presented angiogenic effect.

Synadenium umbellatum Pax, popularmente conhecida como "cola-nota", é uma planta medicinal que cresce em regiões tropicais. O látex desta planta tem sido utilizado no tratamento de várias doenças, como diabetes mellitus, hanseníase, tripanossomíases, leucemia e vários tumores malignos. No presente estudo, a atividade angiogênica do látex de S. umbellatum foi avaliada pelo ensaio da membrana corio-alantóide (MCA) de ovo embrionado de galinha. Os resultados mostraram aumento significativo da rede vascular (p < 0.05) em relação ao controle negativo (H2O). A análise histológica está em concordância com os resultados obtidos. Em conclusão, os dados indicaram que, nas condições experimentais deste estudo, o látex de S. umbellatum exibiu efeito angiogênico.

Avaliação da toxicidade aguda e subaguda, em ratos, do extrato etanólico das folhas e do látex de Synadenium umbellatum Pax / Acute and subacute toxicity studies of the latex and of the ethanolic extract of the leaves of Synadenium umbellatum Pax in rats

O uso de plantas medicinais tem sido muito significativo nos últimos anos, sendo incentivado pela Organização Mundial de Saúde (OMS). Synadenium umbellatum Pax, Euphorbiacea (vulgo cola-nota, cancerola, milagrosa) tem o látex usado empiricamente como antitumoral e antiinflamatório. Por existir espécies tóxicas nesta família e visando à segurança no uso de extratos vegetais, tal estudo avaliou a toxicidade pré-clínica do látex e do extrato etanólico das folhas (EEF) de S. umbellatum, por via oral, em ratas Wistar. O estudo seguiu diretrizes do Guideline 423 (toxicidade aguda) e Guideline 407 (toxicidade subaguda) da OECD (Organisation for Economic Cooperation and Development). Na toxicidade aguda do látex e do EEF, não se observou letalidade nem alterações fisiológicas e comportamentais das ratas na dose de 2000 mg/kg, sendo praticamente atóxico. Porém, na análise histopatológica, o látex ocasionou congestão e infiltrado leucocitário nos rins, fígado e pulmões, efeitos não observados com o EEF. Na toxicidade subaguda, doses de 50, 100 e 200 mg/kg de EEF não produziram alterações dose-dependentes significativas nos parâmetros laboratoriais e fisiológicos, nem alterações macroscópicas e histopatológicas nos órgãos das ratas. Contudo, o uso crônico da planta S. umbellatum merece mais estudos.

The use of medicinal plants has been being very significant in the last years, being the use encouraged by WHO. Synadenium umbellatum Pax, Euphorbiacea (popularly known as cola-note, cancerola, miraculous) has the latex used empirically as anti-cancerous and anti-inflammatory. For there being toxic species in this family and aiming at the safety in the use of vegetable extracts, such study evaluated the pre-clinical toxicity of the latex and of the ethanolic extract of the leaves (EEL) of S. umbellatum, administrated by oral route, in Wistar female rats. The study followed OECD's Guidelines for test of acute toxicity (Guideline 423) and for subacute toxicity (Guideline 407). In the acute toxicity of latex and EEL, behavioral and physiological alterations were not observed neither animal's death in the dose level of 2,000 mg/kg. However, the latex caused congestion and leukocytes infiltration of the kidneys, liver and lungs, effects not observed with EEL. In the subacute toxicity, dose levels of 50, 100 and 200 mg/kg of EEL did not produced significant dose-dependent alterations in the lab results and no physiologic, macroscopic and hystopathological alterations. EEL of S. umbellatum is practically poisonless in acute exposure; already the latex can cause hystological damages. The chronic use of S. umbellatum needs more specific studies.

Synadenium umbellatum: citotoxicidade e danos ao DNA de células da medula óssea de camundongos / Synadenium umbellatum: cytotoxicity and DNA damage to bone marrow cells from mice

No presente estudo investigamos o potencial citotóxico e mutagênico, in vitro e in vivo, respectivamente, do extrato etanólico de Synadenium umbellatum (EESU) sobre células da medula óssea de camundongos. A citotoxicidade in vitro foi avaliada por meio da exposição de células da medula óssea de animais normais a diferentes concentrações (40-0,312 mg/mL) do EESU, por 12, 24 ou 48 h, utilizando os testes de redução do MTT e o de exclusão do azul de tripano. O ensaio de micronúcleo foi realizado para investigar potenciais efeitos mutagênicos do EESU (10, 25 ou 50 mg/kg/dia) sobre a medula óssea de camundongos. Os animais foram expostos a uma única dose, por via oral, e 24 h após à exposição, sacrificados para realização do estudo (n=5/grupo). Os resultados obtidos demonstraram que o EESU possui potencial efeito citotóxico e mutagênico, de forma dose-dependente, sobre as células da medula óssea de camundongos, in vitro e in vivo, respectivamente. Maiores estudos são necessários para expandir o conhecimento acerca do potencial toxicológico/farmacológico do EESU.

In the present study we investigated, in vitro and in vivo, the cytotoxic and mutagenic potential of the Synadenium umbellatum ethanolic extract (SUEE) on the bone marrow cells of mice. In vitro cytotoxicity was assayed incubating bone marrow cells with different concentrations (40-0.312 mg/mL) of SUEE for 12, 24 or 48 h using the MTT tetrazolium reduction test and the trypan blue exclusion test. The micronuclei assay was performed to determine mutagenic effects of mice orally exposed to different doses of SUEE (10, 25 or 50 mg/kg/day) in 24 h. Our results demonstrated that SUEE has, in a dose-dependent manner, potential cytotoxic and mutagenic effects on the bone marrow cells of mice. Further studies are needed to expand the knowledge of the toxicological/pharmacological potential of the SUEE.

Protective effect of lectin from Synadenium carinatum on Leishmania amazonensis infection in BALB/c mice

The protective effect of the Synadenium carinatum latex lectin (ScLL), and the possibility of using it as an adjuvant in murine model of vaccination against American cutaneous leishmaniasis, were evaluated. BALB/c mice were immunized with the lectin ScLL (10, 50, 100 microgram/animal) separately or in association with the soluble Leishmania amazonensis antigen (SLA). After a challenge infection with 10(6) promastigotes, the injury progression was monitored weekly by measuring the footpad swelling for 10 weeks. ScLL appeared to be capable of conferring partial protection to the animals, being most evident when ScLL was used in concentrations of 50 and 100 microgram/animal. Also the parasite load in the interior of macrophages showed significant reduction (61.7%) when compared to the control group. With regard to the cellular response, ScLL 50 and 100 microgram/animal stimulated the delayed-type hypersensitivity (DTH) reaction significantly (P < 0.05) higher than SLA or SLA plus ScLL 10 weeks after the challenge infection. The detection of high levels of IgG2a and the expression of mRNA cytokines, such as IFN-gamma, IL-12, and TNF-alpha (Th1 profiles), corroborated the protective role of this lectin against cutaneous leishmaniasis. This is the first report of the ScLL effect on leishmaniasis and shows a promising role for ScLL to be explored in other experimental models for treatment of leishmaniasis.

Purification and characterisation of a carboxylesterase from the latex of Synadenium grantii Hook,'f'.

The latex of Synadenium grantii was found to contain esterolytic activity. Polyacrylamide gel electrophoretic study coupled with substrate and inhibitor specificity studies revealed the presence of multiple forms of carboxylesterases and cholinesterases in the latex. One of the carboxylesterases of the latex was purified by acetone fractionation, carboxymethyl- Sephadex chromatography and Sepharose-6B gel filtration. The homogeneity of the enzyme was established by polyacrylamide gel electrophoresis, isoelectric focussing and sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The enzyme consists of a single polypeptide chain with a molecular weight of 14,000. The amino acid analysis of the purified enzyme revealed that it contained a greater number of neutral and acidic, compared to basic amino acid residues. The isoelectric pH of the enzyme was found to be 4·0. The enzyme was a glycoprotein as revealed by periodic acid Schiff-staining technique. Studies with different organophosphate and carbamate inhibitors showed that this enzyme was sensitive to organophosphates. The product inhibition studies with this enzyme showed linear competitive inhibition with acetate and linear non-competitive inhibition with 1- naphthol.