As the living cytoplasm of laticiferous cells, Hevea brasiliensis latex is a rich blend of organic substances that include a mélange of proteins. A small number of these proteins have given rise to the problem of latex allergy. The salient characteristics of H. brasiliensis latex allergens that are recognized by the International Union of Immunological Societies (IUIS) are reviewed. These are the proteins associated with the rubber particles, the cytosolic C-serum proteins and the B-serum proteins that originate mainly from the lutoids. Procedures for the isolation and purification of latex allergens are discussed, from latex collection in the field to various preparative approaches adopted in the laboratory. As interest in recombinant latex allergens increases, there is a need to validate recombinant proteins to ascertain equivalence with their native counterparts when used in immunological studies, diagnostics, and immunotherapy.
Natural rubber from hevea brasiliensis trees (Thailand, RRIM 600 clone) of different age (8, 20, and 35 years) were characterized by size exclusion chromatography coupled with online viscometry according to their distribution of molar mass and branching index at a temperature of 70 degrees C using cyclohexane as solvent. Washing with an aqueous solution of sodium dodecylsulfate and subsequent saponification purified the natural rubber samples. With this procedure physical branching points caused by phospholipids, proteins and hydrophobic terminal units, mainly fatty acids, of the natural rubber (cis-1,4-polyisoprene) molecule, could be removed leading to completely soluble polymer samples. All samples investigated possess a very broad (10 to 50,000 kg/mol) and distinct bimodal molar mass distribution. With increasing age the peak area in the low molar mass region decreases favoring the peak area in the high molar mass region. By plotting the branching index as a function of the both, the molar mass and the age of the trees.
The flame retardancy of medium density fiberboard (MDF) made from mixture of rubberwood fibers and recycled old corrugated containers was studied. Aluminum trihydroxide (ATH) was used as a fire retardant additive and mixed with the fibers to manufacture experimental MDF panels using wet process. Phenol formaldehyde (PF) resin in liquid, 2% based on oven dry weight of fibers, was used along with 0%, 10%, 15% and 20% of ATH. The flame retardant test was done using the limiting oxygen index (LOI) test. The other properties investigated include internal bond strength, thickness swelling and water absorption. The results showed that ATH loading increased as the LOI of MDF increased. This demonstrated that ATH could improved the fire retardant property of MDF at sufficient loading. An increase in concentration of ATH showed an increase in the IB values of MDF made without resin. MDF panels made without resin showed a progressive increase in internal bond as the composition of recycled old corrugated containers fiber increased. Addition of resin improved internal bond strength and reduced thickness swelling, and water absorption. Thickness swelling of panel increased as the composition of recycled old corrugated containers fiber increased. Scanning electron microscopy (SEM) and energy dispersive X-ray analysis (EDAX) showed that there is indication of ATH and resin filling the void space in between fibers.
Hev b 4 is an allergenic natural rubber latex (NRL) protein complex that is reactive in skin prick tests and in vitro immunoassays. On SDS-polyacrylamide gel electrophoresis (SDS-PAGE), Hev b 4 is discerned predominantly at 53-55 kDa together with a 57 kDa minor component previously identified as a cyanogenic glucosidase. Of the 13 NRL allergens recognized by the International Union of Immunological Societies, the 53-55 kDa Hev b 4 major protein is the only candidate that lacks complete cDNA and protein sequence information.
We sought to clone the transcript encoding the Hev b 4 major protein, and characterize the native protein and its recombinant form in relation to IgE binding.
The 5'/3' rapid amplification of cDNA ends method was employed to obtain the complete cDNA of the Hev b 4 major protein. A recombinant form of the protein was over-expressed in Escherichia coli. The native Hev b 4 major protein was deglycosylated by trifluoromethane sulphonic acid. Western immunoblots of the native, deglycosylated and recombinant proteins were performed using both polyclonal antibodies and sera from latex-allergic patients.
The cDNA encoding the Hev b 4 major protein was cloned. Its open reading frame matched lecithinases in the conserved domain database and contained 10 predicted glycosylation sites. Detection of glycans on the Hev b 4 lecithinase homologue confirmed it to be a glycoprotein. The deglycosylated lecithinase homologue was discerned at 40 kDa on SDS-PAGE, this being comparable to the 38.53 kDa mass predicted by its cDNA. Deglycosylation of the lecithinase homologue resulted in the loss of IgE recognition, although reactivity to polyclonal rabbit anti-Hev b 4 was retained. IgE from latex-allergic patients also failed to recognize the non-glycosylated E. coli recombinant lecithinase homologue.
The IgE epitopes of the Hev b 4 lecithinase homologue reside mainly in its carbohydrate moiety, which also account for the discrepancy between the observed molecular weight of the protein and the value calculated from its cDNA.
A comparative study on solid substrate fermentation (SSF) of sago 'hampas', oil palm frond parenchyma tissue (OPFPt) and rubberwood sawdust with Pycnoporus sanguineus for laccase production was carried out. Optimal mycelial growth of Pyc. sanguineus was observed on all the substrates studied over a 21 days time-course fermentation. Laccase productivity was highest during degradation of sago 'hampas' and OPFPt and a range from 7.5 to 7.6 U/g substrate on the 11th day of fermentation compared to degradation of rubberwood sawdust with a maximum laccase productivity of 5.7 U/g substrate on day 11 of SSF. Further optimization of laccase production was done by varying the inoculum age, density and nitrogen supplementation. SSF of OPFPt by Pyc. sanguineus gave maximum productivity of laccase of 46.5 U/g substrate on day 6 of fermentation with a 30% (w/w) of 4 weeks old inoculum and 0.92% nitrogen in the form of urea supplemented in the substrate. The extraction of laccase was also optimized in this study. Recovery of laccase was fourfold higher at 30.6 U/g substrate on day 10 of SSF using unadjusted tap water at pH 8.0 as extraction medium at 25+/-2 degrees C compared to laccase recovery of 7.46 U/g substrate using sodium acetate buffer at pH 4.8 at 4 degrees C. Further optimization showed that laccase recovery was increased by 50% with a value of 46.5 U/g substrate on day 10 of SSF when the extraction medium was tap water adjusted to pH 5.0 at 25+/-2 degrees C.
Hevea brasiliensis latex serum is commonly used as the in vivo and in vitro reference antigen for latex allergy diagnosis as it contains the full complement of latex allergens.
This study quantifies the concentrations of the significant allergens in latex serum and examines its suitability as an antigen source in latex allergy diagnosis and immunotherapy.
The serum phase was extracted from centrifuged latex that was repeatedly freeze-thawed or glycerinated. Quantitation of latex allergens was performed by two-site immunoenzymetric assays. The abundance of RNA transcripts of the latex allergens was estimated from the number of their clones in an Expressed Sequence Tags library.
The latex allergens, Hev b 1, 2, 3, 4, 5, 6, 7 and 13, were detected in freeze-thawed and glycerinated latex serum at levels ranging from 75 (Hev b 6) to 0.06 nmol/mg total proteins (Hev b 4). Hev b 6 content in the latex was up to a thousand times higher than the other seven latex allergens, depending on source and/or preparation procedure. Allergen concentration was reflected in the abundance of mRNA transcripts. When used as the antigen, latex serum may bias the outcome of latex allergy diagnostic tests towards sensitization to Hev b 6. Tests that make use of latex serum may fail to detect latex-specific IgE reactivity in subjects who are sensitized only to allergens that are present at low concentrations.
Latex allergy diagnostics and immunotherapy that use whole latex serum as the antigen source may not be optimal because of the marked imbalance of its constituent allergens.
How tropical trees flower synchronously near the equator in the absence of significant day length variation or other meteorological cues has long been a puzzle. The rubber tree (Hevea brasiliensis) is used as a model to investigate this phenomenon. The annual cycle of solar radiation intensity is shown to correspond closely with the flowering of the rubber tree planted near the equator and in the subtropics. Unlike in temperate regions, where incoming solar radiation (insolation) is dependent on both day length and radiation intensity, insolation at the equator is due entirely to the latter. Insolation at the upper atmosphere peaks twice a year during the spring and autumn equinoxes, but the actual solar radiation that reaches the ground is attenuated to varying extents in different localities. The rubber tree shows one or two flowering seasons a year (with major and minor seasons in the latter) in accordance with the solar radiation intensity received. High solar radiation intensity, and in particular bright sunshine (as distinct from prolonged diffuse radiation), induces synchronous anthesis and blooming in Hevea around the time of the equinoxes. The same mechanism may be operational in other tropical tree species.
Hevea brasiliensis is the most widely cultivated species for commercial production of natural rubber (cis-polyisoprene). In this study, 10,040 expressed sequence tags (ESTs) were generated from the latex of the rubber tree, which represents the cytoplasmic content of a single cell type, in order to analyse the latex transcription profile with emphasis on rubber biosynthesis-related genes. A total of 3,441 unique transcripts (UTs) were obtained after quality editing and assembly of EST sequences. Functional classification of UTs according to the Gene Ontology convention showed that 73.8% were related to genes of unknown function. Among highly expressed ESTs, a significant proportion encoded proteins related to rubber biosynthesis and stress or defence responses. Sequences encoding rubber particle membrane proteins (RPMPs) belonging to three protein families accounted for 12% of the ESTs. Characterization of these ESTs revealed nine RPMP variants (7.9-27 kDa) including the 14 kDa REF (rubber elongation factor) and 22 kDa SRPP (small rubber particle protein). The expression of multiple RPMP isoforms in latex was shown using antibodies against REF and SRPP. Both EST and quantitative reverse transcription-PCR (QRT-PCR) analyses demonstrated REF and SRPP to be the most abundant transcripts in latex. Besides rubber biosynthesis, comparative sequence analysis showed that the RPMPs are highly similar to sequences in the plant kingdom having stress-related functions. Implications of the RPMP function in cis-polyisoprene biosynthesis in the context of transcript abundance and differential gene expression are discussed.
Morphological features and Inter Simple Sequence Repeat (ISSR) polymorphism were employed to analyse 21 Corynespora cassiicola isolates obtained from a number of Hevea clones grown in rubber plantations in Malaysia. The C. cassiicola isolates used in this study were collected from several states in Malaysia from 1998 to 2005. The morphology of the isolates was characteristic of that previously described for C. cassiicola. Variations in colony and conidial morphology were observed not only among isolates but also within a single isolate with no inclination to either clonal or geographical origin of the isolates. ISSR analysis delineated the isolates into two distinct clusters. The dendrogram created from UPGMA analysis based on Nei and Li's coefficient (calculated from the binary matrix data of 106 amplified DNA bands generated from 8 ISSR primers) showed that cluster 1 encompasses 12 isolates from the states of Johor and Selangor (this cluster was further split into 2 sub clusters (1A, 1B), sub cluster 1B consists of a unique isolate, CKT05D); while cluster 2 comprises of 9 isolates that were obtained from the other states. Detached leaf assay performed on selected Hevea clones showed that the pathogenicity of representative isolates from cluster 1 (with the exception of CKT05D) resembled that of race 1; and isolates in cluster 2 showed pathogenicity similar to race 2 of the fungus that was previously identified in Malaysia. The isolate CKT05D from sub cluster 1B showed pathogenicity dissimilar to either race 1 or race 2.
The efficiency of sodium hydroxide treated rubber (Hevea brasiliensis) leaves powder (NHBL) for removing copper ions from aqueous solutions has been investigated. The effects of physicochemical parameters on biosorption capacities such as stirring speed, pH, biosorbent dose, initial concentrations of copper, and ionic strength were studied. The biosorption capacities of NHBL increased with increase in pH, stirring speed and copper concentration but decreased with increase in biosorbent dose and ionic strength. The isotherm study indicated that NHBL fitted well with Langmuir model compared to Freundlich and Dubinin-Radushkevich models. The maximum biosorption capacity determined from Langmuir isotherm was 14.97 mg/g at 27 degrees C. The kinetic study revealed that pseudosecond order model fitted well the kinetic data, while Boyd kinetic model indicated that film diffusion was the main rate determining step in biosorption process. Based on surface area analysis, NHBL has low surface area and categorized as macroporous. Fourier transform infrared (FT-IR) analyses revealed that hydroxyl, carboxyl, and amino are the main functional groups involved in the binding of copper ions. Complexation was one of the main mechanisms for the removal of copper ions as indicated by FT-IR spectra. Ion exchange was another possible mechanism since the ratio of adsorbed cations (Cu2+ and H+) to the released cations (Na+, Ca2+, and Mg2+) from NHBL was almost unity. Copper ions bound on NHBL were able to be desorbed at > 99% using 0.05 mol/L HCl, 0.01 mol/L HNO3, and 0.01 mol/L EDTA solutions.
Dielectric properties of natural rubber Hevea brasiliensis latex were measured at frequencies 0.2 to 20 GHz, at temperatures of 2, 15, 25, 35, and 50oC and around 30-98% moisture content. Measurements were done using open-ended coaxial line sensor and automated network analyzer. As expected, results showed that dielectric constant increased with increasing moisture. From 0.2 to 2.6 GHz, the losses were governed by conductive losses but for frequencies greater than 2.6 GHz, these were mainly due to dipolar losses. The former is due to conducting phases in hevea latex, while the latter is mainly governed by the orientation of water molecules. The results were analyzed at 2.6, 10, and 18 GHz, respectively. These were then compared with the values predicted by the dielectric mixture equations recommended by Weiner, Bruggeman and Kraszewski. All the measured values were found to be within the Weiner’s boundaries and close to the upper limit of Weiner’s model. It is also close to the predicted values of Bruggeman’s model with a/b = 0.1. All the models including Kraszewski are suitable for predicting the dielectric properties of hevea latex for frequencies 2.6 to 18 GHz, moisture content 30 to 98% and temperatures 2 to 50oC.
Rubber leaf powder (an agricultural waste) was treated with potassium permanganate followed by sodium carbonate and its performance in the removal of Pb(II) ions from aqueous solution was evaluated. The interactions between Pb(II) ions and functional groups on the adsorbent surface were confirmed by Fourier transform infrared (FT-IR) spectroscopy, scanning electron microscopy (SEM) coupled with X-ray energy dispersive spectroscopy (EDX). The effects of several important parameters which can affect adsorption capacity such as pH, adsorbent dosage, initial lead concentration and contact time were studied. The optimum pH range for lead adsorption was 4-5. Even at very low adsorbent dosage of 0.02 g, almost 100% of Pb(II) ions (23 mg/L) could be removed. The adsorption capacity was also dependent on lead concentration and contact time, and relatively a short period of time (60-90 min) was required to reach equilibrium. The equilibrium data were analyzed with Langmuir, Freundlich and Dubinin-Radushkevich isotherms. Based on Langmuir model, the maximum adsorption capacity of lead was 95.3 mg/g. Three kinetic models including pseudo first-order, pseudo second-order and Boyd were used to analyze the lead adsorption process, and the results showed that the pseudo second-order fitted well with correlation coefficients greater than 0.99.
Hevea brasiliensis extracts could potentially be employed as a relatively low cost resource for various anti-fungal activities due to the simplicity of the extract preparation and its abundance especially in the tropical region. Latex B-serum was reported to have anti-cancer property and its specificity in anti-fungal property has not been elucidated. The present study was conducted to determine the anti-fungal activity of Hevea latex B-serum against Candida (C.) albicans (a rounded cell fungus) and Aspergillus (A.) niger (a filamentous fungus).
The cytosolic mevalonate (MVA) pathway in Hevea brasiliensis latex is the conventionally accepted pathway which provides isopentenyl diphosphate (IPP) for cis-polyisoprene (rubber) biosynthesis. However, the plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway may be an alternative source of IPP since its more recent discovery in plants. Quantitative RT-PCR (qRT-PCR) expression profiles of genes from both pathways in latex showed that subcellular compartmentalization of IPP for cis-polyisoprene synthesis is related to the degree of plastidic carotenoid synthesis. From this, the occurrence of two schemes of IPP partitioning and utilization within one species is proposed whereby the supply of IPP for cis-polyisoprene from the MEP pathway is related to carotenoid production in latex. Subsequently, a set of latex unique gene transcripts was sequenced and assembled and they were then mapped to IPP-requiring pathways. Up to eight such pathways, including cis-polyisoprene biosynthesis, were identified. Our findings on pre- and post-IPP metabolic routes form an important aspect of a pathway knowledge-driven approach to enhancing cis-polyisoprene biosynthesis in transgenic rubber trees.
The rubber tree (Hevea brasiliensis) extracts are becoming increasingly visible in pharmaceutical and therapeutical research. The present study is aimed at examining the specific anti-proliferation property of H. brasiliensis latex B-serum sub-fractions against human breast cancer epithelial cell lines MCF-7 and MDA-MB231. The results showed that the latex whole B-serum and DBP sub-fraction exerted a specific anti-proliferation activity against cancer-origin cells MDA-MB231 but had little effect on non-cancer-origin cells. On the other hand, the anti-proliferative activity was diminished in the pre-heated B-serum fractions. With the low toxicity that the B-serum demonstrated previously in Brine Shrimp Lethality Test (BSLT), the present results suggest the potential use of the B-serum sub-fractions in cancer treatment.
Latex from Hevea brasiliensis (natural rubber tree primarily cultivated for its rubber particles) has no known primary metabolic function, although its biological role is as a plant defence system. The present study has evaluated specific anti-proliferative effects of latex whole C-serum and its subfractions, on human cancer cell lines.
Effects of cultural practice under different habitats, of well-managed monoculture plantation and growing wild under rubber trees, were studied in Aquilaria malaccensis (Karas) leaves. This study was carried out on Karas growing in these two habitats each from Lipis, Pahang and Sepang, Selangor areas in Malaysia; under the control and induced treatments. The parameters studied include wet and dry weight of 50 matured leaves, iron and zinc elemental contents in leaf, iron and zinc uptakes from soil, and leaf and soil moisture contents. Iron and zinc were analysed in Karas leaves and soil by using Instrumental Neutron Activation Analysis (INAA) technique.
The lipid fraction of rubber (Hevea brasiliensis (kunth. Muell)) seed was extracted and analyzed for toxicological effect. The toxicological compound such as linamarin in rubber seed oil (RSO) extracted using different solvents, such as hexane (RSOh), mixture of chloroform + methanol (RSOchl+mth) and ethanol (RSOeth) were also studied. Various methods analysis such as Fourier transforms infrared spectroscopy (FTIR) and colorimetric methods were carried out to determine the present of such compounds.