In Vitro Cytotoxicity and Anti-inflammatory Cytokinine Activity Study of Three Isolated Novel Compounds of Prismatomeris glabra.

OBJECTIVES: The aim of the present study was to isolate and evaluate cytotoxicity and anti-inflammatory activities of new novel compounds isolated from Prismatomeris glabra. MATERIALS AND METHODS: Dried root of P. glabra was extracted under reflux with methyl alcohol, fractionated through the vacuum liquid chromatography technique, and evaporated and then purified the compounds using column chromatography and preparative thin-layer chromatography. THP-1 cells were treated with amentoflavone, 5,7,4'-hydroxyflavonoid, and stigmasterol with various concentrations (0-30 µg/mL) and then incubated with MTS reagent for 2h. Treatment was done for 24, 48, and 72h. Then, effects of these compounds were also tested on PGE2, TNF-α, and IL-6 expression in human THP-1-derived macrophage cells for 24h. RESULTS: Three new compounds such as amentoflavone, 5,7,4'-hydroxyflavonoid, and stigmasterol were isolated. After 24h of incubation, a significant decrease in cell viability was reported with IC50 values of amentoflavone, 5,7,4'- hydroxyflavonoid, and stigmasterol (21 µg/mL ≡ 38 M), (18 µg/mL ≡ 66 M) and (20 µg/mL ≡ 48.5 M), respectively. Whereas for 48 and 72h treatment showed a less decreased cell viability compared with 24h treatment. These compounds also showed a significant reduction in the production of TNF-α, IL-6, and PGE2 in a dose-dependent manner. CONCLUSIONS: The isolated new compounds showed significant cytotoxicity and anti-inflammatory effects.

Asymptomatic neurotoxicity of amyloid ß-peptides (Aß1-42 and Aß25-35) on mouse embryonic stem cell-derived neural cells.

One of the strategies in the establishment of in vitro oxidative stress models for neurodegenerative diseases, such as Alzheimer's disease (AD), is to induce neurotoxicity by amyloid beta (Aß) peptides in suitable neural cells. Presently, data on the neurotoxicity of Aß in neural cells differentiated from stem cells are limited. In this study, we attempted to induce oxidative stress in transgenic 46C mouse embryonic stem cell-derived neurons via treatment with Aß peptides (Aß1-42 and Aß25-35). 46C neural cells were generated by promoting the formation of multicellular aggregates, embryoid bodies in the absence of leukemia inhibitory factor, followed by the addition of all-trans retinoic acid as the neural inducer. Mature neuronal cells were exposed to different concentrations of Aß1-42 and Aß25-35 for 24 h. Morphological changes, cell viability, and intracellular reactive oxygen species (ROS) production were assessed. We found that 100 µM Aß1-42 and 50 µM Aß25-35 only promoted 40% and 10%, respectively, of cell injury and death in the 46C-derived neuronal cells. Interestingly, treatment with each of the Aß peptides resulted in a significant increase of intracellular ROS activity, as compared to untreated neurons. These findings indicate the potential of using neurons derived from stem cells and Aß peptides in generating oxidative stress for the establishment of an in vitro AD model that could be useful for drug screening and natural product studies.

Orthosiphon stamineus Benth. is an Outstanding Food Medicine: Review of Phytochemical and Pharmacological Activities.

Orthosiphon stamineus Benth. (Lamiaceae) is a valued medicinal plant in traditional folk medicine. Many pharmacological studies have demonstrated the ability of this plant to exhibit antimicrobial, antioxidant, hepatoprotection, antigenotoxic, antiplasmodial, cytotoxic, cardioactive, antidiabetic, anti-inflammatory activies. This review is a comprehensive summary of the presently available chemical, pharmacological investigations as well as the traditional and therapeutic uses of this plant. Important and different experimental data have been addressed along with a review of all phytochemicals identified in this plant, including flavonoids, terpenoids, and essential oils. O. stamineus has wide traditional and pharmacological uses in various pathophysiological conditions. Therefore, it is an attractive subject for further experimental and clinical investigations.

Understanding the mode-of-action of Cassia auriculata via in silico and in vivo studies towards validating it as a long term therapy for type II diabetes.

ETHNOPHARMACOLOGICAL RELEVANCE: Cassia auriculata (CA) is used as an antidiabetic therapy in Ayurvedic and Siddha practice. This study aimed to understand the mode-of-action of CA via combined cheminformatics and in vivo biological analysis. In particular, the effect of 10 polyphenolic constituents of CA in modulating insulin and immunoprotective pathways were studied. MATERIALS AND METHODS: In silico target prediction was first employed to predict the probability of the polyphenols interacting with key protein targets related to insulin signalling, based on a model trained on known bioactivity data and chemical similarity considerations. Next, CA was investigated in in vivo studies where induced type 2 diabetic rats were treated with CA for 28 days and the expression levels of genes regulating insulin signalling pathway, glucose transporters of hepatic (GLUT2) and muscular (GLUT4) tissue, insulin receptor substrate (IRS), phosphorylated insulin receptor (AKT), gluconeogenesis (G6PC and PCK-1), along with inflammatory mediators genes (NF-κB, IL-6, IFN-γ and TNF-α) and peroxisome proliferators-activated receptor gamma (PPAR-γ) were determined by qPCR. RESULTS: In silico analysis shows that several of the top 20 enriched targets predicted for the constituents of CA are involved in insulin signalling pathways e.g. PTPN1, PCK-α, AKT2, PI3K-γ. Some of the predictions were supported by scientific literature such as the prediction of PI3K for epigallocatechin gallate. Based on the in silico and in vivo findings, we hypothesized that CA may enhance glucose uptake and glucose transporter expressions via the IRS signalling pathway. This is based on AKT2 and PI3K-γ being listed in the top 20 enriched targets. In vivo analysis shows significant increase in the expression of IRS, AKT, GLUT2 and GLUT4. CA may also affect the PPAR-γ signalling pathway. This is based on the CA-treated groups showing significant activation of PPAR-γ in the liver compared to control. PPAR-γ was predicted by the in silico target prediction with high normalisation rate although it was not in the top 20 most enriched targets. CA may also be involved in the gluconeogenesis and glycogenolysis in the liver based on the downregulation of G6PC and PCK-1 genes seen in CA-treated groups. In addition, CA-treated groups also showed decreased cholesterol, triglyceride, glucose, CRP and Hb1Ac levels, and increased insulin and C-peptide levels. These findings demonstrate the insulin secretagogue and sensitizer effect of CA. CONCLUSION: Based on both an in silico and in vivo analysis, we propose here that CA mediates glucose/lipid metabolism via the PI3K signalling pathway, and influence AKT thereby causing insulin secretion and insulin sensitivity in peripheral tissues. CA enhances glucose uptake and expression of glucose transporters in particular via the upregulation of GLUT2 and GLUT4. Thus, based on its ability to modulate immunometabolic pathways, CA appears as an attractive long term therapy for T2DM even at relatively low doses.

Mesenchymal Stem Cell-Based Therapies against Podocyte Damage in Diabetic Nephropathy.

Injury to podocytes is an early event in diabetic nephropathy leading to proteinuria with possible progression to end-stage renal failure. The podocytes are unique and highly specialized cells that cover the outer layer of kidney ultra-filtration barrier and play an important role in glomerular function. In the past few decades, adult stem cells, such as mesenchymal stem cells (MSCs) with a regenerative and differentiative capacity have been extensively used in cell-based therapies. In addition to their capability for regeneration and differentiation, MSCs contributes to their milieu by paracrine action of a series of growth factors via antiapoptotic, mitogenic and other cytokine actions that actively participate in treatment of podocyte damage through prevention of podocyte effacement, detachment and apoptosis. It is hoped that novel stem cell-based therapies will be developed in the future to prevent podocyte injury, thereby reducing the burden of kidney disease.

Corrigendum to "Stevioside from Stevia rebaudiana Bertoni Increases Insulin Sensitivity in 3T3-L1 Adipocytes".

[This corrects the article DOI: 10.1155/2013/938081.].

Animal Model of Gestational Diabetes Mellitus with Pathophysiological Resemblance to the Human Condition Induced by Multiple Factors (Nutritional, Pharmacological, and Stress) in Rats.

This study attempts to develop an experimental gestational diabetes mellitus (GDM) animal model in female Sprague-Dawley rats. Rats were fed with high fat sucrose diet, impregnated, and induced with Streptozotocin and Nicotinamide on gestational day 0 (D0). Sleeping patterns of the rats were also manipulated to induce stress, a lifestyle factor that contributes to GDM. Rats were tested for glycemic parameters (glucose, C-peptide, and insulin), lipid profiles (total cholesterol, triglycerides, HDL, and LDL), genes affecting insulin signaling (IRS-2, AKT-1, and PCK-1), glucose transporters (GLUT-2 and GLUT-4), proinflammatory cytokines (IL-6, TNF-α), and antioxidants (SOD, CAT, and GPX) on D6 and D21. GDM rats showed possible insulin resistance as evidenced by high expression of proinflammatory cytokines, PCK-1 and CRP. Furthermore, low levels of IRS-2 and AKT-1 genes and downregulation of GLUT-4 from the initial to final phases indicate possible defect of insulin signaling. GDM rats also showed an impairment of antioxidant status and a hyperlipidemic state. Additionally, GDM rats exhibited significantly higher body weight and blood glucose and lower plasma insulin level and C-peptide than control. Based on the findings outlined, the current GDM animal model closely replicates the disease state in human and can serve as a reference for future investigations.

Impact of chitosan composites and chitosan nanoparticle composites on various drug delivery systems: A review.

Chitosan is a promising biopolymer for drug delivery systems. Because of its beneficial properties, chitosan is widely used in biomedical and pharmaceutical fields. In this review, we summarize the physicochemical and drug delivery properties of chitosan, selected studies on utilization of chitosan and chitosan-based nanoparticle composites in various drug delivery systems, and selected studies on the application of chitosan films in both drug delivery and wound healing. Chitosan is considered the most important polysaccharide for various drug delivery purposes because of its cationic character and primary amino groups, which are responsible for its many properties such as mucoadhesion, controlled drug release, transfection, in situ gelation, and efflux pump inhibitory properties and permeation enhancement. This review can enhance our understanding of drug delivery systems particularly in cases where chitosan drug-loaded nanoparticles are applied.

Apoptosis Induction by Polygonum minus is related to antioxidant capacity, alterations in expression of apoptotic-related genes, and S-phase cell cycle arrest in HepG2 cell line.

Polygonum minus (Polygonaceae) is a medicinal herb distributed throughout eastern Asia. The present study investigated antiproliferative effect of P. minus and its possible mechanisms. Four extracts (petroleum ether, methanol, ethyl acetate, and water) were prepared by cold maceration. Extracts were subjected to phytochemical screening, antioxidant, and antiproliferative assays; the most bioactive was fractionated using vacuum liquid chromatography into seven fractions (F1-F7). Antioxidant activity was measured via total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) assays. Antiproliferative activity was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Most active fraction was tested for apoptosis induction and cell cycle arrest in HepG2 cells using flow cytometry and confocal microscopy. Apoptotic-related gene expression was studied by RT-PCR. Ethyl acetate extract was bioactive in initial assays. Its fraction, F7, exhibited highest antioxidant capacity (TPC; 113.16 ± 6.2 mg GAE/g extract, DPPH; EC50: 30.5 ± 3.2 µg/mL, FRAP; 1169 ± 20.3 µmol Fe (II)/mg extract) and selective antiproliferative effect (IC50: 25.75 ± 1.5 µg/mL). F7 induced apoptosis in concentration- and time-dependent manner and caused cell cycle arrest at S-phase. Upregulation of proapoptotic genes (Bax, p53, and caspase-3) and downregulation of antiapoptotic gene, Bcl-2, were observed. In conclusion, F7 was antiproliferative to HepG2 cells by inducing apoptosis, cell cycle arrest, and via antioxidative effects.

Potential Roles of Stevia rebaudiana Bertoni in Abrogating Insulin Resistance and Diabetes: A Review.

Insulin resistance is a key factor in metabolic disorders like hyperglycemia and hyperinsulinemia, which are promoted by obesity and may later lead to Type II diabetes mellitus. In recent years, researchers have identified links between insulin resistance and many noncommunicable illnesses other than diabetes. Hence, studying insulin resistance is of particular importance in unravelling the pathways employed by such diseases. In this review, mechanisms involving free fatty acids, adipocytokines such as TNF α and PPAR γ and serine kinases like JNK and IKK ß , asserted to be responsible in the development of insulin resistance, will be discussed. Suggested mechanisms for actions in normal and disrupted states were also visualised in several manually constructed diagrams to capture an overall view of the insulin-signalling pathway and its related components. The underlying constituents of medicinal significance found in the Stevia rebaudiana Bertoni plant (among other plants that potentiate antihyperglycemic activities) were explored in further depth. Understanding these factors and their mechanisms may be essential for comprehending the progression of insulin resistance towards the development of diabetes mellitus.

Antioxidant properties of phenolic Schiff bases: structure-activity relationship and mechanism of action.

Phenolic Schiff bases are known for their diverse biological activities and ability to scavenge free radicals. To elucidate (1) the structure-antioxidant activity relationship of a series of thirty synthetic derivatives of 2-methoxybezohydrazide phenolic Schiff bases and (2) to determine the major mechanism involved in free radical scavenging, we used density functional theory calculations (B3P86/6-31+(d,p)) within polarizable continuum model. The results showed the importance of the bond dissociation enthalpies (BDEs) related to the first and second (BDEd) hydrogen atom transfer (intrinsic parameters) for rationalizing the antioxidant activity. In addition to the number of OH groups, the presence of a bromine substituent plays an interesting role in modulating the antioxidant activity. Theoretical thermodynamic and kinetic studies demonstrated that the free radical scavenging by these Schiff bases mainly proceeds through proton-coupled electron transfer rather than sequential proton loss electron transfer, the latter mechanism being only feasible at relatively high pH.

Systematic pharmacogenomics analysis of a Malay whole genome: proof of concept for personalized medicine.

BACKGROUND: With a higher throughput and lower cost in sequencing, second generation sequencing technology has immense potential for translation into clinical practice and in the realization of pharmacogenomics based patient care. The systematic analysis of whole genome sequences to assess patient to patient variability in pharmacokinetics and pharmacodynamics responses towards drugs would be the next step in future medicine in line with the vision of personalizing medicine. METHODS: Genomic DNA obtained from a 55 years old, self-declared healthy, anonymous male of Malay descent was sequenced. The subject's mother died of lung cancer and the father had a history of schizophrenia and deceased at the age of 65 years old. A systematic, intuitive computational workflow/pipeline integrating custom algorithm in tandem with large datasets of variant annotations and gene functions for genetic variations with pharmacogenomics impact was developed. A comprehensive pathway map of drug transport, metabolism and action was used as a template to map non-synonymous variations with potential functional consequences. PRINCIPAL FINDINGS: Over 3 million known variations and 100,898 novel variations in the Malay genome were identified. Further in-depth pharmacogenetics analysis revealed a total of 607 unique variants in 563 proteins, with the eventual identification of 4 drug transport genes, 2 drug metabolizing enzyme genes and 33 target genes harboring deleterious SNVs involved in pharmacological pathways, which could have a potential role in clinical settings. CONCLUSIONS: The current study successfully unravels the potential of personal genome sequencing in understanding the functionally relevant variations with potential influence on drug transport, metabolism and differential therapeutic outcomes. These will be essential for realizing personalized medicine through the use of comprehensive computational pipeline for systematic data mining and analysis.

Chemical and pharmacognostical characterization of two Malaysian plants both known as Ajisamat

Ajisamat, an herb commonly used as an aphrodisiac in the Malaysian traditional medicine, corresponds to two different species from different families - Salacia macrophylla Blume, Celastraceae, and Prismatomeris glabra (Korth.) Valeton, Rubiaceae. Macromorphological inspection of the vegetative parts both plants reveals only a slight difference in the arrangement of the petioles. Microscopic investigation of the plants roots used as crude drugs revealed however distinctive anatomical features. Prismatic calcium oxalate crystals and banded paratracheal parenchyma are characteristics of S. macrophylla while P. glabra displays an abundance as crystals. Other features such as vessels diameters and arrangements are also of diagnostic importance. Some of these characters were also identified in the powder of thes e plant materials and proposed for diagnostic purpose. The values for extraction of ethanol and water as well as total ash, acid-insoluble ash, water-soluble ash and sulfated ash were determined for both plants. Phytochemical studies were carried out on hexane and chloroform extracts of S. macrophylla and methanolic extract of P. glabra. S. macrophylla was shown to contain highly oxidized pentacyclic triterpenes while P. glabra contains anthraquinones. The pharmacognostical and hytochemical information can be utilised as the identification tools for Salacia macrophylla and Prismatomeris glabra .

Stevioside from Stevia rebaudiana Bertoni Increases Insulin Sensitivity in 3T3-L1 Adipocytes.

Stevioside from Stevia rebaudiana has been reported to exert antihyperglycemic effects in both rat and human subjects. There have been few studies on these effects in vitro. In this paper, radioactive glucose uptake assay was implemented in order to assess improvements in insulin sensitivity in 3T3-L1 cells by elevation of glucose uptake following treatment with stevioside. Oil Red-O staining and MTT assay were utilized to confirm adipocyte differentiation and cell viability, respectively. Findings from this research showed a significant increase in absorbance values in mature adipocytes following Oil Red-O staining, confirming the differentiation process. Stevioside was noncytotoxic to 3T3-L1 cells as cell viability was reduced by a maximum of 17%, making it impossible to determine its IC50. Stevioside increased glucose uptake activities by 2.1 times (p < 0.001) in normal conditions and up to 4.4 times (p < 0.001) in insulin-resistant states. At times, this increase was higher than that seen in positive control group treated with rosiglitazone maleate, an antidiabetic agent. Expressions of pY20 and p-IRS1 which were measured via Western blot were improved by stevioside treatment. In conclusion, stevioside has direct effects on 3T3-L1 insulin sensitivity via increase in glucose uptake and enhanced expression of proteins involved in insulin-signalling pathway.

Hypoglycemic effect of Octomeles sumatrana aqueous extract in streptozotocin-induced diabetic rats and its molecular mechanisms.

OBJECTIVE: To investigate the hypoglycemic effect of the aqueous extract of Octomeles sumatrana (O. sumatrana) (OS) in streptozotocin-induced diabetic rats (STZ) and its molecular mechanisms. METHODS: Diabetes was induced by intraperitoneal (i.p.) injection of streptozotocin (55 mg/kg) in to male Sprague-Dawley rats. Rats were divided into six different groups; normal control rats were not induced with STZ and served as reference, STZ diabetic control rats were given normal saline. Three groups were treated with OS aqueous extract at 0.2, 0.3 and 0.5 g/kg, orally twice daily continuously for 21 d. The fifth group was treated with glibenclamide (6 mg/kg) in aqueous solution orally continuously for 21 d. After completion of the treatment period, biochemical parameters and expression levels of glucose transporter 2 (Slc2a2), glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PCK1) were determined in liver by quantitative real time PCR. RESULTS: Administration of OS at different doses to STZ induced diabetic rats, resulted in significant decrease (P<0.05) in blood glucose level in a dose dependent manner by 36%, 48%, and 64% at doses of 0.2, 0.3 and 0.5 g/kg, respectively, in comparison to the STZ control values. Treatment with OS elicited an increase in the expression level of Slc2a2 gene but reduced the expression of G6Pase and PCK1 genes. Morefore, OS treated rats, showed significantly lower levels of serum alanine transaminase (ALT), aspartate aminotransferase (AST) and urea levels compared to STZ untreated rats. The extract at different doses elicited signs of recovery in body weight gain when compared to STZ diabetic controls although food and water consumption were significantly lower in treated groups compared to STZ diabetic control group. CONCLUSIONS: O. sumatrana aqueous extract is beneficial for improvement of hyperglycemia by increasing gene expression of liver Slc2a2 and reducing expression of G6Pase and PCK1 genes in streptozotocin-induced diabetic rats.

Enhanced CD4+CD25+ regulatory T cells with splenic proliferation and protection against oxidative stress by nicotinamide in gestational diabetes.

Gestational diabetes mellitus (GDM) is a common complication during pregnancy. Metabolic changes in GDM affect fetal development and fetal glucose homeostasis. Several complications of diabetes are related to increased intracellular oxidative stress where prooxidants exceed antioxidant capacity. The present study was initiated to evaluate the effects of nicotinamide on CD4+CD25+ regulatory T cells (Tregs), proliferation of splenocytes, production of reactive oxygen species (ROS) by neutrophils and serum glucose levels. Changes in mRNA levels of two antioxidant genes in liver, viz, superoxide dismutase (SOD1) and catalase (CAT) were quantified with real-time PCR (QRT-PCR). Nicotinamide (50, 100 and 200 mg/kg) was supplemented p.o. to pregnant diabetic rats from days 6 through 20 of gestation. The highest dose enhanced expression of Tregs and increased splenocytes proliferation in both resting and lipopolysaccharide (LPS)-stimulated cells. Oxidative burst activity of neutrophils in response to phorbol myristate acetate (PMA), N-formyl-methionyl-leucyl-phenylalanine (FMLP) or E. coli activation was reduced. mRNA expressions of superoxide dismutase (SOD) and catalase (CAT) genes were upregulated by nicotinamide. In summary, nicotinamide boosted the immune system through stimulation of adaptive immune cells with enhancement of antioxidant defences and reduced production of ROS. Serum glucose level was normalised by nicotinamide (200 mg/kg). These findings provide evidence for usage of nicotinamide as a supplement or as adjunct to therapeutic agents in gestational diabetes and in pregnant individuals with weakened immune systems.

Immunomodulatory activity of polyphenols derived from Cassia auriculata flowers in aged rats.

The immunomodulatory activity of Cassia auriculata (CA)-derived polyphenols was tested on aged rats. Rats (24-26 months old) were given CA polyphenols supplementation at doses of 25, 50, and 100 mg/kg for 28 days. Flow cytometry analysis of CA polyphenols-treated aged rats showed increased T and B cells percentage along with enhanced proliferation of splenocytes in both resting and LPS-stimulated cells. Increased percentage of pan T cells is further supported by an elevation of CD4+, CD8+, and CD4+CD25+ regulatory cells. In terms of innate immune cell activity, CA polyphenol supplementation reduced the oxidative burst activity of neutrophils in response to PMA and Escherichia coli activation. Our results collectively show that polyphenols derived from CA boost T cell immunity by increasing the number of T cells and its sensitivity towards stimulants and decreasing ROS production by neutrophils that could potentially harm multiple biological systems in aged individuals.

Amino and fatty acid compositions in Haruan traditional extract (HTE) / Composición de aminoácidos y ácidos grasos en extractos de Haruan tradicional

Evaluation of amino and fatty acids compositions in Haruan Traditional Extracts (HTE) was done using HPLC and GC methods. The HTE contained at least 17 amino acids with glutamic acid, glycine, leusine, aspartic acid, proline, alanine and arginine are the most, with values 1.87 - 43.13 mg/g, 21.80 - 80.85 mg/g, 7.85- 40.19 mg/g, 13.85 - 44.07 mg/g, 9.49 - 45.46 mg/g, 11.38 - 35.25 mg/g and 5.99 - 21.79 mg/g, respectively. Meanwhile, the highest percentage of fatty acids is palmitic acid; 3.54 - 26.84 percent of total protein. The others major fatty acids are stearic acid, oleic acid and linoleic acid with values 3.25 - 15.90 percent, 1.40 - 27.68 percent, 0.51 - 7.82 percent of total protein, respectively. HTE also found to have 4 extra bioactive compounds labelled as 1 to 4 on chromatographic tracing which in line with previously finding. It is concluded that the HTE is containing all the important amino acids plus some fatty acids, which is the basis to conduct antioxidant composition in both fresh Haruan and the HTE which was claimed to have wound healing properties. Comparative study was also carried out in various other extraction protocols, including commercial product.

Evaluación de las composiciones de aminoácidos y ácidos grasos en Haruan Extractos tradicional (HTE) se realizó mediante métodos de HPLC y GC. La HTE contenía al menos 17 aminoácidos con ácido glutámico, glicina, leucina, ácido aspártico, prolina, alanina y arginina como mayoritarios, con valores de 1.87 - 43.13 mg/g, 21.80 - 80.85 mg/g, 7.85 - 40.19 mg/g, 13.85 - 44.07 mg/g, 9.49 - 45.46 mg/g, 11.38 - 35.25 mg/g and 5.99 - 21.79 mg/g, respectivamente. Mientras tanto, el mayor porcentaje de ácidos grasos es el ácido palmítico; 3.54 - 26.84 por ciento de la proteína total. Otros ácidos grasos importantes son el ácido esteárico, ácido oleico y ácido linoleico con valores de 3.25 - 15.90 por ciento, 1.40 - 27.68 %, 0.51 - 7.82 por ciento de la proteína total, respectivamente. HTE también encontró cuatro compuestos bioactivos adicionales etiquetados de 1 a 4 en el seguimiento cromatográfico que está de acuerdo con resultados previos. Se concluye que la HTE contiene todos los aminoácidos importantes además de algunos ácidos grasos, que es la base para llevar a cabo la composición antioxidante, tanto en fresco Haruan y la HTE que se afirma poseen propiedades curativas. Estudios comparativos se llevaron a cabo con otros protocolos de extracción, incluido el producto comercial.