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Fulltext Algorithm for rapid identification of flavonoids classes

Noh, C.H.C., Azmin, N.F.M., Amid, A., Asnawi, A.L.

International Food Research Journal, 2017;24(101):0-0.
MyJurnal

Bioactive compounds are one of the natural products used especially for medicinal, pharmaceutical and food application. Increasing research performed on the extraction, isolation and identification of bioactive compounds, however non to date has explored on the identification of flavonoids classes. Therefore, this study was focused on the development of algorithm for rapid identification of flavonoids classes which are flavanone, flavone and flavonol and also their derivatives. Fourier Transform Infrared (FTIR) spectroscopy coupled with multivariate statistical data analysis, which is Principal Component Analysis (PCA) was utilized. The results exhibited that few significant wavenumber range provides the identification and characterization of the flavonoids classes based on PCA algorithm. The study concluded that FTIR coupled with PCA analysis can be used as a molecular fingerprint for rapid identification of flavonoids.
Fulltext Protease purification from Bacillus Amyloliquefaciens B7 using Aqueous Two-Phase System (ATPS)

Abd Samad, N.S., Amid, A., Jimat, D.N., Shukor, N.A.

International Food Research Journal, 2017;24(101):0-0.
MyJurnal

Bacillus amyloliquefaciens B7 was isolated from the fermented fish sauce and identified as protease producer. Generally, in downstream processing, purification of enzymes consumes higher cost regarding reagents and equipment used. Moreover, harsh purification methods used might cause denaturation of the enzymes. Therefore, there is a high demand for efficient and lowcost extraction and purification methods. Aqueous two-phase system (ATPS) is an alternative method that should be considered as it is simple, rapid separation yet cause little denaturation. Protease produced by B. amyloliquefaciens B7 was partitioned using two different ATPS, which were PEG/potassium phosphate and PEG/sodium citrate. Results showed the highest enzyme activity was found in interface phase with the ATPS system of 27% (w/w) PEG1500/ 34% (w/w) sodium citrate. Later, the ATPS conditions (pH, temperature, the concentration of selected salt and PEG) were optimized by using response surface methodology. The optimum conditions for ATPS purification were observed in ATPS conditions at pH 7 and 35°C with the enzyme activity of 0.20± 0.01 U/ml.
Fulltext Effects of process conditions on the ultrasonic extraction of phenolics scavenger from Curcuma Caesia Rhizome

Jemain, S.F.P., Jamal, P., Raus, A. R., Amid, A., Jaswir, I.

International Food Research Journal, 2017;24(101):0-0.
MyJurnal

Medicinal properties of Malaysian Curcuma caesia have not been studied extensively, even though it has been used as a traditional remedy. This study examined the effects of various extraction temperatures (30, 40, 50, 60, 70oC) using a high frequency (40 kHz) ultrasonic extraction method, time (30,60,90 and 120 minutes), pH (1,2,3,4,5,6,7,8,9,10) on the extraction yield of total phenolics and DPPH (2,2-diphenyl-1-picrylhydrazyl) scavenging activities from C. caesia rhizome. Extraction was most efficient at pH 6.0, while the extraction time of 30 minutes and temperature of 60oC was the best in terms of total phenolics content and DPPH scavenging activity. This study is important due to its ability to improve extraction of total phenolics compound using ultrasonic extraction method while maintaining a relatively high DPPH scavenging activity of the extracts.
Fulltext Enzyme aided extraction of sulfated polysaccharides from Turbinaria turbinata brown seaweed

Hammed, A. M., Jaswir, I., Simsek, S., Alam, Z., Amid, A.

International Food Research Journal, 2017;24(4):1660-1666.
MyJurnal

This study involves extraction of sulfated polysaccahride (SP) from brown seaweed (Turbinaria turbinata). Eight processing conditions affecting enzyme aided extraction (EAE) were screened using Plackett-Burman design. Three significant factors (hydrolysis time, enzyme concentration and extraction stage) were optimized using Faced Centred Central Composite Design in Random Surface Methods. Micrograph obtained using Field Emission Scanning Electron Microscopy revealed that cellulase degradation ruptured the seaweed cell matrix thus caused increase in the release of SP. The optimum conditions for extraction of SP from T. turbinata are: extraction stage of 2, hydrolysis time of 19.5 h and enzyme concentration of 1.5 μl/ml to produce 25.13% yield. The SP obtained from cellulase treated T. turbinata is a suitable anti-inflammatory agent for pharmaceutical applications.
Fulltext Xanthine oxidase inhibitory activity from potential Malaysian medicinal plant as remedies for gout

Azmi SMN, Jamal P, Amid A

International Food Research Journal, 2012;19(1):159-165.
MyJurnal

Malaysia has a rich diversity of medicinal plants and some of them inhibit xanthine oxidase (XO), which can be introduced as new natural sources of gout medication and a substitute for synthetic xanthine oxidase inhibitors (XOI). The degree of XO inhibitory activity was determined by measuring the absorbance spectrophotometrically at 295 nm, which is associated with uric acid formation. Our preliminary screening study had employed the use of distilled water, 70% methanol and absolute ethanol to extract XOI from twenty parts of five plant species, namely, Averrhoa carambola, Carica papaya, Dimocarpus longan malesianus, Manilkara zapota and Salacca zalacca. These plants were selected based on their frequent medicinal usages by local folks. The results have shown that an aqueous extract of Carica papaya mature leaves has promising activity to inhibit XO up to 75.68 ± 0.1%. Statistical experimental design were employed to optimize the selected sample (dried Carica papaya leaves: distilled water) on extraction of XOI and the maximum XOI percentage of 86.93 ± 1.9% was obtained, which exhibited only 6.76% less than the activity exhibited by allopurinol (93.69 ± 0.2%), a commercial XOI. The comparison was made between allopurinol and optimized extract on the basis of IC50concentrations. Allopurinol showed IC50 value of 3.74 μg/ml that is considerably lower as compared to the optimized sample (4.33 μg/ml).
Gene expression analysis in MCF-7 breast cancer cells treated with recombinant bromelain

Fouz N, Amid A, Hashim YZ

Appl Biochem Biotechnol, 2014 Aug;173(7):1618-39.
PMID: 24928548 DOI: 10.1007/s12010-014-0947-6

The contributing molecular pathways underlying the pathogenesis of breast cancer need to be better characterized. The principle of our study was to better understand the genetic mechanism of oncogenesis for human breast cancer and to discover new possible tumor markers for use in clinical practice. We used complimentary DNA (cDNA) microarrays to compare gene expression profiles of treated Michigan Cancer Foundation-7 (MCF-7) with recombinant bromelain and untreated MCF-7. SpringGene analysis was carried out of differential expression followed by Ingenuity Pathway Analysis (IPA), to understand the underlying consequence in developing disease and disorders. We identified 1,102 known genes differentially expressed to a significant degree (p<0.001) changed between the treatment. Within this gene set, 20 genes were significantly changed between treated cells and the control cells with cutoff fold change of more than 1.5. These genes are RNA-binding motif, single-stranded interacting protein 1 (RBMS1), ribosomal protein L29 (RPL29), glutathione S-transferase mu 2 (GSTM2), C15orf32, Akt3, B cell translocation gene 1 (BTG1), C6orf62, C7orf60, kinesin-associated protein 3 (KIFAP3), FBXO11, AT-rich interactive domain 4A (ARID4A), COPS2, TBPL1|SLC2A12, TMEM59, SNORD46, glioma tumor suppressor candidate region gene 2 (GLTSCR2), and LRRFIP. Our observation on gene expression indicated that recombinant bromelain produces a unique signature affecting different pathways, specific for each congener. The microarray results give a molecular mechanistic insight and functional effects, following recombinant bromelain treatment. The extent of changes in genes is related to and involved significantly in gap junction signaling, amyloid processing, cell cycle regulation by BTG family proteins, and breast cancer regulation by stathmin1 that play major roles.
Cytokinetic study of MCF-7 cells treated with commercial and recombinant bromelain

Fouz N, Amid A, Hashim YZ

Asian Pac J Cancer Prev, 2014 Jan;14(11):6709-14.
PMID: 24377593

BACKGROUND: Breast cancer is a leading cause of death in women. The available chemotherapy drugs have been associated with many side effects. Bromelain has novel medicinal qualities including anti-inflammatory, anti-thrombotic, fibrinolytic and anti-cancer functions. Commercially available bromelain is obtained through tedious methods; therefore, recombinant bromelain may provide a cheaper and simpler choice with similar quality.
MATERIALS AND METHODS: This study aimed to assess the effects of commercial and recombinant bromelain on the cytokinetic behavior of MCF-7 breast cancer cells and their potential as therapeutic alternatives in cancer treatment. Cytotoxic activities of commercial and recombinant bromelain were determined using (sulforhodamine) SRB assay. Next, cell viability assays were conducted to determine effects of commercial and recombinant bromelain on MCF-7 cell cytokinetic behavior. Finally, the established growth kinetic data were used to modify a model that predicts the effects of commercial and recombinant bromelain on MCF-7 cells.
RESULTS: Commercial and recombinant bromelain exerted strong effects towards decreasing the cell viability of MCF-7 cells with IC50 values of 5.13 μg/mL and 6.25 μg/mL, respectively, compared to taxol with an IC50 value of 0.063 μg/mL. The present results indicate that commercial and recombinant bromelain both have anti-proliferative activity, reduced the number of cell generations from 3.92 to 2.81 for commercial bromelain and to 2.86 for recombinant bromelain, while with taxol reduction was to 3.12. Microscopic observation of bromelain-treated MCF-7 cells demonstrated detachment. Inhibition activity was verified with growth rates decreased dynamically from 0.009 h-1 to 0.0059 h-1 for commercial bromelain and to 0.0063 h-1 for recombinant bromelain.
CONCLUSIONS: Commercial and recombinant bromelain both affect cytokinetics of MCF-7 cells by decreasing cell viability, demonstrating similar strength to taxol.
Fulltext Identification of troponin I and actin, alpha cardiac muscle 1 as potential biomarkers for hearts of electrically stimulated chickens

Amid A, Samah NA, Yusof F

Proteome Sci, 2012;10(1):1.
PMID: 22230661 DOI: 10.1186/1477-5956-10-1

In this study, proteomics methods have been used to study the effects of different currents and voltages used to stun chickens. Protein profiles of chicken hearts were constructed to detect differences in protein expression and modification. The different voltages studied were 10 V, 40 V and 70 V, while the currents examined were 0.25 A, 0.5 A, and 0.75 A. The profiles obtained from these stunning conditions were compared to the non-stunned (0 A, 0 V) sample.
Microarray and quantitative PCR analysis of gene expression profiles in response to treatment with tomato leaf extract in mcf-7 breast cancer cells

Amid A, Wan Chik WD, Jamal P, Hashim YZ

Asian Pac J Cancer Prev, 2012;13(12):6319-25.
PMID: 23464452

We previously found cytotoxic effects of tomato leaf extract (TLE) on the MCF-7 breast cancer cell line. The aim of this study was to ascertain the molecular mechanisms associated with the usage of TLE as an anticancer agent by microarray analysis using mRNA from MCF-7 breast cancer cells after treatment with TLE for 1 hr and 48 hrs. Approximately 991 genes out of the 30,000 genes in the human genome were significantly (p<0.05) changed after the treatment. Within this gene set, 88 were significantly changed between the TLE treated cells and the untreated MCF-7 cells (control cells) with a cut-off fold change >2.00. In order to focus on genes that were involved in cancer cell growth, only twenty-nine genes were selected, either down-regulated or up-regulated after treatment with TLE. Microarray assay results were confirmed by analyzing 10 of the most up and down regulated genes related to cancer cells progression using real-time PCR. Treatment with TLE induced significant up-regulation in the expression of the CRYAB, PIM1, BTG1, CYR61, HIF1-α and CEBP-β genes after 1 hr and 48 hrs, whereas the TXNIP and THBS1 genes were up-regulated after 1 hr of treatment but down-regulated after 48 hrs. In addition both the HMG1L1 and HIST2H3D genes were down-regulated after 1 hr and 48 hrs of treatment. These results demonstrate the potent activity of TLE as an anticancer agent.
Fulltext Cocoa pod husk, a new source of hydrolase enzymes for preparation of cross-linked enzyme aggregate

Yusof F, Khanahmadi S, Amid A, Mahmod SS

Springerplus, 2016;5:57.
PMID: 26904389 DOI: 10.1186/s40064-015-1621-3

Cocoa pod husk (CPH) is a by-product of cocoa production obtained after removing the beans from the fruit. The analysis of CPH has shown that it contains high amounts of protein. This study is aimed to utilize this protein source in hydrolase enzyme production. In this study, seven hydrolase enzymes (amylase, fructosyltransferase, mannanase, glucosidase, glucanase, lipase and protease) were screened from CPH for the first time for feasible industrial production. Among these hydrolases, lipase was chosen for the next steps of experiments as it has a lot of applications in different industries. The extraction of high active lipase from CPH has been done under optimum conditions. The condition that was optimum for the three major factors was achieved using Face centered central composite design (FCCCD) with response surface methodology (RSM) to obtain the highest enzyme activity of crude lipase from CPH. The optimum condition of extraction is used for preparation of cross-linked enzyme aggregate (CLEA). For the production of immobilized biocatalyst, the technique of CLEA is considered as an effective technique for its industrially attractive advantages. Referring to the results of OFAT, CLEA-lipase was prepared in the best condition at the presence of 30 mM ammonium sulphate, 70 mM glutaraldehyde with 0.23 mM Bovine serum albumin as an additive. Immobilization effectively improved the stability of lipase against various organic solvents.
Understanding thermostability factors of Aspergillus niger PhyA phytase: a molecular dynamics study

Noorbatcha IA, Sultan AM, Salleh HM, Amid A

Protein J, 2013 Apr;32(4):309-16.
PMID: 23636517 DOI: 10.1007/s10930-013-9489-y

Molecular dynamics simulation was used to study the dynamic differences between native Aspergillus niger PhyA phytase and a mutant with 20 % greater thermostability. Atomic root mean square deviation, radius of gyration, and number of hydrogen bonds and salt bridges are examined to determine thermostability factors. The results suggest that, among secondary structure elements, loops have the most impact on the thermal stability of A. niger phytase. In addition, the location rather than the number of hydrogen bonds is found to have an important contribution to thermostability. The results also show that salt bridges may have stabilizing or destabilizing effect on the enzyme and influence its thermostability accordingly.
Fulltext Recombinant bromelain production in Escherichia coli: process optimization in shake flask culture by response surface methodology

Muntari B, Amid A, Mel M, Jami MS, Salleh HM

AMB Express, 2012;2:12.
PMID: 22336426 DOI: 10.1186/2191-0855-2-12

Bromelain, a cysteine protease with various therapeutic and industrial applications, was expressed in Escherichia coli, BL21-AI clone, under different cultivation conditions (post-induction temperature, L-arabinose concentration and post-induction period). The optimized conditions by response surface methodology using face centered central composite design were 0.2% (w/v) L-arabinose, 8 hr and 25°C. The analysis of variance coupled with larger value of R2 (0.989) showed that the quadratic model used for the prediction was highly significant (p < 0.05). Under the optimized conditions, the model produced bromelain activity of 9.2 U/mg while validation experiments gave bromelain activity of 9.6 ± 0.02 U/mg at 0.15% (w/v) L-arabinose, 8 hr and 27°C. This study had innovatively developed cultivation conditions for better production of recombinant bromelain in shake flask culture.
Optimized preparation and characterization of CLEA-lipase from cocoa pod husk

Khanahmadi S, Yusof F, Amid A, Mahmod SS, Mahat MK

J Biotechnol, 2015 May 20;202:153-61.
PMID: 25481099 DOI: 10.1016/j.jbiotec.2014.11.015

Cross-linked enzyme aggregate (CLEA) is easily prepared from crude enzyme and has many advantages to the environment and it is considered as an economic method in the context of industrial biocatalysis compared to free enzyme. In this work, a highly active and stable CLEA-lipase from cocoa pod husk (CPH) which is a by-product after removal of cocoa beans, were assayed for their hydrolytic activity and characterized under the optimum condition successfully. Face centered central composite design (FCCCD) under response surface methodology (RSM) was used to get the optimal conditions of the three significant factors (concentration of ammonium sulfate, concentration of glutaraldehyde and concentration of additive) to achieve higher enzyme activity of CLEA. From 20 runs, the highest activity recorded was around 9.407U (83% recovered activity) under the condition of using 20% saturated ammonium sulfate, 60mM glutaraldehyde as cross-linker and 0.17mM bovine serum albumin as feeder. Moreover, the optimal reaction temperature and pH value in enzymatic reaction for both crude enzyme and immobilized were found to be 45°C at pH 8 and 60°C at pH 8.2, respectively. A systematic study of the stability of CLEA and crude enzyme was taken with regards to temperature (25-60°C) and pH (5-10) value and in both factors, CLEA-lipase showed more stability than free lipase. The Km value of CLEA was higher compared to free enzyme (0.55mM vs. 0.08mM). The CLEA retained more than 60% of the initial activity after six cycles of reuse compared to free enzyme. The high stability and recyclability of CLEA-lipase from CPH make it efficient for different industrial applications.
Cocoa pod husk: A new source of CLEA-lipase for preparation of low-cost biodiesel: An optimized process

Khanahmadi S, Yusof F, Chyuan Ong H, Amid A, Shah H

J Biotechnol, 2016 Aug 10;231:95-105.
PMID: 27184429 DOI: 10.1016/j.jbiotec.2016.05.015

Enzymatic reactions involving lipases as catalyst in transesterification can be an excellent alternative to produce environmental-friendly biodiesel. In this study, lipase extracted from Cocoa Pod Husk (CPH) and immobilized through cross linked enzyme aggregate (CLEA) technology catalysed the transesterification of Jatropha curcas oil successfully. Face centered central composite design (FCCCD) under response surface methodology (RSM) was used to get the optimal conditions of 3% (w/w) enzyme loading, 4h reaction time and 1:6 oil/ethanol ratio to achieve the highest conversion of free fatty acid and glycerides into biodiesel (93%). The reusability of CLEA-lipase was tested and after seven cycles, the conversion percentage reduced to 58%. The results revealed that CLEA lipase from CPH is a potential catalyst for biodiesel production.
Evaluation of metabolomics behavior of human colon cancer HT29 cell lines treated with ionic liquid graviola fruit pulp extract

Daddiouaissa D, Amid A, Abdullah Sani MS, Elnour AAM

J Ethnopharmacol, 2021 Apr 24;270:113813.
PMID: 33444719 DOI: 10.1016/j.jep.2021.113813

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants have been used by indigenous people across the world for centuries to help individuals preserve their wellbeing and cure diseases. Annona muricata L. (Graviola) which is belonging to the Annonaceae family has been traditionally used due to its medicinal abilities including antimicrobial, anti-inflammatory, antioxidant and cancer cell growth inhibition. Graviola is claimed to be a potential antitumor due to its selective cytotoxicity against several cancer cell lines. However, the metabolic mechanism information underlying the anticancer activity remains limited.
AIM OF THE STUDY: This study aimed to investigate the effect of ionic liquid-Graviola fruit pulp extract (IL-GPE) on the metabolomics behavior of colon cancer (HT29) by using an untargeted GC-TOFMS-based metabolic profiling.
MATERIALS AND METHODS: Multivariate data analysis was used to determine the metabolic profiling, and the ingenuity pathway analysis (IPA) was used to predict the altered canonical pathways after treating the HT29 cells with crude IL-GPE and Taxol (positive control).
RESULTS: The principal components analysis (PCA) identified 44 metabolites with the most reliable factor loading, and the cluster analysis (CA) separated three groups of metabolites: metabolites specific to the non-treated HT29 cells, metabolites specific to the treated HT29 cells with the crude IL-GPE and metabolites specific to Taxol treatment. Pathway analysis of metabolomic profiles revealed an alteration of many metabolic pathways, including amino acid metabolism, aerobic glycolysis, urea cycle and ketone bodies metabolism that contribute to energy metabolism and cancer cell proliferation.
CONCLUSION: The crude IL-GPE can be one of the promising anticancer agents due to its selective inhibition of energy metabolism and cancer cell proliferation.
Anti-Proliferative Activity of Triterpenes Isolated from Clinicanthus nutans on Hep-G2 Liver Cancer Cells

Zakaria KN, Amid A, Zakaria Z, Jamal P, Ismail A

Asian Pac J Cancer Prev, 2019 Feb 26;20(2):563-567.
PMID: 30803221

Problem statement: Clinicanthus nutans has been used by Malaysian since long time ago. It is used to treat many diseases including cancer. Many studies carried out on its crude extract but no clear report on the specific secondary metabolites responsible for its nature in treating selected diseases. Objective: This study aims to confirm the practice carried out by many people on the usage of Clinicanthus nutans in treating cancer. Methods: C. nutans leaves were extracted by methanol. Thin layer chromatography was used to identify the suitable solvent for fractions separation. The fractions were then separated at larger volume using gravity column chromatography. Each fraction was tested on its anti-proliferative activity on Hep-G2 liver cancer cells by MTT assay. The phytochemical screening was carried out to identify the bioactive compound based on qualitative analysis. Results: The fraction 2 (F2) of C. nutans showed the lowest IC50 value of 1.73 μg/ml against Hep-G2 cancer cells, and it is identified as triterpenes. Conclusion: The fraction F2 identified as triterpenes isolated from C. nutans has potential as an anti-proliferative agent against liver cancer.
Fulltext Statistical Analysis of Metal Chelating Activity of Centella asiatica and Erythroxylum cuneatum Using Response Surface Methodology

Mohd Salim RJ, Adenan MI, Amid A, Jauri MH, Sued AS

Biotechnol Res Int, 2013;2013:137851.
PMID: 23533781 DOI: 10.1155/2013/137851

The purpose of the study is to evaluate the relationship between the extraction parameters and the metal chelating activity of Centella asiatica (CA) and Erythroxylum cuneatum (EC). The response surface methodology was used to optimize the extraction parameters of methanolic extract of CA and EC with respect to the metal chelating activity. For CA, Run 17 gave optimum chelating activity with IC50 = 0.93 mg/mL at an extraction temperature of 25°C, speed of agitation at 200 rpm, ratio of plant material to solvent at 1 g : 45 mL and extraction time at 1.5 hour. As for EC, Run 13 with 60°C, 200 rpm, 1 g : 35 mL and 1 hour had metal chelating activity at IC50 = 0.3817 mg/mL. Both optimized extracts were further partitioned using a solvent system to evaluate the fraction responsible for the chelating activity of the plants. The hexane fraction of CA showed potential activity with chelating activity at IC50 = 0.090 and the ethyl acetate fraction of EC had IC50 = 0.120 mg/mL. The study showed that the response surface methodology helped to reduce the extraction time, temperature and agitation and subsequently improve the chelating activity of the plants in comparison to the conventional method.
Fulltext Metabolomics profiling of extracellular metabolites in CHO-K1 cells cultured in different types of growth media

Mohmad-Saberi SE, Hashim YZ, Mel M, Amid A, Ahmad-Raus R, Packeer-Mohamed V

Cytotechnology, 2013 Aug;65(4):577-86.
PMID: 23179090 DOI: 10.1007/s10616-012-9508-4

An efficient mammalian cell system for producing bioproducts should retain high cell viability and efficient use of energy sources rendering the need to understand the effects of various variables on the cell system. In this study, global metabolite (metabolomics) analysis approach was used to try and understand the relationships between types of media used, culture growth behavior and productivity. CHO-KI cells producing IGF-1 were obtained from ATCC and grown in T-flask (37 °C, 5 % CO2) until 70-80 % confluent in RPMI 1640 and Ham's F12, respectively. Samples were taken at 8-hourly intervals for routine cell counting, biochemical responses, insulin like growth factor-1 (IGF-1) protein concentration and global metabolite analysis (gas chromatography mass spectrometry, GCMS). Conditioned media from each time point were spun down before injection into GCMS. Data from GCMS were then transferred to SIMCA-P + Version 12 for chemometric evaluation using principal component analysis. The results showed that while routine analysis gave only subtle differences between the media, global metabolite analysis was able to clearly separate the culture based on growth media with growth phases as confounding factor. Different types of media also appeared to affect IGF-1 production. Asparagine was found to be indicative of healthiness of cells and production of high IGF-1. Meanwhile identification of ornithine and lysine in death phase was found to be associated with apoptosis and oversupplied nutrient respectively. Using the biomarkers revealed in the study, several bioprocessing strategies including medium improvement and in-time downstream processing can be potentially implemented to achieve efficient CHO culture system.
Bromelain: an overview of industrial application and purification strategies

Arshad ZI, Amid A, Yusof F, Jaswir I, Ahmad K, Loke SP

Appl Microbiol Biotechnol, 2014 Sep;98(17):7283-97.
PMID: 24965557 DOI: 10.1007/s00253-014-5889-y

This review highlights the use of bromelain in various applications with up-to-date literature on the purification of bromelain from pineapple fruit and waste such as peel, core, crown, and leaves. Bromelain, a cysteine protease, has been exploited commercially in many applications in the food, beverage, tenderization, cosmetic, pharmaceutical, and textile industries. Researchers worldwide have been directing their interest to purification strategies by applying conventional and modern approaches, such as manipulating the pH, affinity, hydrophobicity, and temperature conditions in accord with the unique properties of bromelain. The amount of downstream processing will depend on its intended application in industries. The breakthrough of recombinant DNA technology has facilitated the large-scale production and purification of recombinant bromelain for novel applications in the future.
Antiproliferative activity of ionic liquid-graviola fruit extract against human breast cancer (MCF-7) cell lines using flow cytometry techniques

Daddiouaissa D, Amid A, Kabbashi NA, Fuad FAA, Elnour AM, Epandy MAKMS

J Ethnopharmacol, 2019 May 23;236:466-473.
PMID: 30853648 DOI: 10.1016/j.jep.2019.03.003

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants have been used for ages by indigenous communities around the world to help humankind sustain its health. Graviola (Annona muricata), also called soursop, is a member of the Annonaceae family and is an evergreen plant that is generally distributed in tropical and subtropical areas of the world. Graviola tree has a long history of traditional use due to its therapeutic potential including anti-inflammatory, antimicrobial, antioxidant, insecticide and cytotoxic to tumor cells.
AIM OF THE STUDY: This study aimed to investigate the in vitro antiproliferative effects and apoptotic events of the ionic liquid extract of Graviola fruit (IL-GFE) on MCF-7 breast cancer cells and their cytokinetics behaviour to observe their potential as a therapeutic alternative in cancer treatment.
MATERIALS AND METHODS: The cell viability assay of the extract was measured using tetrazolium bromide (MTT assay) to observe the effects of Graviola fruit extract. Then the cytokinetics behaviour of MCF-7 cells treated with IL-GFE is observed by plotting the growth curve of the cells. Additionally, the cell cycle distribution and apoptosis mechanism of IL-GFE action on MCF-7 cancer cells were observed by flow cytometry.
RESULTS: IL-GFE exhibited anti-proliferative activity on MCF-7 with the IC50 value of 4.75 μg/mL, compared to Taxol with an IC50 value of 0.99 μg/mL. IL- GFE also reduced the number of cell generations from 3.71 to 1.67 generations compared to 2.18 generations when treated with Taxol. Furthermore, the anti-proliferative activities were verified when the growth rate was decreased dynamically from 0.0077 h to 1 to 0.0035 h-1. Observation of the IL-GFE-treated MCF-7 under microscope demonstrated detachment of cells and loss of density. The growth inhibition of the cells by extracts was associated with cell cycle arrest at the G0/G1 phase, and phosphatidylserine externalisation confirms the anti-proliferation through apoptosis.
CONCLUSIONS: ionic liquid Graviola fruit extract affect the cytokinetics behaviour of MCF-7 cells by reducing cell viability, induce apoptosis and cell cycle arrest at the G0/G1 phase.

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