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Fulltext Optimization of ultraviolet ozone treatment process for improvement of polycaprolactone (PCL) microcarrier performance

Samsudin N, Hashim YZH, Arifin MA, Mel M, Salleh HM, Sopyan I, et al.

Cytotechnology, 2017 Aug;69(4):601-616.
PMID: 28337561 DOI: 10.1007/s10616-017-0071-x

Growing cells on microcarriers may have overcome the limitation of conventional cell culture system. However, the surface functionality of certain polymeric microcarriers for effective cell attachment and growth remains a challenge. Polycaprolactone (PCL), a biodegradable polymer has received considerable attention due to its good mechanical properties and degradation rate. The drawback is the non-polar hydrocarbon moiety which makes it not readily suitable for cell attachment. This report concerns the modification of PCL microcarrier surface (introduction of functional oxygen groups) using ultraviolet irradiation and ozone (UV/O3) system and investigation of the effects of ozone concentration, the amount of PCL and exposure time; where the optimum conditions were found to be at 60,110.52 ppm, 5.5 g PCL and 60 min, respectively. The optimum concentration of carboxyl group (COOH) absorbed on the surface was 1495.92 nmol/g and the amount of gelatin immobilized was 320 ± 0.9 µg/g on UV/O3 treated microcarriers as compared to the untreated (26.83 ± 3 µg/g) microcarriers. The absorption of functional oxygen groups on the surface and the immobilized gelatin was confirmed with the attenuated total reflectance Fourier transformed infrared spectroscopy (ATR-FTIR) and the enhancement of hydrophilicity of the surface was confirmed using water contact angle measurement which decreased (86.93°-49.34°) after UV/O3 treatment and subsequently after immobilization of gelatin. The attachment and growth kinetics for HaCaT skin keratinocyte cells showed that adhesion occurred much more rapidly for oxidized surfaces and gelatin immobilized surface as compared to untreated PCL.
Optimization of ultraviolet/ozone (UVO3) process conditions for the preparation of gelatin coated polystyrene (PS) microcarriers

Arifin MA, Mel M, Swan SY, Samsudin N, Hashim YZH, Salleh HM

Prep Biochem Biotechnol, 2022;52(2):181-196.
PMID: 34010098 DOI: 10.1080/10826068.2021.1923031

The aim of this study was to develop gelatin coated polystyrene (PS) microcarriers with good cell adhesion and proliferation properties. PS microspheres, prepared using oil-in water (o/w) solvent evaporation method, were loaded with oxygen containing functional groups using an ultraviolet/ozone (UVO3) system. Using water-soluble carbodiimide chemistry, gelatin was subsequently immobilized on UVO3 treated PS microspheres. The amount of immobilized gelatin was found to be directly proportional to the surface carboxyl (COOH) concentration on PS microspheres. Face Centered Central Composite Design (FCCD) was employed to optimize the process conditions of UVO3 treatment to maximize the surface COOH concentration on PS microspheres for allowing higher gelatin immobilization. Statistical results revealed that, the optimized process conditions were ozone flow rate of ∼64,603 ppm, exposure time of ∼60 minutes and sample amount of 5.05 g. Under these conditions, the surface COOH concentration on PS microspheres was ∼1,505 nmol/g with the corresponding amount of immobilized gelatin was ∼2,725 µg/g. Characterization analyses strongly suggest that the optimized UVO3 treatment and successive gelatin immobilization have successfully improved surface wettability and dispersion stability of PS microspheres. Moreover, gelatin coated PS microcarriers were also proven as able to support the growth of CHO-K1 cells in high cell density culture.
Fulltext Isotopic Tracer for Absolute Quantification of Metabolites of the Pentose Phosphate Pathway in Bacteria

Mohd Kamal K, Mahamad Maifiah MH, Zhu Y, Abdul Rahim N, Hashim YZH, Abdullah Sani MS

Metabolites, 2022 Nov 09;12(11).
PMID: 36355168 DOI: 10.3390/metabo12111085

The pentose phosphate pathway (PPP) plays a key role in many metabolic functions, including the generation of NADPH, biosynthesis of nucleotides, and carbon homeostasis. In particular, the intermediates of PPP have been found to be significantly perturbed in bacterial metabolomic studies. Nonetheless, detailed analysis to gain mechanistic information of PPP metabolism remains limited as most studies are unable to report on the absolute levels of the metabolites. Absolute quantification of metabolites is a prerequisite to study the details of fluxes and its regulations. Isotope tracer or labeling studies are conducted in vivo and in vitro and have significantly improved the analysis and understanding of PPP. Due to the laborious procedure and limitations in the in vivo method, an in vitro approach known as Group Specific Internal Standard Technology (GSIST) has been successfully developed to measure the absolute levels of central carbon metabolism, including PPP. The technique adopts derivatization of an experimental sample and a corresponding internal standard with isotope-coded reagents to provide better precision for accurate identification and absolute quantification. In this review, we highlight bacterial studies that employed isotopic tracers as the tagging agents used for the absolute quantification analysis of PPP metabolites.
Fulltext Bacterial Metabolomics: Sample Preparation Methods

Mohd Kamal K, Mahamad Maifiah MH, Abdul Rahim N, Hashim YZH, Abdullah Sani MS, Azizan KA

Biochem Res Int, 2022;2022:9186536.
PMID: 35465444 DOI: 10.1155/2022/9186536

Metabolomics is a comprehensive analysis of metabolites existing in biological systems. As one of the important "omics" tools, the approach has been widely employed in various fields in helping to better understand the complex cellular metabolic states and changes. Bacterial metabolomics has gained a significant interest as bacteria serve to provide a better subject or model at systems level. The approach in metabolomics is categorized into untargeted and targeted which serves different paradigms of interest. Nevertheless, the bottleneck in metabolomics has been the sample or metabolite preparation method. A custom-made method and design for a particular species or strain of bacteria might be necessary as most studies generally refer to other bacteria or even yeast and fungi that may lead to unreliable analysis. The paramount aspect of metabolomics design comprises sample harvesting, quenching, and metabolite extraction procedures. Depending on the type of samples and research objective, each step must be at optimal conditions which are significantly important in determining the final output. To date, there are no standardized nor single designated protocols that have been established for a specific bacteria strain for untargeted and targeted approaches. In this paper, the existing and current developments of sample preparation methods of bacterial metabolomics used in both approaches are reviewed. The review also highlights previous literature of optimized conditions used to propose the most ideal methods for metabolite preparation, particularly for bacterial cells. Advantages and limitations of methods are discussed for future improvement of bacterial metabolomics.
Fulltext A study on volatile organic compounds emitted by in-vitro lung cancer cultured cells using gas sensor array and SPME-GCMS

Thriumani R, Zakaria A, Hashim YZH, Jeffree AI, Helmy KM, Kamarudin LM, et al.

BMC Cancer, 2018 04 02;18(1):362.
PMID: 29609557 DOI: 10.1186/s12885-018-4235-7

BACKGROUND: Volatile organic compounds (VOCs) emitted from exhaled breath from human bodies have been proven to be a useful source of information for early lung cancer diagnosis. To date, there are still arguable information on the production and origin of significant VOCs of cancer cells. Thus, this study aims to conduct in-vitro experiments involving related cell lines to verify the capability of VOCs in providing information of the cells.
METHOD: The performances of e-nose technology with different statistical methods to determine the best classifier were conducted and discussed. The gas sensor study has been complemented using solid phase micro-extraction-gas chromatography mass spectrometry. For this purpose, the lung cancer cells (A549 and Calu-3) and control cell lines, breast cancer cell (MCF7) and non-cancerous lung cell (WI38VA13) were cultured in growth medium.
RESULTS: This study successfully provided a list of possible volatile organic compounds that can be specific biomarkers for lung cancer, even at the 24th hour of cell growth. Also, the Linear Discriminant Analysis-based One versus All-Support Vector Machine classifier, is able to produce high performance in distinguishing lung cancer from breast cancer cells and normal lung cells.
CONCLUSION: The findings in this work conclude that the specific VOC released from the cancer cells can act as the odour signature and potentially to be used as non-invasive screening of lung cancer using gas array sensor devices.
Fulltext Fabrication and characterization of Agarwood extract-loaded nanocapsules and evaluation of their toxicity and anti-inflammatory activity on RAW 264.7 cells and in zebrafish embryos

Eissa MA, Hashim YZH, Mohd Nasir MH, Nor YA, Salleh HM, Isa MLM, et al.

Drug Deliv, 2021 Dec;28(1):2618-2633.
PMID: 34894947 DOI: 10.1080/10717544.2021.2012307

Aquilaria malaccensis has been traditionally used to treat several medical disorders including inflammation. However, the traditional claims of this plant as an anti-inflammatory agent has not been substantially evaluated using modern scientific techniques. The main objective of this study was to evaluate the anti-inflammatory effect of Aquilaria malacensis leaf extract (ALEX-M) and potentiate its activity through nano-encapsulation. The extract-loaded nanocapsules were fabricated using water-in-oil-in-water (w/o/w) emulsion method and characterized via multiple techniques including DLS, TEM, FTIR, and TGA. The toxicity and the anti-inflammatory activity of ALEX-M and the extract-loaded nanocapsules (ALEX-M-PNCs) were evaluated in-vitro on RAW 264.7 macrophages and in-vivo on zebrafish embryos. The nanocapsules demonstrated spherical shape with mean particle diameter of 167.13 ± 1.24 nm, narrow size distribution (PDI = 0.29 ± 0.01), and high encapsulation efficiency (87.36 ± 1.81%). ALEX-M demonstrated high viability at high concentrations in RAW 264.7 cells and zebrafish embryos, however, ALEX-M-PNCs showed relatively higher cytotoxicity. Both free and nanoencapsulated extract expressed anti-inflammatory effects through significant reduction of the pro-inflammatory mediator nitric oxide (NO) production in LPS/IFNγ-stimulated RAW 264.7 macrophages and zebrafish embryos in a concentration-dependent manner. The findings highlight that ALEX-M can be recognized as a potential anti-inflammatory agent, and its anti-inflammatory activity can be potentiated by nano-encapsulation. Further studies are warranted toward investigation of the mechanistic and immunomodulatory roles of ALEX-M.