Kefir is a homemade, natural fermented product comprised of a probiotic bacteria and yeast complex. Kefir consumption has been associated with many advantageous properties to general health, including as an antioxidative, anti-obesity, anti-inflammatory, anti-microbial, and anti-tumor moiety. This beverage is commonly found and consumed by people in the United States of America, China, France, Brazil, and Japan. Recently, the consumption of kefir has been popularized in other countries including Malaysia. The microflora in kefir from different countries differs due to variations in culture conditions and the starter media. Thus, this study was aimed at isolating and characterizing the lactic acid bacteria that are predominant in Malaysian kefir grains via macroscopic examination and 16S ribosomal RNA gene sequencing. The results revealed that the Malaysian kefir grains are dominated by three different strains of Lactobacillus strains, which are Lactobacillus harbinensis, Lactobacillusparacasei, and Lactobacillus plantarum. The probiotic properties of these strains, such as acid and bile salt tolerances, adherence ability to the intestinal mucosa, antibiotic resistance, and hemolytic test, were subsequently conducted and extensively studied. The isolated Lactobacillus spp. from kefir H maintained its survival rate within 3 h of incubation at pH 3 and pH 4 at 98.0 ± 3.3% and 96.1 ± 1.7% of bacteria growth and exhibited the highest survival at bile salt condition at 0.3% and 0.5%. The same isolate also showed high adherence ability to intestinal cells at 96.3 ± 0.01%, has antibiotic resistance towards ampicillin, penicillin, and tetracycline, and showed no hemolytic activity. In addition, the results of antioxidant activity tests demonstrated that isolated Lactobacillus spp. from kefir G possessed high antioxidant activities for total phenolic content (TPC), total flavonoid content (TFC), ferric reducing ability of plasma (FRAP), and 1,1-diphenyl-2-picryl-hydrazine (DPPH) assay compared to other isolates. From these data, all Lactobacillus spp. isolated from Malaysian kefir serve as promising candidates for probiotics foods and beverage since they exhibit potential probiotic properties and antioxidant activities.

Patients with irritable bowel syndrome (IBS) are increasingly presenting with a wide range of neuropsychiatric symptoms, such as deterioration in gastroenteric physiology, including visceral hypersensitivity, altered intestinal membrane permeability, and gastrointestinal motor dysfunction. Functional imaging of IBS patients has revealed several abnormalities in various brain regions, such as significant activation of amygdala, thinning of insular and anterior cingulate cortex, and increase in hypothalamic gray matter, which results in poor psychiatric and cognitive outcomes. Interrelations between the enteric and central events in IBS-related gastrointestinal, neurological, and psychiatric pathologies have compelled researchers to study the gut-brain axis—a bidirectional communication that maintains the homeostasis of the gastrointestinal and central nervous system with gut microbiota as the protagonist. Thus, it can be disrupted by any alteration owing to the gut dysbiosis or loss of diversity in microbial composition. Available evidence indicates that the use of probiotics as a part of a balanced diet is effective in the management of IBS and IBS-associated neurodegenerative and psychiatric comorbidities. In this review, we delineate the pathogenesis and complications of IBS from gastrointestinal and neuropsychiatric standpoints while also discussing the neurodegenerative events in enteric and central nervous systems of IBS patients and the therapeutic potential of gut microbiota-based therapy established on clinical and preclinical data.

Citral is an active compound naturally found in lemongrass, lemon, and lime. Although this pale-yellow liquid confers low water solubility, the compound has been reported to possess good therapeutic features including antiproliferative and anticancer modalities. The self nano-emulsifying drug delivery system (SNEDDS) is a type of liquid-lipid nanocarrier that is suitable for the loading of insolubilized oil-based compound such as Citral. This study reports the design and optimization of a SNEDDS formulation, synthesis and characterization as well as loading with Citral (CIT-SNEDDS). Further assessment of theantiproliferative effects of CIT-SNEDDS towards colorectal cancer cells was also conducted. SNEDDS composed of coconut oil, dimethyl sulfoxide (DMSO) and Tween 80. CIT-SNEDDS was prepared via gentle agitation of SNEDDS with 0.5% Citral for 72 h at room temperature. Physicochemical characterization was performed using several physicochemical analyses. The average particle size of CIT-SNEDDS was16.86 ± 0.15 nm, zeta potential of 0.58 ± 0.19 mV, and polydispersity index (PDI) of 0.23 ± 0.01. In vitro drug release of Citral from CIT-SNEDDS was 79.25% of release, and for Citral the release percentage was 93.56% over 72 h. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was done to determine the cytotoxicity effect of CIT-SNEDDS in human colorectal cancer cell lines HT29 and SW620. The half maximal inhibitory concentrations (IC₅₀) for 72 hof CIT-SNEDDS and Citral on SW620 were 16.50 ± 0.87 µg/mL and 22.50 ± 2.50 µg/mL, respectively. The IC₅₀ values of CIT-SNEDDS and Citral after 72 h of treatment on HT29 were 34.10 ± 0.30 µg/mL and 21.77 ± 0.23 µg/mL, respectively. This study strongly suggests that CIT-SNEDDS has permitted the sustained release of Citral and that CIT-SNEDDS constitutes a potential soluble drug nanocarrier that is effective against colorectal cancer cells.

Background: Eucalyptol is an active compound of eucalyptus essential oil and was reported to have many medical attributes including cytotoxic effect on breast cancer cells. However, it has low solubility in aqueous solutions which limits its bioavailability and cytotoxic efficiency. In this study, nanostructured lipid carrier loaded with eucalyptol (NLC-Eu) was formulated and characterized and the cytotoxic effect of NLC-Eu towards breast cancer cell lines was determined. In addition, its toxicity in animal model, BALB/c mice was also incorporated into this study to validate the safety of NLC-Eu. Methods: Eucalyptol, a monoterpene oxide active, was used to formulate the NLC-Eu by using high pressure homogenization technique. The physicochemical characterization of NLC-Eu was performed to assess its morphology, particle size, polydispersity index, and zeta potential. The in vitro cytotoxic effects of this encapsulated eucalyptol on human (MDA MB-231) and murine (4 T1) breast cancer cell lines were determined using the MTT assay. Additionally, acridine orange/propidium iodide assay was conducted on the NLC-Eu treated MDA MB-231 cells. The in vivo sub-chronic toxicity of the prepared NLC-Eu was investigated using an in vivo BALB/c mice model. Results: As a result, the light, translucent, milky-colored NLC-Eu showed particle size of 71.800 ± 2.144 nm, poly-dispersity index of 0.258 ± 0.003, and zeta potential of − 2.927 ± 0.163 mV. Furthermore, the TEM results of NLC-Eu displayed irregular round to spherical morphology with narrow size distribution and relatively uniformed particles. The drug loading capacity and entrapment efficiency of NLC-Eu were 4.99 and 90.93%, respectively. Furthermore, NLC-Eu exhibited cytotoxic effects on both, human and mice, breast cancer cells with IC50 values of 10.00 ± 4.81 μg/mL and 17.70 ± 0.57 μg/mL, respectively at 72 h. NLC-Eu also induced apoptosis on the MDA MB-231 cells. In the sub-chronic toxicity study, all of the studied mice did not show any signs of toxicity, abnormality or mortality. Besides that, no significant changes were observed in the body weight, internal organ index, hepatic and renal histopathology, serum biochemistry, nitric oxide and malondialdehyde contents. Conclusions: This study suggests that the well-characterized NLC-Eu offers a safe and promising carrier system which has cytotoxic effect on breast cancer cell lines.

Osteosarcoma is one of the primary malignant bone tumors that confer low survival rates for patients even with intensive regime treatments. Therefore, discovery of novel anti-osteosarcoma drugs derived from natural products that are not harmful to the normal cells remains crucial. Curcumin is one of the natural substances that have been extensively studied due to its anti-cancer properties and is pharmacologically safe considering its ubiquitous consumption for centuries. However, curcumin suffers from a poor circulating bioavailability, which has led to the development of a chemically synthesized curcuminoid analog, namely (Z)-3-hydroxy-1-(2-hydroxyphenyl)-3-phenylprop-2-en-1-one (DK1). In this study, the cytotoxic effects of the curcumin analog DK1 was investigated in both U-2OS and MG-63 osteosarcoma cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and cell death was microscopically examined via acridine orange/propidium iodide (AO/PI) double staining. Flow cytometer analysis including Annexin V/Fluorescein isothiocyanate (FITC), cell cycle analysis and JC-1 were adapted to determine the mode of cell death. Subsequently in order to determine the mechanism of cell death, quantitative polymerase chain reaction (qPCR) and proteome profiling was carried out to measure the expression of several apoptotic-related genes and proteins. Results indicated that DK1 induced U-2 OS and MG-63 morphological changes and substantially reduced cell numbers through induction of apoptosis. Several apoptotic genes and proteins were steadily expressed after treatment with DK1; including caspase 3, caspase 9, and BAX, which indicated that apoptosis occurred through a mitochondria-dependent signaling pathway. In conclusion, DK1 could be considered as a potential candidate for an anti-osteosarcoma drug in the near future, contingent upon its ability to induce apoptosis in osteosarcoma cell lines.

Colorectal cancer (CRC) is the third most common type of cancer worldwide and a leading cause of cancer death. According to the Malaysian National Cancer Registry Report 2012–2016, colorectal cancer was the second most common cancer in Malaysia after breast cancer. Recent treatments for colon cancer cases have caused side effects and recurrence in patients. One of the alternative ways to fight cancer is by using natural products. Curcumin is a compound of the rhizomes of Curcuma longa that possesses a broad range of pharmacological activities. Curcumin has been studied for decades but due to its low bioavailability, its usage as a therapeutic agent has been compromised. This has led to the development of a chemically synthesized curcuminoid analogue, (2E,6E)-2,6-bis(2,3-dimethoxybenzylidine) cyclohexanone (DMCH), to overcome the drawbacks. This study aims to examine the potential of DMCH for cytotoxicity, apoptosis induction, and activation of apoptosis-related proteins on the colon cancer cell lines HT29 and SW620. The cytotoxic activity of DMCH was evaluated using the [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) cell viability assay on both of the cell lines, HT29 and SW620. To determine the mode of cell death, an acridine orange/propidium iodide (AO/PI) assay was conducted, followed by Annexin V/FITC, cell cycle analysis, and JC-1 assay using a flow cytometer. A proteome profiler angiogenesis assay was conducted to determine the protein expression. The inhibitory concentration (IC₅₀) of DMCH in SW620 and HT29 was 7.50 ± 1.19 and 9.80 ± 0.55 µg/mL, respectively. The treated cells displayed morphological features characteristic of apoptosis. The flow cytometry analysis confirmed that DMCH induced apoptosis as shown by an increase in the sub-G0/G1 population and an increase in the early apoptosis and late apoptosis populations compared with untreated cells. A higher number of apoptotic cells were observed on treated SW620 cells as compared to HT29 cells. Human apoptosis proteome profiler analysis revealed upregulation of Bax and Bad proteins and downregulation of Livin proteins in both the HT29 and SW620 cell lines. Collectively, DMCH induced cell death via apoptosis, and the effect was more pronounced on SW620 metastatic colon cancer cells, suggesting its potential effects as an antimetastatic agent targeting colon cancer cells.