An exploratory investigation into the physicochemical, antioxidant and cellular effects of a selection of honey samples from the Southern African region

The unique floral biodiversity of Southern Africa would be reflected in the phenolic acid and flavonoid composition as well as the antioxidant activity of honeys from this region. In this exploratory investigation the total polyphenolic (TPC) and flavonoid (TFC) content, antioxidant activity as well as the cellular protective effects of a selection of honeys collected in this region was evaluated. Thirteen honey samples representative of the Western Cape (WCa, WCb and WCc), Eastern Cape (ECa, ECb and ECc), South East Mozambique (SEMa, SEMb and SEMc) and Agricultural: A-E (Eucalyptus) (A-E1 and A-E2), A-L (Litchi) and A-O (Orange) were collected. These samples were subjected to physicochemical analysis, the antioxidant content (TPC and TFC) and both enzymatic (catalase activity) and non-enzymatic activity, using the 2,2-diphenyl-2-picrylhydrazyl (DPPH), trolox equivalent antioxidant capacity (TEAC) and oxygen radical antioxidant capacity (ORAC) assays was determined. From the DPPH, TEAC and ORAC data the Relative Antioxidant Capacity Index (RACI) was calculated. To determine whether high antioxidant activity translates into significant cellular protection, biological and cellular assays were undertaken. Using the pBR322 plasmid assay and the erythrocyte haemolysis assay the ability of honeys to protect against 2,2’-Azobis(2-amidinopropane) dihydrochloride (AAPH) oxidative damage was evaluated. Further evaluation was undertaken in the SC-1 fibroblast cell line and the physiologically more relevant Caco-2 cell line. Toxicity and antioxidant effects were evaluated in the SC-1 cell line while antioxidant effects were only evaluated in the Caco-2 cell line. The long-term mitogenic and toxic effects were determined in the SC-1 cell line using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Neutral Red (NR) and Crystal Violet (CV) assays. Short term, total- and intracellular antioxidant effects were determined in both cell lines using the dichlorofluorescein diacetate assay (DCFH-DA) assay. For all cellular experiments honey at concentrations of 0.01% and 1% were used. The physiochemical properties of the honeys evaluated fulfilled the regulatory standards compiled in the Codex Alimentarius (CODEX STAN12-1981 revision 2001). The results were as follows: SEMb had the highest TPC (167.96 mg GAE/100g) and TFC (51.60 mg CE/100g) while A-E2 had the highest catalase (38.48 µmol H2O2/g) activity. RACI revealed that WCb had the highest antioxidant activity.SEMc showed the highest protection of plasmid DNA against oxidative-induced strand breaks while SEMa showed the highest protection of erythrocytes against AAPH-induced haemolysis. Although correlations were found between antioxidant content and antioxidant activity assays, no correlation was found these parameters and the biological assays. For the long-term cytotoxicity assay, AAPH showed significant cytoxicity at 0.78mM, 1.56mM and 0.28mM when measured using the MTT, NR and CV assays, respectively. Some honeys 4/13 and 3/13 showed a mitogenic effect at a concentration of 0.01% and 1% respectively. Toxic effects, were observed for 1/13 and 8/13 at 0.01% and 1% honey respectively. Toxicity after 72 h exposure varied from 10-30% (CV assay). The same concetrations of honey was used to determine the short-term, 2h, antioxidant effects in both the SC-1 and Caco-2 cell lines. No oxidative effect was found for all honeys at these concentrations. For the DCFH-DA assay using the SC-1 cell line at 1%, 12/13 and 7/13 honeys showed total and intracellular protection respectively. The highest extracellular protection was for SEMa (% Protection (%P) = 95) and SEMb (%P = 93). Intracellular protection was the highest for SEMc (%P = 21) and A-L (%P = 20). At 0.01%, 7/13 and 8/13 honeys exhibited total and intracellular protection, respectively. For both the highest protection was found for SEMc (%P = 43, total and %P = 30, intracellular). For the Caco-2 cell line at 1%, 11/13 and 4/13 showed total and intracellular protection, respectively. Of these the highest extracellular protection was for SEMb (% Protection (%P) = 90). Intracellular protection was the highest for ECa (%P = 28) and WCc (%P = 26). At 0.01%, 4/13 and 8/13 honeys showed total and intracellular protection respectively. The highest extracellular protection was found for SEMc (%P = 62) and intracellular protection was ECc (%P = 28). The SC-1 cell line was found to be the most sensitive to the antioxidant effects of honey compared to the Caco-2 cell line. The honeys SEMa, SEMb and SEMc showed protection against oxidative damage in both cell lines. In conclusion, the antioxidant activity of honeys from Southern Africa is of a high quality. The WC, SEM and EC honeys showed the highest antioxidant effects and could provide health benefits against diseases associated with oxidative stress as indicated by these results. CopyrightDissertation (MSc)--University of Pretoria, 2011.Anatomyunrestricte

Stability, morphology, and effects of in vitro digestion on the antioxidant properties of polyphenol inclusion complexes with -cyclodextrin

DATA AVAILABILTY STATEMENT : Some data is available as supplementary material and the rest under the UPSpace institutional repository, URI: http://hdl.handle.net/2263/84453 (date dissertation was available online 11 March 2022).SUPPLEMENTARY MATERIAL : FIGURE S1: Enhanced product ions scan mass spectrum of CD inclusion complexes with (a) CAT, (b) GA, and (c) EGCG. CD—beta cyclodextrin, CAT—catechin, GA—gallic acid, EGCG—epigallocatechin gallate; FIGURE S2: Inhibition of AGEs formation by non-digested (ND) and following simple (SD) and complex (CD) digestion of nonencapsulated and encapsulated CAT/GA, CAT/EGCG, and GA/EGCG combinations evaluated with the (a) BSA-MGO and (b) BSA-FRU models at 100 M for each polyphenol in each sample. The data is represented as the mean SEM of at least five experiments performed in duplicates. ND—nondigested, SD—simple digestion, CD—complex digestion, AGEs—advanced glycation end-products, BSA—bovine serum albumin, MGO—methylglyoxal, FRU—fructose, CAT—catechin, GA—gallic acid, EGCG—epigallocatechin gallate. The * and + represent significant (p < 0.05) differences between non-encapsulated and encapsulated ND compared with the respective SD or CD samples. The denotes significant (p < 0.05) differences between the non-encapsulated and encapsulated for each treatment; TABLE S1: Polyphenol–polyphenol interactions on antioxidant activity (ORAC assay ( M TE)); TABLE S2: Polyphenol–polyphenol interactions on antiglycation activity (AGEs assay (%)); TABLE S3: Polyphenol–polyphenol interactions on cellular antioxidant activity (DCFH-DA assay (%)).Polyphenols are inversely associated with the incidence of chronic diseases, but therapeutic use is limited by poor stability and bioaccessibility. Encapsulation has been shown to overcome some of these limitations. A selection of polyphenols (catechin, gallic acid, and epigallocatechin gallate) and their combinations were encapsulated in beta-cyclodextrin (CD). Encapsulation was characterized and the thermal and storage stability was evaluated using the 2,2-azinobis (3-ethylbenzothiazoline- 6-sulfonic acid) (ABTS) assay. The samples were then subjected to in vitro digestion using a simple digestion (SD) model (gastric and duodenal phases) and a more complex digestion (CD) model (oral, gastric, and duodenal phases). Thereafter, the chemical (oxygen radical absorbance capacity assay) and cellular (dichlorofluorescein diacetate assay in Caco-2 cells) antioxidant and antiglycation (advanced glycation end-products assay) activities were determined. Inclusion complexes formed at a 1:1 molar ratio with a high encapsulation yield and efficiency. Encapsulation altered the morphology of the samples, increased the thermal stability of some and the storage stability of all samples. Encapsulation maintained the antioxidant activity of all samples and significantly improved the antiglycation and cellular antioxidant activities of some polyphenols following SD. In conclusion, the formed inclusion complexes of CD with polyphenols had greater storage stability, without altering the beneficial cellular effects of the polyphenols.The National Research Foundation South Africa and the APC was funded by the University of Pretoria, Faculty of Health Sciences Library and the University of Pretoria, Faculty of Health Sciences.https://www.mdpi.com/journal/moleculesam2023AnatomyPharmacolog

An in vitro investigation of l-kynurenine, quinolinic acid, and kynurenic acid on B16 F10 melanoma cell cytotoxicity and morphology

DATA AVAILABILITY STATEMENT : The data that support the findings of this study are available from the corresponding author upon reasonable request.The metastatic behavior of melanoma has accentuated the need for specific therapy targets. Compounds, namely L‐kynurenine (L‐kyn), quinolinic acid (Quin), and kynurenic acid (KA) previously displayed antiproliferative and cytotoxic effects in vitro against cancer cells. Despite the growing interest in these compounds there are limited studies examining the in vitro effects on melanoma. In B16 F10 melanoma cells, RAW 264.7 macrophage cells, and HaCat keratinocyte cells, postexposure to the compounds, crystal violet staining was used to determine the half‐maximal inhibitory concentration (IC50), whereas polarization‐optical transmitted light differential interference contrast and light microscopy after hematoxylin and eosin (H&E) staining was used to assess morphological changes. L‐kyn, Quin, and KA‐induced cytotoxicity in all cell lines, with L‐kyn being the most cytotoxic compound. L‐kyn and KA at IC50‐induced morphological changes in B16 F10, RAW 264.7, and HaCat cell lines, whereas Quin had effects on B16 F10 and RAW 264.7 cells but did not affect HaCat cells. L‐kyn, Quin, and KA each display different levels of cytotoxicity, which were cell line specific. L‐kyn was shown to be the most potent compound against all cell lines and may offer future treatment strategies when combined with other viable treatments against melanoma.The University of Pretoria and the National Research Foundation (NRF).wileyonlinelibrary.com/journal/cbfam2024AnatomyPhysiologySDG-03:Good heatlh and well-bein

Chemokines as possible therapeutic targets in metastatic melanoma

DATA AVAILABILITY STATEMENT : Data sharing not applicable to this article as no datasets were generated or analysed during the current study.BACKGROUND : Cutaneous melanoma is a relentless form of cancer which continues to rise in incidence. Currently, cutaneous melanoma is the leading cause of skin cancer-related mortality, which can mainly be attributed to its metastatic potential. The activation of chemokine axes is a major contributor to melanoma metastasis through its involvement in promoting tumour cell migration, proliferation, survival, and adhesion. This review will focus on the role of chemokines in melanoma and possible therapeutic strategies to alter chemokine activation and subsequently inhibit the activation of signalling cascades that may promote metastasis. METHODS : A literature review was conducted to evaluate chemokines as possible therapeutic targets in metastatic melanoma. RESULTS : The crosstalk between signalling pathways and immune responses in the melanoma microenvironment resembles a complex and dynamic system. Therefore, the involvement of governing chemokine axes in the promotion of cutaneous and metastatic melanoma demands a proper understanding of the tumour microenvironment in order to identify possible targets and develop appropriate treatments against melanoma. CONCLUSION : Even though chemokine axes are regarded as promising therapeutic targets, it has become increasingly evident that chemokines can play a critical role in both tumour inhibition and promotion. The inhibition of chemokine axes to inhibit signalling cascades in target cells that regulate metastasis should, therefore, be carefully approached.The National Research Foundation (NRF); the School of Medicine Research Committee (RESCOM) and the University of Pretoria.http://www.wileyonlinelibrary.com/journal/cam4hj2023AnatomyPhysiolog

The tryptophan–kynurenine pathway in immunomodulation and cancer metastasis

DATA AVAILABILITY STATEMENT : Data sharing not applicable to this article as no datasets were generated or analysed during the current study.INTRODUCTION : The activation of the kynurenine pathway in cancer progression and metastasis through immunomodulatory pathways has drawn attention to the potential for kynurenine pathway inhibition. The activation of the kynurenine pathway, which results in the production of kynurenine metabolites through the degradation of tryptophan, promotes the development of intrinsically malignant properties in cancer cells while facilitating tumour immune escape. In addition, kynurenine metabolites act as biologically active substances to promote cancer development and metastasis. METHODS : A literature review was conducted to investigate the role of the tryptophan-kynurenine pathway in immunomodulation and cancer metastasis. RESULTS : Evidence suggests that several enzymes and metabolites implicated in the kynurenine pathway are overexpressed in various cancers. As such, the tryptophan pathway represents a promising target for cancer treatment. However, downstream signalling pathways, including aryl hydrocarbon receptor activation, have previously induced diverse biological effects in various malignancies, which resulted in either the promotion or the inhibition of metastasis. CONCLUSION : As a result, a thorough investigation of the kynurenine pathway and its regulatory mechanisms is necessary in order to properly comprehend the effects of kynurenine pathway activation involved in cancer development and metastasis.The University of Pretoria, School of Medicine Research Committee (RESCOM) and National Research Foundation (NRF).http://wileyonlinelibrary.com/journal/cam4hj2023AnatomyPhysiolog

L-kynurenine and quinolinic acid inhibited markers of cell survival in B16 F10 melanoma cells in vitro

DATA AVAILABILITY STATEMENT : The data that support the findings of this study are available from the corresponding author upon reasonable request.Melanoma is an aggressive malignancy and remains a major cause of skin cancer mortality, highlighting the need for new treatment strategies. Recent findings revealed that L-kynurenine and quinolinic acid induce cytotoxicity and morphological changes in B16 F10 melanoma cells in vitro. This paper highlights the effects of L-kynurenine and quinolinic acid at previously determined half-maximal inhibitory concentrations on cell cycle progression, cell death and extracellular signal-regulated protein kinase inhibition. Melanoma, B16 F10 and murine macrophages, RAW 264.7 cells were used in this study, as both cell lines express all the enzymes associated with the kynurenine pathway. Post exposure to the compounds at half-maximal inhibitory concentrations, transmission electron microscopy was used to assess intracellular morphological changes. Flow cytometry was used to analyse cell cycle progression and quantify apoptosis via the dual staining of Annexin V and propidium iodide and cell survival via extracellular signal-regulated protein kinase. L-kynurenine and quinolinic acid at half-maximal inhibitory concentrations induced intracellular morphological changes representative of cell death. Flow cytometry revealed alterations in cell cycle distribution, increased apoptosis and significantly inhibition of cell survival. L-kynurenine and quinolinic acid are exogenous kynurenine compounds which inhibited cell survival through extracellular signal-regulated protein kinase inhibition, induced cell cycle alterations and induced apoptosis in B16 F10 melanoma cells. SIGNIFICANCE STATEMENT : Data obtained in the current study revealed information regarding the in vitro effects of L-kynurenine and quinolinic acid on B16 F10 melanoma cell cycle progression, intracellular morphology, extracellular signal-regulated kinase 1/2 inhibition and cell death. Evaluating the therapeutic effects of kynurenine metabolites against melanoma may potentially result in improved future cancer treatment options to manage the global melanoma burden.National Research Foundation; Struwig-Germeshuysen Kankernavorsingstrust; RESCOM; RDP.https://onlinelibrary.wiley.com/journal/10958355hj2024AnatomyPhysiologySDG-03:Good heatlh and well-bein

The potential antidiabetic properties of green and purple tea [Camellia sinensis (L.) O Kuntze], purple tea ellagitannins, and urolithins

DATA AVAILABILITY : Data will be made available on request.ETHNOPHARMACOLOGICAL RELEVANCE : Tea (Camellia sinensis) has been consumed for centuries as traditional medicine for various diseases, including diabetes. The mechanism of action of many traditional medicines, including tea, often requires elucidation. Purple tea is a natural mutant of Camellia sinensis, grown in China and Kenya, and is rich in anthocyanins and ellagitannins. AIM OF THE STUDY : Here we aimed to determine whether commercial green and purple teas are a source of ellagitannins and whether green and purple teas, purple tea ellagitannins and their metabolites urolithins have antidiabetic activity. MATERIALS AND METHODS : Targeted UPLC-MS/MS was employed to quantify the ellagitannins corilagin, strictinin and tellimagrandin I, in commercial teas. The inhibitory effect of commercial green and purple teas and purple tea ellagitannins was evaluated on α-glucosidase and α-amylase. The bioavailable urolithins were then investigated for additional antidiabetic effects, by evaluating their effect on cellular glucose uptake and lipid accumulation. RESULTS : Corilagin, strictinin and tellimagrandin I (ellagitannins) were identified as potent inhibitors of α-amylase and α-glucosidase, with Ki values significantly lower (p 0.05) as metformin in increasing glucose uptake in adipocytes, muscle cells and hepatocytes. In addition, similar (p > 0.05) to metformin, both urolithin A and urolithin B reduced lipid accumulation in adipocytes and hepatocytes. CONCLUSIONS : This study identified green-purple teas as an affordable widely available natural source with antidiabetic properties. Furthermore, additional antidiabetic effects of purple tea ellagitannins (corilagin, strictinin and tellimagrandin I) and urolithins were identified.The University of Pretoria.α-glucosidase inhibitionam2024AnatomyBiochemistryGeneticsMicrobiology and Plant PathologyPharmacologySDG-03:Good heatlh and well-bein

Gastrointestinal effects on the antioxidant and immunomodulatory properties of South African fynbos honey

DATA AVAILABILITY STATEMENT: Data is available on request from the corresponding author.Please read abstract in article.The Department of Anatomy, University of Pretoria. Open Access funding is enabled and organized by SANLiC Gold.http://www.hindawi.com/journals/ijfs/AnatomySDG-03:Good heatlh and well-beingSDG-15:Life on lan

Antioxidant and anti-inflammatory properties of Ilex guayusa tea preparations : a comparison to Camellia sinensis teas

Ilex guayusa tea preparations are now commercially available as Runa tea. Little is known regarding the antioxidant and anti-inflammatory bioactivities of this tea. The I. guayusa teas had a total polyphenolic content between 54.39 and 67.23 mg GAE/g dry mass and peroxyl radical scavenging capacities between 1773.41 and 2019 µmol TE/g dry mass, nearly half of that for the Camellia sinensis teas. The I. guayusa teas afforded 60-80% protection from oxidative stress in the Caco-2 cellular antioxidant assay, comparable to the C. sinensis teas. The anti-inflammatory activity in lipopolysaccharide-stimulated RAW 264.7 cells of I. guayusa teas was similarly comparable to the C. sinensis teas with nitric oxide production reduced by 10-30%. Major compounds identified by mass spectrometry were the phenolic mono- and dicaffeoylquinic acid derivatives. I. guayusa teas are a good source of dietary phenolic compounds with cellular antioxidant and anti-inflammatory properties.http://pubs.rsc.org/en/journals/journalissues/fo#!recentarticles&adv2018-12-13hj2017AnatomyBiochemistr

The dipeptidyl peptidase IV inhibitory activity and multifunctional antidiabetic properties of SQSPA : structure – activity relationship evaluated with alanine scanning

Type 2 diabetes is a multifactorial disease and drugs with multifunctional properties are required. The peptide, SQSPA, was reported to be a potent and gastrointestinally stable α-glucosidase inhibitory peptide. In this study, the structure-activity relationship of this peptide was studied using alanine scanning. Four analogs; AQSPA, SASPA, SQAPA and SQSAA were designed and investigated for multifunctional antidiabetic effects. Molecular docking studies on human dipeptidyl peptidase-IV (DPP-IV) suggested that the binding affinities were in the order; AQSPA>SASPA>SQSPA>SQSAA>SQAPA while for in vitro DPP-IV inhibitory activity, it was SQSPA>SQSAA>AQSPA>SASPA>SQAPA. Enzyme kinetic studies revealed that the peptides are uncompetitive inhibitors with the exception of SQSAA and SQSPA. In 3T3-L1 differentiated adipocytes, SASPA was the only analog that significantly (p < 0.05) reduced and prevented lipid accumulation and did not induce cytotoxicity to differentiated 3T3-L1 cells. All peptides, especially SASPA scavenged methylglyoxal and peroxyl radicals thereby preventing advanced glycosylated end products formation and oxidative stress. The nitric oxide scavenging activity of all peptides was comparable to IPI and glutathione. Findings indicate that the amide side chain of Q2 is probably the most critical functional group for modulating the multifunctional antidiabetic effects of SQSPA while SASPA has been identified, as a novel peptide with enhanced multifunctional antidiabetic activity.The National Research Foundation of South Africa and the University of Pretoria.http://www.elsevier.com/locate/ijbiomac2021-10-01hj2020AnatomyBiochemistryGeneticsMicrobiology and Plant Patholog