Photoelectrochemical detection of microRNAs based on target-triggered self-assembly of energy band position-matched CdS QDs and C3N4 nanosheets.

An ultrasensitive photochemical biosensor based on the target miRNA-triggered catalytic hairpin assembly (CHA) reaction between Au nanoparticles (AuNPs)/C3N4 nanosheets and CdS quantum dots (QDs) was developed for the determination of miRNAs. Firstly, AuNPs/C3N4 nanosheets were immobilized onto a working glassy carbon electrode. Then, the hairpin probe 1 (H1) was loaded through Au-S bonding. Afterward, the unbound sites were blocked with 6-mercaptohexanol to avoid nonspecific adsorption. In the presence of the target miRNA, the CHA reaction between the H1 and hairpin probe 2-CdS QDs (H2-CdS QDs) could be triggered. As a result, the AuNPs/C3N4 nanosheet and CdS QDs were linked by the double helix structure H1-H2. Unlike the other CHA reactions, H2 used in this work is longer than H1 so that the AuNPs/C3N4 nanosheets could touch the CdS QDs. Given the matched energy band positions between the C3N4 nanosheet and CdS QDs, a strong photocurrent could be obtained after the CHA reaction was triggered by the target miRNA. In addition, p-type C3N4 nanosheets and n-type CdS QDs presented reduction photocurrents and oxidation photocurrents, respectively. Therefore, the photocurrents were vectors in this design that can eliminate the interference of nonspecific adsorption and avoid the generation of false-positive signals. Under the optimal conditions, the limit of detection was 92 aM. The constructed photoelectrochemical biosensor showed good reproducibility and selectivity in the analysis of serum samples, which indicates its great prospects in disease diagnostics and bioanalysis.

A label-free electrochemical platform based on a thionine functionalized magnetic Fe-N-C electrocatalyst for the detection of microRNA-21.

Taking a composite of a nanomaterial and a signal molecule as a substrate material can provide a label-free electrochemical platform. Besides, the nanomaterial with a high catalytic activity towards the signal molecule can improve the sensitivity of the platform. Herein, a thionine functionalized Fe-N-C nanocomposite was employed as the substrate. Firstly, the electrocatalytic activity of Fe-N-C towards the electroreduction of thionine was explored. Then, an immobilization-free and label-free electrochemical platform for the determination of microRNA-21 based on Fe-N-C-thionine/Fe3O4@AuNPs was constructed. A magnetic glassy carbon electrode (MGCE) was used to keep the magnetic Fe-N-C-thionine/Fe3O4@AuNPs modified onto the surface of the MGCE. Fe-N-C and Fe3O4 nanoparticles can co-catalyze the electroreduction of thionine and a strong electrochemical reduction signal of thionine could be realized in the differential pulse voltammetry (DPV) test. Also, a catalytic hairpin assembly (CHA) reaction was utilized to enhance the sensitivity of the developed electrochemical biosensor. Besides, the developed biosensor shows excellent specificity and reproducibility in the test of human serum samples, indicating its wide application prospects in the early-stage diagnosis of tumors.

Mitochondria-targeted delocalized lipophilic cation complexed with human serum albumin for tumor cell imaging and treatment.

Small molecule 5BMF is a novel mitochondria-targeted delocalized lipophilic cation (DLC) with good anti-tumor activity and fluorescence emission suitable for bioimaging. In this study, human serum albumin (HSA) complexed with 5BMF (5BMF@HSA) has been developed to further improve its solubility (from 1.61 to 5.41 mg/mL), and the fluorescent intensity of 5BMF@HSA was improved over 2 times. Nearly 10-fold 5BMF was released from 5BMF@HSA complex in acidic condition when compared with neutral/basic environment. Intracellular distribution of 5BMF was altered by HSA as its signals were observed in lysosomes where free 5BMF barely localized. Both 5BMF@HSA and 5BMF showed selective toxicity toward tumor cells in µM and nM range and effectively inhibited tumor growth in mice model. In summary, 5BMF@HSA shows improved solubility in aqueous buffer and enhanced fluorescence emission, and maintains tumor inhibition capability. It is a promising protein complex for tumor cell imaging and tumor treatment.

Anti-HIV Activity of Ocimum labiatum Extract and Isolated Pheophytin-a.

Ocimum plants are traditionally used to manage HIV/AIDS in various African countries. The effects of Ocimum labiatum extract on HIV-1 protease (PR) and reverse transcriptase (RT) is presented here along with characterization of an identified bioactive compound, achieved through ¹H- and 13C-NMR. The extract's effect on HIV-1 replication was assessed by HIV-1 p24 antigen capture. Cytotoxicity of samples was evaluated using tetrazolium dyes and real-time cell electronic sensing (RT-CES). Ocimum labiatum inhibited HIV-1 PR with an IC50 value of 49.8 ± 0.4 µg/mL and presented weak inhibition (21%) against HIV-1 RT. The extract also reduced HIV-1 replication in U1 cells at a non-cytotoxic concentration (25 µg/mL). The CC50 value of the extract in U1 cells was 42.0 ± 0.13 µg/mL. The HIV-1 PR inhibiting fraction was purified using prep-HPLC and yielded a chlorophyll derivative, pheophytin-a (phy-a). Phy-a inhibited HIV-1 PR with an IC50 value of 44.4 ± 1.5 µg/mL (51 ± 1.7 µM). The low cytotoxicity of phy-a in TZM-bl cells was detected by RT-CES and the CC50 value in U1 cells was 51.3 ± 1.0 µg/mL (58.9 ± 1.2 µM). This study provides the first in vitro evidence of anti-HIV activity of O. labiatum and isolated phy-a, supporting further investigation of O. labiatum for lead compounds against HIV-1.

Triterpenoids from Ocimum labiatum Activates Latent HIV-1 Expression In Vitro: Potential for Use in Adjuvant Therapy.

Latent HIV reservoirs in infected individuals prevent current treatment from eradicating infection. Treatment strategies against latency involve adjuvants for viral reactivation which exposes viral particles to antiretroviral drugs. In this study, the effect of novel triterpenoids isolated from Ocimum labiatum on HIV-1 expression was measured through HIV-1 p24 antigen capture in the U1 latency model of HIV-1 infection and in peripheral blood mononuclear cells (PBMCs) of infected patients on combination antiretroviral therapy (cART). The mechanism of viral reactivation was determined through the compound's effect on cytokine production, histone deacetylase (HDAC) inhibition, and protein kinase C (PKC) activation. Cytotoxicity of the triterpenoids was determined using a tetrazolium dye and flow cytometry. The isolated triterpene isomers, 3-hydroxy-4,6a,6b,11,12,14b-hexamethyl-1,2,3,4,6,6a,6b,7,8,8a,9,10,11,12,12a,14,14a,14b-octadecahydropicene-4,8a-dicarboxylic acid (HHODC), significantly (p < 0.05) induced HIV-1 expression in a dose-dependent manner in U1 cells at non-cytotoxic concentrations. HHODC also induced viral expression in PBMCs of HIV-1 infected patients on cART. In addition, the compound up-regulated the production of interleukin (IL)-2, IL-6, tumour necrosis factor (TNF)-α, and interferon (IFN)-γ but had no effect on HDAC and PKC activity, suggesting cytokine upregulation as being involved in latency activation. The observed in vitro reactivation of HIV-1 introduces the adjuvant potential of HHODC for the first time here.

In vitro anti-HIV and antioxidant activity of Hoodia gordonii (Apocynaceae), a commercial plant product.

BACKGROUND: Hoodia gordonii products are widely commercialized for anti-obesity purposes; however, minimal research is available on the other health properties demonstrated by this popular herbal plant. METHODS: H. gordonii crude extracts (ethanol and ethyl acetate) were assayed for in vitro anti-HIV-1 protease (PR), reverse transcriptase (RT) and integrase activity. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) and reducing power assays were used for the antioxidant analysis. In addition, qualitative and quantitative phytochemical analyses of the extracts were determined using standard methods. RESULTS: H. gordonii extract demonstrated good inhibition against HIV RT with IC50 values of 73.55 ± 0.04 and 69.81 ± 9.45 µg/mL for ethanol and ethyl acetate extracts, respectively. Both extracts also demonstrated inhibitory activity against HIV PR with IC50 values of 97.29 ± 0.01 and 63.76 ± 9.01 µg/mL for ethanol and ethyl acetate extracts. In addition, H. gordonii also showed good antioxidant activity with IC50 values of 124.6 ± 11.3 and 126.2 ± 3.15 µg/mL obtained for ethanol and ethyl acetate extracts, respectively. The reducing power of H. gordonii extracts increased as the concentration increased which confirmed the presence of antioxidants (reductants) in the extracts. Phytochemical screening of H. gordonii revealed the presence of phenolics, alkaloids, terpenes, steroids, cardiac glycosides and tannins in the ethanolic extract, while the ethyl acetate extract only showed the presence of phenolics, cardiac glycosides and steroids. The total phenolic content was 420 ± 0.17 and 319.9 ± 0.2 mg GAE/g for the ethanol and ethyl acetate extracts, respectively. The ethanol extract, which revealed the presence of tannins, had a tannin content of 330 ± 0.2 mg TAE/g extract. CONCLUSION: This data suggests that H. gordonii has good in vitro inhibition against selected HIV-1 enzymes as well as antioxidant properties, suggesting new potential uses for this commercial plant.

Tylosema esculentum (Marama) Tuber and Bean Extracts Are Strong Antiviral Agents against Rotavirus Infection.

Tylosema esculentum (marama) beans and tubers are used as food, and traditional medicine against diarrhoea in Southern Africa. Rotaviruses (RVs) are a major cause of diarrhoea among infants, young children, immunocompromised people, and domesticated animals. Our work is first to determine anti-RV activity of marama bean and tuber ethanol and water extracts; in this case on intestinal enterocyte cells of human infant (H4), adult pig (CLAB) and adult bovine (CIEB) origin. Marama cotyledon ethanolic extract (MCE) and cotyledon water extract (MCW) without RV were not cytotoxic to all cells tested, while seed coat and tuber extracts showed variable levels of cytotoxicity. Marama cotyledon ethanolic and water extracts (MCE and MCW, resp.) (≥0.1 mg/mL), seed coat extract (MSCE) and seed coat water extract (MSCW) (0.01 to 0.001 mg/mL), especially ethanolic, significantly increased cell survival and enhanced survival to cytopathic effects of RV by at least 100% after in vitro co- and pre-incubation treatments. All marama extracts used significantly enhanced nitric oxide release from H4 cells and enhanced TER (Ω/cm(2)) of enterocyte barriers after coincubation with RV. Marama cotyledon and seed coat extracts inhibited virion infectivity possibly through interference with replication due to accumulation of nitric oxide. Marama extracts are therefore promising microbicides against RV.

p-Co-BDC/AuNPs-based multiple signal amplification for ultra-sensitive electrochemical determination of miRNAs.

In this work, we report AuNPs-decorated pyrolyzed Co-BDC nanosheets (p-Co-BDC/AuNPs) as high-performance electrocatalyst for developing an electrochemical platform. p-Co-BDC/AuNPs as a new electrocatalyst showed superior electrocatalytic activity towards the electrochemical oxidation of methylene blue (MB). Besides, magnetic p-Co-BDC/AuNPs can be well immobilized on the magnetic glassy carbon electrode without further assistance. The oxidation of MB can be reduced by ascorbic acid. Inspired by this phenomenon, an electrochemical biosensor was constructed based on multiple signal amplification for the diagnosis of miRNAs. Firstly, p-Co-BDC/AuNPs enhanced the electrochemical oxidation of MB. Then, strand displacement amplification reaction can form lots of double helix structure DNA to embed more MB molecules. Finally, ascorbic acid in the electrolyte was utilized to reduce the oxidation of MB and improve the electrochemical signal of MB electro-oxidation. The linear detection range for the detection of miRNAs is 100 aM to 10 nM, and the limit of detection is 86 aM. Furthermore, the constructed biosensor also displayed satisfactory selectivity, good reproducibility, and excellent recovery in the detection of real samples. We are convinced that our proposed multiple signal amplification strategy will provide more promising methods for the diagnosis of cancer.

Electrochemical detection of microRNAs based on AuNPs/CNNS nanocomposite with Duplex-specific nuclease assisted target recycling to improve the sensitivity.

MicroRNAs (miRNAs) are important biomarkers in early diagnosis of disease. In this work, we developed a simple and effective electrochemical biosensor based on Au nanoparticles (AuNPs)/Carbon nitride nanosheet (CNNS) nanocomposite for the miRNAs detection. Duplex-specific nuclease (DSN) and hairpin structure probe were utilized to improve the sensitivity and selectivity respectively. In the presence of miRNA-21, the signal molecule could be released from the surface of the electrode and decrease the current peak in square wave voltammetry (SWV) test. Under the optimal conditions, the reported biosensor showed that the detection range of miRNA-21 is from 10 fM to 1 nM and detection limit is as low as 2.9 fM. Furthermore, the detection of miRNA-21 added in serum samples indicates the developed biosensor with good selectivity, stability and reproducibility which verify its potential to be used in the early diagnosis of diseases.

Tuning the near infrared II emitting wavelength of small molecule dyes by single atom alteration.

A series of small molecule dyes demonstrate the feasibility of manipulating Near Infrared II emission by simply altering the donors' heteroatoms, which involved both electronegativity and intramolecular steric effects. Furthermore, these dyes show high resolution and stability for in vivo imaging after being complexed with human serum albumin.

Characterization of Acanthosicyos horridus and Citrullus lanatus seed oils: two melon seed oils from Namibia used in food and cosmetics applications.

The physicochemical characteristics, fatty acid, tocopherol, stigmasterol, ß-sitosterol, and 1H NMR profiles of Citrullus lanatus and Acanthosicyos horridus melon seed oils were determined and compared among different extraction methods (cold pressing, traditional, and Soxhlet). The oil content was 40.2 ± 3.45 and 37.8 ± 7.26% for C. lanatus and A. horridus, respectively. Significant differences (p < 0.05) were observed among the different extraction methods in the characteristics studied. Physicochemical characteristics of the melon seed oils were saponification value, 180.48-189.86 mg KOH/g oil; iodine value, 108.27-118.62 g I2/100 g oil; acid value, 0.643-1.63 mg KOH/g oil; peroxide value; 1.69-2.98 mequiv/kg oil; specific gravity, 0.901-0.922; and refractive indices, 1.4676-1.4726. The dominant tocopherol was γ-tocopherol with total tocopherol in the range 27.61-74.39 mg/100 g. The dominant fatty acid was linoleic acid in the range 52.57-56.96%. The favorable oil yield, physicochemical characteristics, tocopherol, and fatty acid composition have the potential to replace or improve major commercial vegetable oils and to be used for various applications in the food industry and nutritive medicines.