Supramolecular biomimetic binding of the DNA-dye Hoechst 33258 by a synthetic macrocycle

Dissertação de mest., Ciências Biomédicas, Faculdade de Ciências e Tecnologia, Univ. do Algarve, 2010The supramolecular interaction between the well-known DNA-binder Hoechst 33258 dye (guest) and cucurbit[7]uril (CB[7], host) was investigated in detail. The formation of the 1:1 complexes was verified by various methodologies such as Job’s plot method and electrospray ionization mass spectrometry. The binding is characterized by a high association constant, K = 3.5 × 106 M-1, which was determined by UV-Vis and fluorescence titrations. The formation of supramolecular 1:1 complexes is accompanied by drastic changes in the fluorescence quantum yield (Φf) of the dye. While the free dye is barely fluorescent in water at pH 7 (Φf= 0.01), the emission is considerably increased in the host-guest assembly (Φf= 0.74). The binding characteristics at pH 4.5 are similar to the ones at pH 7. However, the fluorescence enhancement at the former pH is less dramatic, because the free dye has here already its highest emission (Φf= 0.29). Proton magnetic resonance spectroscopy clearly demonstrated that CB[7] is accommodated around the piperazine-benzimidazole unit. A mechanism that explains this peculiar fluorescence behavior, which involves intramolecular proton transfer and charge transfer, internal rotation of the free and complexed dye is proposed. The competitive binding to CB[7] of several polyamines (putrescine, cadaverine, spermidine, spermine, and histamine) towards Hoechst 33258 was evaluated as well. For some of these polyamines the CB[7] binding constant has been determined the first time to the best of my knowledge. Finally, the well-characterized binding of the dye with calf-thymus DNA was used in competition experiments. It was found that the known unspecific binding of the dye to DNA leads to strong fluorescence quenching, which becomes appreciable through the initially high fluorescence of the dye in its CB[7] complex. Hence, the strongly emissive Hoechst 33258/CB[7] platform may be further used for the characterization of unspecific DNA binding through fluorescence

A supramolecular keypad lock

The reversible photoswitching between an anthracene derivative and its [4+4] dimer, using the template effect of the CB8 macrocycle, was demonstrated. This example of supramolecular chemistry in water was harnessed to demonstrate the operation of a keypad lock device that is driven by means of light and chemicals as inputs

Cucurbiturils as supramolecular inhibitors of DNA restriction by type II endonucleases

Cucurbiturils (CB6 and CB7) were shown to inhibit the enzymatically catalyzed restriction of plasmids and linear DNA. This effect can be inverted by supramolecular masking of the macrocycles through competitive complexation with polyamines. These experiments provide supramolecular control of biocatalytic processes.Spanish MINECO [CTQ2011-28390]; FEDER; COST [CM1005]; DFG [NA-686/5]; Portuguese FCT [SFRH/BD/81628/2011, PEst-OE/EQB/LA0023/2013

A photoinduced pH jump applied to drug release from cucurbit[7]uril

A proof-of-principle for the application of a photoinduced pH jump for delivery of the Hoechst 33258 drug by disassembly of its host-guest complex with cucurbit[7]uril is described

An aminonaphthalimide-putrescine conjugate as fluorescent probe for cucurbituril host-guest complexes

A fluorophore-anchor dye based on the 4-amino-1,8-naphthalimide chromophore was designed and characterised with respect to its ability of forming supramolecular hostguest complexes with cucurbit[6]uril (CB6) and cucurbit[7]uril (CB7) hosts. It was found that CB6 encapsulates the anchor of the dye with a high binding constant [K = (1.11.4) X 107 M- 1], which was independent of acidic or neutral pH conditions. The 1:1 binding was accompanied by fluorescence quenching (ca. 20%) and a bathochromic shift (?? = +22 nm) of the charge-transfer absorption band of the dye. This is indicative of the occurrence of hydrogen bonding between a carbonyl-lined host portal and the aromatic NH group of the guest dye. In comparison, CB7 encapsulated the dye much less efficiently (K = 4.8 X 104 M- 1) and showed a significant fluorescence enhancement (Ff = 0.15 versus 0.52 for free and CB7-complexed dye at pH 4, respectively). The occurrence of hydrogen bonding with the aromatic NH was much less evident for CB7 as judged by the only minor bathochromic shift (?? = +4 nm) of the charge-transfer absorption band of the dye. The dyeCB6 assembly has the potential for the detection of biogenic amines under physiological pH conditions and at low analyte concentrations

Validation of SmartVA using conventional autopsy: A study of adult deaths in Brazil.

BACKGROUND: Accurate cause of death data are essential to guide health policy. However, mortality surveillance is limited in many low-income countries. In such settings, verbal autopsy (VA) is increasingly used to provide population-level cause of death data. VAs are now widely interpreted using the automated algorithms SmartVA and InterVA. Here we use conventional autopsy as the gold standard to validate SmartVA methodology. METHODS: This study included adult deaths from natural causes in São Paulo and Recife for which conventional autopsy was indicated. VA was conducted with a relative of the deceased using an amended version of the SmartVA instrument to suit the local context. Causes of death from VA were produced using the SmartVA-Analyze program. Physician coded verbal autopsy (PCVA), conducted on the same questionnaires, and Global Burden of Disease Study data were used as additional comparators. Cause of death data were grouped into 10 broad causes for the validation due to the real-world utility of VA lying in identifying broad population cause of death patterns. FINDINGS: The study included 2,060 deaths in São Paulo and 1,079 in Recife. The cause specific mortality fractions (CSMFs) estimated using SmartVA were broadly similar to conventional autopsy for: cardiovascular diseases (46.8% vs 54.0%, respectively), cancers (10.6% vs 11.4%), infections (7.0% vs 10.4%) and chronic respiratory disease (4.1% vs 3.7%), causes accounting for 76.1% of the autopsy dataset. The SmartVA CSMF estimates were lower than autopsy for “Other NCDs” (7.8% vs 14.6%) and higher for diabetes (13.0% vs 6.6%). CSMF accuracy of SmartVA compared to autopsy was 84.5%. CSMF accuracy for PCVA was 93.0%. INTERPRETATION: The results suggest that SmartVA can, with reasonable accuracy, predict the broad cause of death groups important to assess a population's epidemiological transition. VA remains a useful tool for understanding causes of death where medical certification is not possible

Characterisation of microbial attack on archaeological bone

As part of an EU funded project to investigate the factors influencing bone preservation in the archaeological record, more than 250 bones from 41 archaeological sites in five countries spanning four climatic regions were studied for diagenetic alteration. Sites were selected to cover a range of environmental conditions and archaeological contexts. Microscopic and physical (mercury intrusion porosimetry) analyses of these bones revealed that the majority (68%) had suffered microbial attack. Furthermore, significant differences were found between animal and human bone in both the state of preservation and the type of microbial attack present. These differences in preservation might result from differences in early taphonomy of the bones. © 2003 Elsevier Science Ltd. All rights reserved