Organic & Biomolecular Chemistry Blog

Over the past several decades, advances in cell imaging have dramatically transformed biology and medicine. Fluorescence spectroscopy and microscopy are currently the most popular imaging techniques however, there are intrinsic limitations; many substrates are non-fluorescent or weakly fluorescent, fluorescent labels are often perturbative for small molecules and peptides and, perhaps most importantly, labelling or staining with fluorophores are not recommended for in vivo medicinal applications in humans. Hence, the search for highly sensitive optical imaging methods is increasingly desirable in biomedical and material sciences.

Nonlinear optical imaging is an emerging technology that encompasses a range of optical phenomena. In a recent study by Prof. Koen Clays of the University of Leuven and Prof. Harry Anderson of Oxford University, a new group of chromophores based on pyropheophorbide-a methyl ester (PPa-OMe) was developed for the linear and nonlinear optical imaging of membrane potentials as well as biological imaging of structures through two-photon excited fluorescence (TPEF) and second harmonic generation (SGH) microscopy.

In TPEF, a fluorophore is excited by the simultaneous absorption of two photons in the infrared spectral range. In conventional one-photon fluorescence, the same transition to higher energy levels requires photons in the ultraviolet or visible range. The longer incident wavelength in TPEF leads to improved depth penetration in tissues, with reduced potential for photolytic damage. SHG, on the other hand, is a nonlinear process where two photons interact with a nonlinear material and are effectively combined to generate new photons with twice the energy. This process does not involve absorption of photons but relies on virtual energy states and can only occur in materials that exhibit a non-centrosymmetric structure.

Unlike incoherent processes such as fluorescence, coherent nonlinear optical spectroscopies generate different optical signals depending on the underlying processes. They have broad utility as biomedical tools, offering contrasting mechanisms to fluorescence emissions and provide a useful alternative to label-based imaging.

In this OBC publication, the electronic structure of PPa-OMe (1a) was altered to tune it’s linear and nonlinear optical properties. Porphyrins and related porphyrinoid chromophores inherently possess excellent linear and nonlinear optical properties due to their large, conjugated π-system. By incorporating both electron-donating and –accepting groups, a push-pull type system was generated in which greater SHG intensity was observed due to the increased polarization of its π-system. A hydrophilic group, bis-triethyleneglycol (TEG) amide, was attached to make PPa-OMe amphiphilic and was then suspended in lipid-based water in oil monolayer droplets—a simple model system used to probe potentials across cellular membranes. TPEF and SHG images of the bis-TEG amide attached dyes revealed that the TPEF and SHG involve transition dipole moments in different orientations. While TPEF is detectable in all directions around the sample, SHG is detected in the forward direction of the incident light meaning there is an overall cancelling of the SHG signal from anti-parallel dyes. In order to improve these systems, control over orientation within cell membranes is crucial however, chromophores based on these PPa-OMe derivatives are promising prototypes for future cell imaging studies.

To find out more see:

Push-pull pyropheophorbides for nonlinear optical imaging
Anjul Khadria, Yovan de Coene, Przemyslaw Gawel, Cécile Roche, Koen Clays and Harry L. Anderson
DOI: 10.1039/C6OB02319C

Victoria Corless is currently completing her Ph.D. in organic chemistry with Prof. Andrei Yudin at The University of Toronto. Her research is centred on the synthesis of kinetically amphoteric molecules, which offer a versatile platform for the development of chemoselective transformations with particular emphasis on creating novel biologically active molecules.

For years, radiotherapy has been an essential mode of noninvasive cancer therapy and advancements have led to life saving treatments for patients. In contrast to other conventional radiotherapies, boron neutron capture therapy (BNCT) is unique in its selective destruction of cancerous cells. BNCT is based on nuclear capture and fission reactions when nonradioactive ¹⁰B is irradiated with neutrons to yield excited ¹¹B* which decays into high energy alpha particles and ⁷Li nuclei. Boron is preferentially accumulated into tumour cells though non-toxic carriers and the short length of the generated neutron beams (5-9 µm) destroys nearby cells leaving the surrounding healthy tissue intact.

The development of carrier systems that deliver sufficient amounts of boron to carry out effective destruction of all vicinal tumour cells has been a significant area of BNCT research for many years. A recent breakthrough made by Professors Atsushi Ikeda of Hiroshima University, Takeshi Nagasaki of Osaka City University and Kazuya Koumoto of Konan University has led to the development of a novel method for incorporating boron-containing moieties within hydrophobic lipid membranes.

Liposomes have been studied extensively as boron carriers and are promising candidates for BNCT as lipids display low toxicity. In the past, boron-containing liposomes have been prepared by dissolution of boron compounds in an internal water phase (Figure, A) or by using amphiphilic boron compounds embedded in the liposomal bilayer such that boron coats the outer surface of the lipid membrane (Figure, B). Unfortunately, even when these two methods are used in conjunction, the concentration of boron within the liposome is insufficient for effective BNCT and increasing liposomal dosing is problematic as high concentrations of lipids interfere with uptake mechanisms in the liver.

A novel method for boron transport is therefore of high value and in their recent OBC publication, the first example of boron incoporation into liposomal lipid membranes was successfully demonstrated. Key was the preparation of aryl pinacolato boronate esters bearing methyl groups at both ortho positions (Figure, R¹ positions) of the phenyl rings which helped in stabilizing the boronate esters against hydrolysis.

While the concentration of boron within the lipid membrane was not particularly high, this study has unlocked yet another avenue through which non-toxic liposomal boron carrier concentrations can be improved and when used with previous boron loading methods, produces liposomes with sufficient boron concentrations to carry out BCNT. This result will no doubt enhance BNCT and lead to critical, life saving therapies for cancer patients.

To find out more see:

Lipid-membrane-incorporated arylboronate esters as agents for boron neutron capture therapy
Masafumi Ueda, Kengo Ashizawa, Kouta Sugikawa, Kazuya Koumoto, Takeshi Nagasaki and Atsushi Ikeda
DOI:10.1039/C6OB02142E

Victoria Corless is currently completing her Ph.D. in organic chemistry with Prof. Andrei Yudin at The University of Toronto. Her research is centred on the synthesis of kinetically amphoteric molecules, which offer a versatile platform for the development of chemoselective transformations with particular emphasis on creating novel biologically active molecules.