IntroductionGlycoporphyrins offer fascinating prospects for medicinal chemistry and glycobiology. [1a-1d] Carbohydrates not only improve the solubility of porphyrins in an aqueous environment but also offer improved targeting of porphyrin therapeutics.[2] For photodynamic therapy (PDT), [3] carbohydrates can bind to tumour associated lectins displayed on the surface of cancer cells therefore offering the potential for enhanced efficiency and improved selectivity in cancer treatment. Since lectin-carbohydrate interactions are relatively weak it would be advantageous to have access to a PDT photosensitiser with a defined cluster displaying more than one carbohydrate unit. Even more advantageous would be a defined system that displays more than one 'type' of carbohydrate. The synthesis of selectively modified, heterogeneous glycoporphyrins has not previously been reported. To date, the synthesis of glycoporphyrins has relied predominantly on the condensation reaction between a glycosylated aldehyde and a glycosylated aldehyde and a pyrrole.[4] This reaction is often low yielding and is unsuitable for systems where small quantities of a carbohydrate are available. A more general strategy involves the introduction of carbohydrates at specific sites predefined by a regioselective chemical modification. [5] A number of methodologies for the functionalisation of porphyrins with carbohydrates have been investigated including the use of Sonogashira [6] and olefin metathesis [7] cross coupling. Results and DiscussionFor this study we chose to investigate if the Cu(I)-catalysed 1,3-dipolar 'click' reaction [8] could be employed as a robust methodology to access a highly defined, multifunctionalised glycoporphyrin system. Click chemistry has previously been applied to the functionalisation of porphyrins, [9] but to date there is only one literature example describing a click reaction involving a monocarbohydrate modification of a chlorin [10] . 5,10,15,20-Tetraphenylporphyrin was chosen as a common starting material for all modifications. This symmetrical porphyrin can be readily accessed in good yield via a condensation reaction of benzaldehyde and pyrrole. Porphyrin (1), displaying a single azide was prepared according to the literature procedure.[11] The coupling reaction with commercially available β-propargyl glucose (2) (Scheme 1) was investigated under both conventional and microwave mediated heating. The use of MW heating conditions reduced the reaction time from 3 days to 20 mins. A range of reaction conditions were screened. The results of this initial study are outlined in Table 1. Scheme 1. Synthesis of mono-glycoporphyrin by microwave mediated 'click' reaction.It was found that copper chloride in toluene/water (4:1) at 120 o C for 20 mins furnished the desired mono-glycoporphyrin (5) in excellent yield (93%). THF/water (4:1) also gave a very good yield of 81% at a lower temperature but longer times (40 mins) were required for complete consumption of the starting material. It was found that zinc porphyrin was re...

The advantage of stepped vitrification (SV) is avoiding ice crystal nucleation, while decreasing the toxic effects of high cryoprotectant concentrations. We aimed to test this method for human ovarian tissue cryopreservation. Ovarian cortex was taken from 7 fertile adult women. Samples were subjected to an SV protocol performed in an automatic freezer, which allowed sample transfer to ever higher concentrations of dimethyl sulfoxide (DMSO) as the temperature was reduced. Histological evaluation of the vitrified-warmed tissue showed large numbers of degenerated follicles after 24 hours of in vitro culture. We therefore evaluated DMSO perfusion rates by X-ray computed tomography, ice crystal formation by freeze-substitution, and cell toxicity by transmission electron microscopy, seeking possible reasons why follicles degenerated. Although cryoprotectant perfusion was considered normal and no ice crystals were formed in the tissue, ultrastructural analysis detected typical signs of DMSO toxicity, such as mitochondria degeneration, alterations in chromatin condensation, cell vacuolization and extracellular matrix swelling in both stromal and follicular cells. The findings indicated that the method failed to preserve follicles due to the high concentrations of DMSO used. However, adaptations can be made to avoid toxicity to follicles caused by elevated levels of cryoprotectants.

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