The carotenoids lutein (L), zeaxanthin (Z), and meso-zeaxanthin (MZ) accumulate in the central retina, where they are collectively known as macular pigment (MP). Each of these three compounds exhibit a regional dominance, with MZ, Z, and L being the dominant carotenoids at the epicentre, mid-periphery, and periphery of the macula, respectively. There is a growing and evidencebased consensus that MP is important for optimal visual performance, because of its blue light-filtering properties and consequential attenuation of chromatic aberration, veiling luminance, and blue haze. It has also been hypothesised that MP may protect against age-related macular degeneration because of the same optical properties and also because of the antioxidant capacity of the three macular carotenoids. Challenges inherent in the separation and quantification of MZ have resulted in a paucity of data on the content of this carotenoid in foodstuffs, and have rendered the study of tissue concentrations of this compound problematic. As a consequence, the few studies that have investigated MZ have, perhaps, been disproportionately influential in the ongoing debate about the origins of this macular carotenoid. Certainly, the narrative that retinal MZ is derived wholly and solely from retinal L needs to be revisited.

Molecularly Imprinted Polymers (MIPs) against imiquimod, a highly potent immune response modifier used in the treatment of skin cancer, were synthesised using a template analogue strategy and were compared with imprints of the drug itself. An investigation of the complexation between the functional monomer and the template analogue revealed an association constant of 1376 AE 122 M À1 , significantly higher than previously reported values for similar systems. The binding characteristics of the synthesised imprinted polymers were evaluated and extremely strong binding for imiquimod was observed while imprinting factors as high as 17 were calculated. When applied as sorbents in solid-phase extraction of imiquimod from aqueous, urine and blood serum samples, clean extracts and recoveries up to 95% were achieved, and it is concluded that while imiquimod imprints exhibited higher capacity for the drug, template analogue imprints are more selective. The results obtained suggest potential applications of imiquimod imprints as sorbents in rapid extraction and monitoring of undesirable systemic release of the drug.

Sir, Response to Bernstein et alWe welcome the letter by Bernstein et al 1 in response to our publication 'What is meso-zeaxanthin, and where does it come from?' in Eye 2013. 2 In their letter, Bernstein and colleagues argue that our review article contains 'several critical errors that need to be considered. ' Bernstein and colleagues endeavour to make their points under the following headings:1. Quantitation of xanthophylls using reverse-and normal-phase HPLC. 2. The role of saponification in the quantitation of xanthophylls in food and supplements. 3. Meso-zeaxanthin in lutein supplements.4. Additional evidence supporting lutein as the precursor of meso-zeaxanthin.In our letter below, we reply directly to these points in normal font. Statements made by Bernstein and colleagues are presented in bold font for clarity.1. Quantitation of xanthophylls using reverse-and normal-phase HPLC. 'Nolan et al argue that the two-step HPLC method used for MZ quantitation by Johnson et al is limited because of the labor involved in the manual collection of the total Z þ MZ fraction in the first step. The authors suggest that this process is prone to human error, that only a portion of the Z þ MZ fraction would be collected, and that this fraction typically is contaminated with L carryover. ' We thank Bernstein et al for summarising the two-step method in their correspondence, commonly used for quantifying MZ. We are very familiar with this method, as we have used it in several of our recently published studies. [3][4][5][6] In our review article, we point out the limitations of the standard 'two-step method' commonly used by many laboratories to quantify MZ. These limitations include the following: its labour intensive nature due to manual collection; operator dependency and potential for human error; and a very long sample run time, rendering it difficult to perform bulk analysis (eg, for clinical trials). Our concerns with respect to the traditional 'two-step method' remain, and we believe that it is important to recognise these limitations when discussing published methodology and findings from papers, and that is why we included these points in our review.Bernstein et al premise their defence of the methodology of carotenoid quantification in the paper by Johnson et al 7 on the basis that:'The fact that L, MZ and Z appear on the subsequent normal-phase, chiral column chromatogram verifies that the desired peaks were collected, and this was also confirmed by absorption spectra.'Bernstein et al attempt to address our concerns with respect to the unknown peak that was found to co-elute with the Z fractions of retinal samples in the report by Johnson et al 7 by stating that 'ythe peak also appeared in the reverse phase HPLC of retinal samples from the carotenoid-free monkeys. ' We agree that identifying the peaks and confirming their presence by assessing their absorbance spectra are important. However, it is clear from the Johnson et al 7 paper that the already challenging method used to analyse MZ was made more diffic...