Physiological Significance of the Heterogeneous Distribution of Zeaxanthin and Lutein in the Retina of the Human Eye.

Zeaxanthin and lutein are xanthophyll pigments present in the human retina and particularly concentrated in its center referred to as the yellow spot (macula lutea). The fact that zeaxanthin, including its isomer meso-zeaxanthin, is concentrated in the central part of the retina, in contrast to lutein also present in the peripheral regions, raises questions about the possible physiological significance of such a heterogeneous distribution of macular xanthophylls. Here, we attempt to address this problem using resonance Raman spectroscopy and confocal imaging, with different laser lines selected to effectively distinguish the spectral contribution of lutein and zeaxanthin. Additionally, fluorescence lifetime imaging microscopy (FLIM) is used to solve the problem of xanthophyll localization in the axon membranes. The obtained results allow us to conclude that one of the key advantages of a particularly high concentration of zeaxanthin in the central part of the retina is the high efficiency of this pigment in the dynamic filtration of light with excessive intensity, potentially harmful for the photoreceptors.

Macular Pigment Optical Density and Skin Carotenoids in a Childhood Sample.

BACKGROUND: Carotenoids are plant pigments with light filtering and antioxidant properties that deposit in human tissues, including retina and skin. Descriptive characteristics and covariates of carotenoid status in macula and skin have been examined in adults; however, similar studies in children are limited. Thus, this study aimed to delineate how factors of age, sex, race, weight status, and dietary carotenoid intake relate to macular and skin carotenoids in children. METHODS: Children (7-13 y, N = 375) completed heterochromatic flicker photometry to assess macular pigment optical density (MPOD). Participants underwent anthropometrics to measure weight status (BMI percentile [BMI%]), and parent/guardian provided demographic information. Subsample data were available for skin carotenoids (N = 181), assessed using reflection spectroscopy, and dietary carotenoids (N = 101) using the Block Food Frequency Questionnaire. Relationships between skin and macular carotenoids were assessed using partial Pearson's correlations controlling for age, sex, race, and BMI%. Relationships between dietary carotenoids and macular and skin carotenoids were assessed using stepwise linear regression including age, sex, race, and BMI% in the model. RESULTS: Mean MPOD was 0.56 ± 0.22 and skin carotenoid score was 282 ± 94.6. There was no significant correlation between MPOD and skin carotenoids (r = 0.02, P = 0.76). BMI% was negatively associated with skin (stdß = -0.42, P < 0.001), but not macular carotenoids (stdß = -0.04, P = 0.70). Neither MPOD nor skin carotenoids were associated with age, sex, or race (all P > 0.10). MPOD was positively associated with energy-adjusted reported lutein + zeaxanthin intake (stdß = 0.27, P = 0.01). Skin carotenoids were positively associated with energy-adjusted reported carotenoid intake (stdß = 0.26, P = 0.01). CONCLUSIONS: The mean MPOD values in children were higher than what has been reported in adult populations. Previous studies in adult samples report an average MPOD of 0.21. Although macular and skin carotenoids were not related, they were associated with dietary carotenoids relevant to the respective tissues; however, skin carotenoids may be more susceptible negative influence from higher weight status.

Biochemical and Immunological implications of Lutein and Zeaxanthin.

Throughout history, nature has been acknowledged for being a primordial source of various bioactive molecules in which human macular carotenoids are gaining significant attention. Among 750 natural carotenoids, lutein, zeaxanthin and their oxidative metabolites are selectively accumulated in the macular region of living beings. Due to their vast applications in food, feed, pharmaceutical and nutraceuticals industries, the global market of lutein and zeaxanthin is continuously expanding but chemical synthesis, extraction and purification of these compounds from their natural repertoire e.g., plants, is somewhat costly and technically challenging. In this regard microbial as well as microalgal carotenoids are considered as an attractive alternative to aforementioned challenges. Through the techniques of genetic engineering and gene-editing tools like CRISPR/Cas9, the overproduction of lutein and zeaxanthin in microorganisms can be achieved but the commercial scale applications of such procedures needs to be done. Moreover, these carotenoids are highly unstable and susceptible to thermal and oxidative degradation. Therefore, esterification of these xanthophylls and microencapsulation with appropriate wall materials can increase their shelf-life and enhance their application in food industry. With their potent antioxidant activities, these carotenoids are emerging as molecules of vital importance in chronic degenerative, malignancies and antiviral diseases. Therefore, more research needs to be done to further expand the applications of lutein and zeaxanthin.

Macular Pigment Reflectometry: Developing Clinical Protocols, Comparison with Heterochromatic Flicker Photometry and Individual Carotenoid Levels.

The study was designed to: (1) Analyze and create protocols of obtaining measurements using the Macular Pigment Reflectometry (MPR). (2) To assess the agreement of MPOD measurements obtained using the heterochromatic flicker photometry (MPS II) and MPR. (3) To obtain the lutein and zeaxanthin optical density obtained using the MPR in the central one-degree of the macula. The measurements were performed using the MPR and heterochromatic flicker photometry. The MPR measurements were performed twice without pupillary dilation and twice following pupillary dilation. The MPR measurements were performed for a 40-s period and the spectrometer signal was parsed at different time points: 10-20, 10-30, 10-40, 20-30, 20-40, and 30-40 s. The MPR analyzes the high-resolution spectrometer signal and calculates MPOD, lutein optical density and zeaxanthin optical density automatically. The MPR-MPOD data was compared with MPPS II-MPOD results. The MPR-MPOD values are highly correlated and in good agreement with the MPS II-MPOD. Of the various parsing of the data, the data 10-30 interval was the best at obtaining the MPOD, lutein, and zeaxanthin values (8-12% coefficient of repeatability). The lutein to zeaxanthin ratio in the central one-degree of the macula was 1:2.40. Dilation was not needed to obtain the MPOD values but provided better repeatability of lutein and zeaxanthin optical density. MPR generates MPOD measurements that is in good agreement with MPS II. The device can produce lutein and zeaxanthin optical density which is not available from other clinical devices.

Effect of an antioxidant supplement containing high dose lutein and zeaxanthin on macular pigment and skin carotenoid levels.

The effect of a high dose lutein/zeaxanthin supplement on macular pigment optical density (MPOD) and skin carotenoid (SC) levels in healthy subjects was investigated. This is a prospective, single-arm, open-label study. Subjects were 16 Japanese, age 26-57 years. Subjects took a supplement containing 20 mg/day of lutein, 4 mg/day of zeaxanthin, and other antioxidants (vitamin C, vitamin E, zinc, copper) for 16 weeks. MPOD levels were measured by a two-wavelength autofluorescence imaging technique. SC levels were measured by reflection spectroscopy. Total volume of MPOD within 9° eccentricity significantly increased by week 8 and continued to increase until week 16 (p < 0.0001, two-way factorial ANOVA). The increase rate of MPOD was significantly higher in subjects with body mass index (BMI) less than 25 kg/m2 (n = 13) compared to those of 25 kg/m2 and higher (n = 3). SC levels increased significantly by week 4 and continued to increase until week 16 (p < 0.0001, two-way factorial ANOVA). All subjects completed the study without any serious adverse events. These results demonstrated the effectiveness of a high dose lutein/zeaxanthin supplement for MPOD volume and SC levels without serious adverse events.

Lutein Supplementation for Eye Diseases.

Lutein is one of the few xanthophyll carotenoids that is found in high concentration in the macula of human retina. As de novo synthesis of lutein within the human body is impossible, lutein can only be obtained from diet. It is a natural substance abundant in egg yolk and dark green leafy vegetables. Many basic and clinical studies have reported lutein's anti-oxidative and anti-inflammatory properties in the eye, suggesting its beneficial effects on protection and alleviation of ocular diseases such as age-related macular degeneration, diabetic retinopathy, retinopathy of prematurity, myopia, and cataract. Most importantly, lutein is categorized as Generally Regarded as Safe (GRAS), posing minimal side-effects upon long term consumption. In this review, we will discuss the chemical structure and properties of lutein as well as its application and safety as a nutritional supplement. Finally, the effects of lutein consumption on the aforementioned eye diseases will be reviewed.

Serum Lutein is related to Relational Memory Performance.

Dietary carotenoids, plant pigments with anti-oxidant properties, accumulate in neural tissue and are often found in lower concentrations among individuals with obesity. Given previous evidence of negative associations between excess adiposity and memory, it is possible that greater carotenoid status may confer neuroprotective effects among persons with overweight or obesity. This study aimed to elucidate relationships between carotenoids assessed in diet, serum, and the macula (macular pigment optical density (MPOD)) and relational memory among adults who are overweight or obese. Adults aged 25-45 years (N = 94) completed a spatial reconstruction task. Task performance was evaluated for accuracy of item placement during reconstruction relative to the location of the item during the study phase. Dietary carotenoids were assessed using 7-day diet records. Serum carotenoids were measured using high-performance liquid chromatography. Hierarchical linear regression analyses were used to determine the relationship between carotenoids and task performance. Although initial correlations indicated that dietary lutein, beta-carotene, and serum beta-carotene were positively associated with memory performance, these relationships were not sustained following adjustment for age, sex, and BMI. Serum lutein remained positively associated with accuracy in object binding and inversely related to misplacement error after controlling for covariates. Macular carotenoids were not related to memory performance. Findings from this study indicate that among the carotenoids evaluated, lutein may play an important role in hippocampal function among adults who are overweight or obese.

Mechanisms enhancing the protective functions of macular xanthophylls in the retina during oxidative stress.

Macular xanthophylls (MXs) are distinguished from other dietary carotenoids by their high membrane solubility and preferential transmembrane orientation. Additionally, these properties enhance the chemical and physical stability of MXs in the eye retina, and maximize their protective activities. The effectiveness of MXs' protection is also enhanced by their selective accumulation in the most vulnerable domains of retinal membranes. The retina is protected by MXs mainly through blue-light filtration, quenching of the excited triplet states of potent photosensitizers, and physical quenching of singlet oxygen. To perform these physical, photo-related actions, the structure of MXs should remain intact. However, the conjugated double-bond structure of MXs makes them highly chemically reactive and susceptible to oxidation. Chemical quenching of singlet oxygen and scavenging of free radicals destroy their intact structure and consume MXs. Consequently, their physical actions, which are critical to the protection of retina, are diminished. Thus, it is timely and important to identify mechanisms whereby the chemical destruction (bleaching) of MXs in retinal membranes can be reduced. It was shown that nitroxide free radicals (spin labels) located in membranes protect MXs against destruction, and their effect is especially pronounced during the light-induced formation of singlet oxygen. That should extend and enhance their positive action in the retina through physical processes. In this review, we will discuss possible applications of this new strategy during ophthalmological procedures, which can cause acute bleaching of MXs and damage the retina through oxidative processes.

Macular Xanthophylls Are Related to Intellectual Ability among Adults with Overweight and Obesity.

Excess adiposity or obesity has been inversely related to cognitive function and macular xanthophyll status. However, whether the neuroprotective effects of macular xanthophylls on cognitive function are independent of excess adiposity is unclear. We investigated the relationship between macular xanthophylls and intellectual ability among adults (N = 114) between 25 and 45 years with overweight and obesity (≥25 kg/m²). Dual energy X-ray absorptiometry and heterochromatic flicker photometry were used to assess whole body adiposity (%Fat) and macular pigment optical density (MPOD), respectively. Dietary xanthophylls (lutein and zeaxanthin) were assessed using 7-day diet records. The Kaufman Brief Intelligence Test-2 (KBIT-2) was used to assess general intelligence (IQ) as well as fluid and crystallized intelligence. Bivariate correlations revealed that MPOD was inversely related to %Fat and positively associated with IQ and fluid intelligence. Although %Fat was inversely correlated to IQ and fluid intelligence, this relationship did not persist following adjustment for sex and MPOD. Further, MPOD was an independent predictor of IQ and fluid intelligence. However, no significant relationships were observed between MPOD and crystalized intelligence. These results suggest that macular xanthophylls are selectively related to fluid intelligence, regardless of degree of adiposity among adults with overweight and obesity.

Two heterozygous mutations identified in one Chinese patient with bilateral macular coloboma.

Congenital macular coloboma is characterized by defined punched out atrophic lesions of the macula. The present study aimed to investigate the genetic alterations of one Chinese sporadic patient with bilateral large macular coloboma. Complete ophthalmic examinations, including best­corrected visual acuity, slit­lamp examination, fundus examination, fundus photograph and fundus fluorescein angiography imaging, Pentacam, and optical coherence tomography were performed on the patient. Genomic DNA was extracted from leukocytes in a peripheral blood sample collected from the patient, the patient's unaffected family members and from 200 unrelated control subjects from the same population. Next­generation sequencing of the known genes involved in ocular disease was performed. The functional effects of the mutation were analyzed using Polymorphism Phenotyping (PolyPhen) and Sorting Intolerant From Tolerant (SIFT). One heterozygous bestrophin 1 (BEST1) mutation c.1037C>A (p.Pro346His, p.P346H) in exon 9 and one heterozygous regulating synaptic membrane exocytosis 1 (RIMS1) mutation c.3481A>G (p.Arg1161Gly, p.R1161G) in exon 23 were identified in the patient being investigated, but not in the unaffected family members or unrelated control subjects. Polyphen and SIFT predicted that the amino acid substitution p.P346H in the BEST1 protein is damaging. In addition, Polyphen predicted that the amino acid substitution p.R1161G in the RIM1 protein is damaging. The results of the current study have increased the mutation spectrums of BEST1 and RIMS1, and are valuable for improving the current genetic counseling process and developing novel therapeutic interventions for patients with macular coloboma.

Evidence of macular pigment in the central macula in albinism.

PURPOSE: Albinism represents a spectrum of disorders with diminished to absent amounts of melanin pigmentation including the posterior segment of the eye. Macular pigment (MP) consists of two main carotenoids, lutein and zeaxanthin, concentrated in the macula. MP serves as blue light absorbent, antioxidant, and may reduce chromatic aberration and glare. It remains unclear if albinos have detectable MP. The purpose was to investigate the distribution of MP in albino patients with psychophysical and imaging techniques. METHODS: MP was measured at the eccentricity of 0.5° by heterochromatic flicker perimetry (QuantifEye(®); Tinsley Precision Instruments Ltd.) or by scanning laser ophthalmoscopy (MPOD module, MultiColor Spectralis(®), Heidelberg Engineering, Heidelberg, Germany) in four albino patients, who were also investigated with multimodal ophthalmic imaging. RESULTS: Visual acuity ranged from 20/32 to 20/125, nystagmus was present in three patients, and all patients showed typical foveal hypoplasia on fundus exam and optical coherence tomography. Fundus autofluorescence (FAF) demonstrated various degrees of central FAF signal attenuation. Genetic testing was available in three patients and confirmed the diagnosis. Measurable amounts of MP were detected in all four patients and ranged from 0.05 to 0.24, which is below the normal range. CONCLUSIONS: We conclude that MP can be demonstrated and measured in albinos. Further studies are needed to investigate MP accumulation following carotenoid supplementation and its impact on visual performance.

Association of macular pigment optical density with serum concentration of oxidized low-density lipoprotein in healthy adults.

PURPOSE: To analyze the association between macular pigment optical density (MPOD), which reflects lutein (L), zeaxanthin (Z), and meso-zeaxanthin (MZ) in the macula, and background characteristics. METHODS: Fifty-five healthy adult volunteers were analyzed. Macular pigment optical density was measured using a heterochromatic flicker photometry technique, and serum concentrations of carotenoids and lipoproteins were by high-performance liquid chromatography and enzyme-linked immunosorbent assay, respectively. Dietary intake of nutrient was determined by a validated self-administered questionnaire on ingestion frequency. RESULTS: Macular pigment optical density was positively correlated with serum concentrations of L and Z and dietary L intake and inversely correlated with serum oxidized low-density lipoprotein (LDL). Although MPOD decreased with age (95% confidence interval, -0.011 to -0.002; correlation coefficient, -0.269; P = 0.007), serum L/Z and dietary L intake did not. In contrast, serum oxidized LDL was positively correlated with age (95% confidence interval, 0.69-2.34; correlation coefficient, 0.333; P = 0.0004). After adjusting for age, sex, and oxidized LDL, serum L was positively correlated with MPOD (95% confidence interval, 0.88-1.69; P = 0.000001). After adjusting for age, sex, and serum L, serum oxidized LDL was inversely correlated with MPOD (95% confidence interval, -0.002 to -0.0004; P = 0.006). CONCLUSION: Macular pigment optical density was inversely correlated with serum oxidized LDL. Further study to know the impact of oxidized LDL on MPOD may be warranted.

Solubilization and stabilization of macular carotenoids by water soluble oligosaccharides and polysaccharides.

Xanthophyll carotenoids zeaxanthin and lutein play a special role in the prevention and treatment of visual diseases. These carotenoids are not produced by the human body and must be consumed in the diet. On the other hand, extremely low water solubility of these carotenoids and their instability restrict their practical application as components of food or medicinal formulations. Preparation of supramolecular complexes of zeaxanthin and lutein with glycyrrhizic acid, its disodium salt and the natural polysaccharide arabinogalactan allows one to minimize the aforementioned disadvantages when carotenoids are used in food processing as well as for production of therapeutic formulations with enhanced solubility and stability. In the present study, the formation of supramolecular complexes was investigated by NMR relaxation, surface plasmon resonance (SPR) and optical absorption techniques. The complexes increase carotenoid solubility more than 1000-fold. The kinetics of carotenoid decay in reactions with ozone molecules, hydroperoxyl radicals and metal ions were measured in water and organic solutions, and significant increases in oxidation stability of lutein and zeaxanthin in arabinogalactan and glycyrrhizin complexes were detected.

A multiscale optimization approach to detect exudates in the macula.

Pathologies that occur on or near the fovea, such as clinically significant macular edema (CSME), represent high risk for vision loss. The presence of exudates, lipid residues of serous leakage from damaged capillaries, has been associated with CSME, in particular if they are located one optic disc-diameter away from the fovea. In this paper, we present an automatic system to detect exudates in the macula. Our approach uses optimal thresholding of instantaneous amplitude (IA) components that are extracted from multiple frequency scales to generate candidate exudate regions. For each candidate region, we extract color, shape, and texture features that are used for classification. Classification is performed using partial least squares (PLS). We tested the performance of the system on two different databases of 652 and 400 images. The system achieved an area under the receiver operator characteristic curve (AUC) of 0.96 for the combination of both databases and an AUC of 0.97 for each of them when they were evaluated independently.

Comparison of two methods to measure macular pigment optical density in healthy subjects.

PURPOSE: To evaluate and compare macular pigment optical density (MPOD) measurements obtained using the modified Heidelberg Retina Angiograph (HRA) and the Visucam 200. METHODS: Healthy young subjects were included in this prospective study. MPOD was measured with the modified HRA at 0° and 0.5°, 1°, 2°, and 6° eccentricities from the fovea. The parameters obtained with the Visucam 200 (maximum, mean, area, and volume) were recorded the same day on the same subjects. Intraclass correlation coefficients (ICCs) and correlation coefficients were used to evaluate the agreement between the two devices. The repeatability and the reproducibility of each method were also assessed. RESULTS: Sixty-seven subjects were included whose median (interquartile ratio) age was 25 years (range, 23-30 years). The MPODs as measured with the modified HRA were higher than those measured with the Visucam 200 (P < 0.0001). The ICCs were low, ranging from 0.020 to 0.188. The correlation coefficients between the two methods were very low and ranged from 0.05 to 0.22. Repeatability and reproducibility were good with both methods, with ICCs ranging from 0.697 to 0.923. CONCLUSIONS: Agreement between the modified HRA and the Visucam in measuring MPOD was rather low. These results suggest that the two methods are not interchangeable. Before using the Visucam 200 in clinical and research setting, further evaluation seems mandatory (http://ansm.sante.fr/ number, 2009-A00448-49).

Macular pigment optical density is related to cognitive function in older people.

BACKGROUND: the xanthophylls lutein (L) and zeaxanthin (Z) exist in relatively high concentration in multiple central nervous tissues (e.g. cortex and neural retina). L + Z in macula (i.e. macular pigment, MP) are thought to serve multiple functions, including protection and improvement of visual performance. Also, L + Z in the macula are related to L + Z in the cortex. OBJECTIVE: to determine whether macular pigment optical density (MPOD, L + Z in the macula) is related to cognitive function in older adults. METHODS: participants were older adults (n = 108, 77.6 ± 2.7 years) sampled from the age-related maculopathy ancillary study of the Health Aging and Body Composition Study (Memphis, TN, USA). Serum carotenoids were measured using high performance liquid chromatography. MPOD was assessed using heterochromatic flicker photometry. Eight cognitive tests designed to evaluate several cognitive domains including memory and processing speed were administered. Partial correlation coefficients were computed to determine whether cognitive measures were related to serum L + Z and MPOD. RESULTS: MPOD levels were significantly associated with better global cognition, verbal learning and fluency, recall, processing speed and perceptual speed, whereas serum L + Z was significantly related to only verbal fluency. CONCLUSION: MPOD is related to cognitive function in older people. Its role as a potential biomarker of cognitive function deserves further study.

Innovative Troxler-free measurement of macular pigment and lens density with correction of the former for the aging lens.

Simplified measurement of macular pigment optical density (MPOD) is important because of the ocular health benefits that are attributed to these retinal carotenoids. Here, we describe a novel instrument designed for this purpose, based on heterochromatic flicker photometry (HFP), which removes a number of difficulties that subjects often experience with traditional HFP. The instrument generates 1.5- and 15-deg diameter, centrally viewed stimuli that alternate between blue and green colors generated by light emitting diodes (LED). The 15 deg stimulus replaces the small, eccentrically viewed stimulus used in traditional HFP. Subjects adjust the blue LED intensity until flicker is eliminated in the case of the 1.5 deg stimulus and eliminated around the periphery in the case of the 15 deg stimulus. A microprocessor computes the subject's MPOD, in addition to the lens OD, and uses the latter to correct the MPOD. Good repeatability was confirmed through test-retest measurements on 52 subjects. The overwhelming majority of them stated that they found the test easy. The importance of the lens correction on MPOD measurements was confirmed in a simulation study. The study showed that, without the correction, MPOD would show an apparent age-related decline in a population for whom there was no real age dependence.

Concordance of macular pigment measurements obtained using customized heterochromatic flicker photometry, dual-wavelength autofluorescence, and single-wavelength reflectance.

This study compares in vivo measurements of macular pigment (MP) obtained using customized heterochromatic flicker photometry (cHFP; Macular Metrics Densitometer(™)), dual-wavelength fundus autofluorescence (Heidelberg Spectralis(®) HRA + OCT MultiColor) and single-wavelength fundus reflectance (Zeiss Visucam(®) 200). MP was measured in one eye of 62 subjects on each device. Data from 49 subjects (79%) was suitable for analysis. Agreement between the Densitometer and Spectralis was investigated at various eccentricities using a variety of quantitative and graphical methods, including: Pearson correlation coefficient to measure degree of scatter (precision), accuracy coefficient, concordance correlation coefficient (ccc), paired t-test, scatter and Bland-Altman plots. The relationship between max MP from the Visucam and central MP from the Spectralis and Densitometer was investigated using regression methods. Agreement was strong between the Densitometer and Spectralis at all central eccentricities (e.g. at 0.25° eccentricity: accuracy = 0.97, precision = 0.90, ccc = 0.87). Regression analysis showed a very weak relationship between the Visucam and Densitometer (e.g. Visucam max on Densitometer central MP: R(2) = 0.008, p = 0.843). Regression analysis also demonstrated a weak relationship between MP measured by the Spectralis and Visucam (e.g. Visucam max on Spectralis central MP: R(2) = 0.047, p = 0.348). MP values obtained using the Heidelberg Spectralis are comparable to MP values obtained using the Densitometer. In contrast, MP values obtained using the Zeiss Visucam are not comparable with either the Densitometer or the Spectralis MP measuring devices. Taking cHFP as the current standard to which other MP measuring devices should be compared, the Spectralis is suitable for use in a clinical and research setting, whereas the Visucam is not.