In the early 1990s, we reported that MZ, along with lutein and zeaxanthin comprise the macular pigment; these carotenoids constitute the macula lutea, or yellow spot in the human retina.^3-5 Located in the Henle fibers (receptor axons)6 and outer segments,^7,8 macular pigment has a protective effect against AMD. Oxidative stress is considered to be an important, but modifiable, factor in the etiology of AMD.⁹ Oxidized lipids, particularly, polyunsaturated fatty acids, are incompletely eliminated through the choroidal circulation and accumulate under the photoreceptors in the form of drusen that characterize early-stage maculopathy. Blue light, acting on photosensitizers, such as A2E, is able to promote the formation of reactive oxygen species (ROS) in the retina.¹⁰ The macular carotenoids, in addition to their antioxidant properties, act as a blue-light filter. Macular pigment is known to exert its protective influence by the combination of these two attributes.^11,12 Fortunately, the macular carotenoids deactivate singlet oxygen, one of the principal chemical villains associated with photo-oxidative events, mainly by energy transfer, with the result that the carotenoids are not “used up.”^12,13 These carotenoids can also directly reduce the destructive capability of other ROS, such as peroxy radicals.¹⁴

In the central 3 mm of the macula, MZ, lutein and zeaxanthin are present in approximately equal amounts. Elsewhere in the retina, these proportions change. Relative to zeaxanthin, the proportion of MZ decreases while that of lutein increases at points more removed from the fovea. This surprising observation led us to hypothesize that an enzyme-mediated isomerization of lutein into MZ occurs within the central-most regions of the retina.¹⁵ Experimental evidence from macaques supplemented with either lutein or zeaxanthin has confirmed our hypothesis and demonstrates that MZ present in the retina is not derived from the blood.¹⁶ Whether MZ is present in the blood of subjects who have not been supplemented with MZ has yet to be demonstrated unambiguously. Results are equivocal, with one study reporting the absence of MZ in the blood3 and another reporting the presence of a small, incompletely identified candidate that may be MZ.¹⁷ Suffice it to say, almost all MZ in the macula is derived from a conversion process of lutein or zeaxanthin to MZ. As we will demonstrate below, this conversion can be bypassed by supplementing the diet directly with MZ (safely) thereby avoiding a potentially deficient unique individual conversion processes or frankly deficient diets.


Stereoisomers  

How Bone and Landrum Discovered Meso-zeaxanthin  Meso-zeaxanthin, a subtle structural variation on commonly encountered dietary zeaxanthin — now considered a critical carotenoid for protecting the macula — was discovered in the retina by chance. In the late 1980s, we thought it would be a worthwhile endeavor to establish conclusively which steroisomers of the xanthophylls compose the macular pigment. Analytic methodologies developed in Japan, Norway and Switzerland had been used to separate these nearly identical species and to detect their presence in tissues in many marine organisms. Using these same methodologies we set out to analyze the xanthophylls in the human macula, fully expecting to see only lutein and zeaxanthin, the carotenoids found abundantly in the diet. To our surprise, we found that meso-zeanthin (uncommon in the diet) comprises 33 percent of the total carotenoid content of the macula. Our discovery demonstrates that serendipity, as much as careful planning, still accounts for many illuminating scientific discoveries. —J.L., R.B.

The unifying motif of all carotenoids is a linear, conjugated chain of double bonds. This feature is responsible for the brilliant reds, oranges and yellows that typify these pigments in nature. Higher animals cannot synthesize the carbon framework of carotenoids but they can metabolize them in a variety of ways.^18,19 Lutein is one of the most abundant carotenoids and can exist in eight stereo-molecular forms. Higher plants predominantly produce one of these forms, referred to here as lutein. This is the form of lutein found in the human retina.
Zeaxanthin and its stereoisomers have identical chemical formulas to lutein but their absorption spectrum is shifted noticeably to the red. An added benefit of this is that the light filtering ability of the macular pigment spans a greater portion of the spectrum than if only a single carotenoid were present. Zeaxanthin exists as three stereoisomers called (R,R), (S,S) and (R,S-meso) zeaxanthin. All three occur in nature.^15,19 Higher plants produce solely the (R,R) form, referred to here as zeaxanthin. Although this is the most commonly encountered zeaxanthin isomer in the diet, the human macular pigment contains all three.^3,15


MZ Is Produced in Nature 

 
Birds make MZ and concentrate it and other carotenoids in their retinas within brightly colored oil-droplets. In chicken retinas, 47 percent of the total zeaxanthin is MZ and in turkeys it is 28 percent.^20,21 As with humans, MZ is formed in the retina from lutein.
In landmark research, the deposition of carotenoids in fish was thoroughly investigated.¹⁸ It is well known) that in both rainbow trout and salmon the characteristic salmon color of the flesh is due to deposition of astaxanthin. Surprisingly, in addition to astaxanthin, trout contain up to 0.2 µg/g total of lutein and zeaxanthin in their flesh and levels of 5 to 10 µg/g in fins and skin.²² Analysis of skin from trout fed a diet rich in astaxanthin revealed significant quantities of both MZ and SS-Z formed from the astaxanthin. Like the rainbow trout, the Atlantic salmon also deposits MZ and SS-Z within its skin.²¹ These isomers of zeaxanthin are present in many aquatic species. This research showed that MZ, zeaxanthin and SS-Z are present in 21 species of edible fish, shrimp and sea turtles.


Transforming Lutein into MZ 

 
Major suppliers of pigments for poultry feed produce MZ-containing carotenoid mixtures. MZ in these products makes them more efficient at pigmenting chickens and egg yolk than lutein.^19,23 Such feeds have been in use in North America for many years and available in the United States through at least 2008.²⁴ The chemical transformation of lutein, derived from marigold blooms, into MZ is carried out commercially by a catalytic method involving the treatment of lutein with base at elevated temperature. This simple process produces an equilibrium mixture of MZ and lutein with as much as 80 percent MZ. This equilibrium accounts for the absence of products containing exclusively MZ.


MZ in the Diet 

 
In a healthy diet, we consume both lutein and zeaxanthin in relatively large quantities compared to other carotenoids. Lutein and zeaxanthin enter the Western diet chiefly as components of green and yellow vegetables and fruits, although they are also present in milk and meat (chiefly in fat), albeit at lower levels. An especially rich dietary source of lutein and zeaxanthin is egg yolks.²⁵ Humans ingest relatively low levels of MZ, setting the stage for a potential deficiency, especially among individuals who are inefficient in synthesizing MZ, or lack the ability. There has been no exhaustive assessment of the amounts of MZ in a normal diet. Brightly colored egg yolks from hens fed MZ are the richest potential human dietary source.²⁶ Fish skin is a dietary source of MZ (and also the highly desirable polyunsaturated fatty acids) as well as potentially the fats and oils that extract MZ during cooking.


MZ Is Safe 

 
The safety of MZ has been evaluated in a recent toxicity trial using a small animal model. The results of this trial verified that the “No-Observed-Adverse-Effect-Level” (NOAEL) was in excess of 200 mg/kg/day,²⁷ far greater than doses used in dietary supplements, which are typically less than 0.5 mg/kg/day. Absence of mutagenicity was confirmed by the Ames test.²⁸ MZ is also a regular dietary component in countries where it is a major pigment used by the poultry industry, particularly Mexico, and no adverse effects have been reported. A recent study of supplemental MZ in human subjects has confirmed, by serum analysis, that renal and liver function, as well as lipid profile, hematological parameters, and indicators of inflammation, are unaffected by such supplementation.²⁹


A Synergistic Antioxidant Effect 

Noteworthy experiments conducted with lutein and zeaxanthin in vitro indicate that zeaxanthin is a more potent antioxidant than lutein.³⁰ According to one study, zeaxanthin was approximately twice as effective at deactivating singlet oxygen when compared with lutein.31 This is due to the extended conjugation of the zeaxanthin molecule in comparison with that of lutein. Like zeaxanthin, MZ shares this electronic feature. However, it has also been reported that MZ, in association with a zeaxanthin-binding protein—the pi isoform of glutathione S-transferase—provides slightly better protection against lipid membrane oxidation than zeaxanthin.³² Of particular relevance to this review are the results of another recent study in which the singlet oxygen deactivating abilities of all three macular carotenoids, individually and in combination, were evaluated.² A mixture of equal amounts of MZ, lutein and zeaxanthin, as found in the center of the retina, was found to have the highest singlet oxygen deactivating ability (a measure of its antioxidant potential): 37-percent more effective than MZ alone, 91-percent more effective than zeaxanthin alone, and 140-percent more effective than lutein alone.²


Human Studies 

 
The first placebo-controlled study to evaluate the effects of a dietary supplement containing predominantly MZ was conducted in our Miami research laboratory.³² The supplement contained all three macular carotenoids in a ratio, MZ:lutein:zeaxanthin, of ~11:4:1 (total 21.8 mg) and was consumed daily over a 120-day period. MZ was effectively absorbed into the serum, and macular pigment density was increased significantly in many subjects in the supplementation group. No such increases were observed in the placebo group. In another study, 19 subjects consumed a supplement also composed of all three macular carotenoids in a ratio, MZ:lutein:zeaxanthin, of 7:9:1 (total 20 mg) over a period of 22 days. Results demonstrated that MZ was absorbed and reached serum concentrations of approximately 0.24 µmol/L.³⁴ Interestingly, serum levels of MZ among women were almost three times higher than men. At the Waterford Institute of Technology, the Meso-zeaxanthin Ocular Supplementation Trial (MOST), was conducted to evaluate macular pigment response and serum carotenoid response in subjects with and without AMD, following consumption of a supplement containing all three macular carotenoids in which MZ was predominant.¹⁷ This study identified statistically significant increases in serum concentrations of MZ and lutein from baseline. Significant increases in central macular pigment levels were also observed after just two weeks of supplementation. Furthermore, in patients who had an atypical macular pigment spatial profile (i.e., a central dip) at baseline, the more typical macular pigment profile (i.e. highest macular pigment at the center) was observed after eight weeks of supplementation with this MZ-dominant supplement.


MZ and AMD 

In a case-control study, we affirmed by comparing eyes of individuals with and without AMD that high levels of the macular carotenoids measured in the retina are associated with a reduction in the risk of AMD.35 In a similar small exploratory study, we investigated differences in the amounts of individual macular carotenoids between AMD and healthy donor retinas.³⁶ In the central 3 mm, the AMD group had, on average, ~ 30-percent less MZ and ~ 30-percent less lutein, but only ~ 20 percent less zeaxanthin than the healthy group. A larger study is needed to further investigate this differential deficiency in MZ and lutein in AMD eyes.

In 2001, AREDS, the Age-Related Eye Disease Study, a randomized, placebo-controlled trial of more than 5,000 patients, demonstrated that supplemental dietary antioxidants were of visual benefit to patients at risk of advanced AMD.³⁷ This important finding represented the first proof of principle that dietary antioxidants could retard the progression of AMD. Subsequently, because MZ, lutein and zeaxanthin have the ideal anatomic, optical and biochemical properties to confer protection against AMD, supplements containing these macular carotenoids have been recommended by eye-care professionals for the last decade, and studies have confirmed their benefit in terms of visual improvement and morphological progression of the condition.^38,39

Given the synergistic antioxidant properties of the three macular carotenoids, and that MZ is a major carotenoid component in the foveal center where a lack of macular pigment is associated with increased risk of AMD, one is driven to conclude that a supplement must contain all three macular carotenoids. This rationale, supported by recent clinical evidence, has prompted many eye-care professionals to recommend a formulation containing MZ, lutein and zeaxanthin to patients with AMD or at high risk of developing the condition. 

References are avaliable on request.


Dr. Bone is a distinguished experimental biophysicist and professor in the department of physics at Florida International University in Miami. Dr. Landrum is an internationally recognized research scientist and professor of chemistry and biochemistry at FIU. 

Dr. Beatty is the director of the Institute of Vision Research and the Macular Pigment Research Group, Waterford, Ireland. Dr. Nolan is a senior scientist and director at the same institution.