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Keywords:

  • antioxidant;
  • beer;
  • fermentation;
  • kimchi;
  • melatonin;
  • melatonin isomer;
  • nomenclature;
  • wine

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

Abstract:  Melatonin was considered to be the sole member of this natural family. The emergence of naturally occurring melatonin isomers (MIs) has opened an exciting new research area. Currently, several MIs have been identified in wine, and these molecules are believed to be synthesized by either yeasts or bacteria. A tentative nomenclature for the MIs is proposed in this article. It will be important to explore whether all organisms have the capacity to synthesize MIs, especially under the conditions of environmental stress. These isomers probably share many of the biological functions of melatonin, but their activities seem to exceed those of melatonin. On basis of the limited available information, it seems that MIs differ in their biosynthetic pathways from melatonin. Especially in those compounds in which the aliphatic side chain is not attached to ring atom 3, the starting material may not be tryptophan. Also, the metabolic pathways of MIs remain unknown. This, therefore, is another promising area of research to explore. It is our hypothesis that MIs would increase the performance of yeasts and probiotic bacteria during the processes of fermentation. Therefore, yeasts producing elevated levels of these isomers might have a superior alcohol tolerance and be able to produce higher levels of alcohol. This can be tested by comparing existing yeast strains differing in alcohol tolerance. Selection for MIs may become a strategy for isolating more resistant yeast and Lactobacillus strains, which can be of interest for industrial alcohol production and quality improvements in bacterially fermented foods such as kimchi.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

The chemical structure of melatonin (N-acetyl-5-methoxytryptamine) was identified in 1958 by Lerner et al. [1]. Since then, the biosynthetic pathway of melatonin in vertebrates has been extensively studied, well documented, and widely accepted in many disciplines by scientists [2, 3]. Basically, four enzymes including tryptophan hydroxylase (TPH), aromatic amino acid decarboxylase (AAAD), arylalkylamine N-acetyltransferase (AANAT), and N-acetylserotonin methyltransferase (ASMT) [formerly called hydroxylindole-O-methyltransferase (HIOMT)] are involved; their successive activities lead to the formation of melatonin from its precursor, tryptophan (Fig. 1). In addition, an alternate pathway, in which the last two enzymatic steps of melatonin biosynthesis are reversed, is also possible, but usually of minor importance [4] (Fig. 1).

image

Figure 1.  The classic biosynthetic pathways of melatonin in vertebrates. TPH, tryptophan hydroxylase; AAAD, aromatic amino acid decarboxylase; NAT, N-acetyltransferase; HIOMT, hydroxyindole-O-methyltransferase; MelDA, melatonin deacetylase; the dashed line represents a minor pathway of melatonin biosynthesis.

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Originally, melatonin was considered as an exclusive product of the pineal gland in vertebrates, and it was, therefore, referred to as a neurohormone [5]. Subsequently, however, many cells, organs, and tissues were found to have the capacity to synthesize melatonin [6]. These include neutrophils, lymphocytes, astrocytes, microglia, retina, pancreas, ovary, testes, skin, bone marrow, and gut. The gastrointestinal tract is reported to produce significantly more melatonin than does the pineal gland [7, 8]. In addition to vertebrates, in the last two decades, melatonin has also been identified in a wide range of invertebrates [9–11] and in bacteria [12], algae [13, 14], fungi [15], and plants [16–21]. Considering this evidence, the concept of melatonin being a neurohormone has been challenged and is no longer tenable [22]. Evidence supports the conclusion that melatonin is a ubiquitously occurring substance present in almost all or all living organisms.

Little information is, however, available related to the biosynthesis of melatonin in organisms other than vertebrates, with a few exceptions. In the dinoflagellate Lingulodinium polyedrum (syn. Gonyaulax polyedra), the pathway of melatonin formation is identical with that of vertebrates [23], and enzymes involved in tryptophan hydroxylation [24], serotonin acetylation, and hydroxylindole methylation [25] have been studied in detail. Moreover, substitution experiments using the intermediate metabolites after inhibition of tryptophan hydroxylase have been conducted in this organism [24, 26]. In yeast, starvation/re-feeding experiments with tryptophan metabolites revealed that melatonin can be formed from tryptophan, serotonin, and N-acetylserotonin, but also from 5-methoxytryptamine (5-MT) [15]. In both Lingulodinium [23] and Saccharomyces [15], melatonin deacetylation to 5-MT has been identified as a catabolic pathway. In dinoflagellates, 5-MT is further metabolized to 5-methoxytryptophol and to the major end product, 5-methoxyindole-3-acetic acid, which is released to the water [23].

It does appear that the biosynthetic pathway of melatonin in plants, particularly in rice, is different from that in the pineal gland of vertebrates. In rice, the first enzymatic step of melatonin synthesis is tryptophan decarboxylation rather than hydroxylation as it occurs in animals [27]. In addition, the homolog of a rate-limiting enzyme, AANAT, which is present in vertebrates, has not yet been identified in higher plants to date [28, 29]. Likewise, knowledge of metabolic pathway(s) of melatonin, unlike in animals, is also essentially unknown in plants. The only melatonin metabolite which has been identified in plant is N1-acetyl-N2-formyl-5-methoxykynuramine (AFMK) [30]. AFMK is also one of the major melatonin metabolites in animals, which is generated by the interaction of melatonin with free radicals, singlet oxygen or via its conversion by enzymes such as 2,3-indolamine dioxygenase (IDO), myeloperoxidase, some hemoperoxidases, and also cytochrome c [6, 31]. In rice plants transgenic for IDO, there are much lower melatonin levels than in its wild-type counterpart [17]. This indicates that animals and plants may, at least, share a similar metabolic pathway regarding the formation of AFMK.

In addition to its many actions in animals [32–37], melatonin functions as an antioxidant in its capacity as a first line to defend against a variety of oxidative stressors in a organism [38]. This has been well documented in organisms including vertebrates [39–42], algae [14, 43, 44], and plants [45–47]. Other functions of melatonin including its role as the chemical expression of darkness [48], which is obviously true for vertebrates, have yielded inconsistent results in plants. For example, in at least one plant, the peak level of melatonin appears at night [49]; however, in other plants, these peaks occur during the day [30, 50, 51]. Moreover, it is even reported that high light intensity, including ultraviolet light, has a positive association with melatonin production in some plant species [50, 52]. Again, this contrasts with the situation in animals where light suppresses melatonin production [37].

The discoveries of melatonin in bacteria, algae, fungi, and plants open a new area of exploration and potentially novel aspects regarding the biosynthesis, metabolism, and other functional roles of this ubiquitously distributed indolamine. As new information related to melatonin accumulates daily, it is necessary for the scientists to reconsider the classic approaches to melatonin investigations and instead consider different perspectives. In the current article, we primarily discuss a completely new topic related to melatonin, that is, the naturally occurring melatonin isomers (MIs) and their proposed nomenclature.

Naturally occurring melatonin isomers

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

Following the characterization of melatonin in 1958 [1], little consideration was given to naturally occurring MIs. This was not surprising for several reasons. First, there were no commercially available MIs which could be used as assay standards. Secondly, the antibodies used for radioimmunoassay were developed using a melatonin-like 5-methoxylated, 3-substituted hapten, for example, N-acetyl-5-methoxytryptophan, coupled via carbodiimide to a carrier protein such as thyroglobulin. Therefore, isomers carrying methoxy residues and aliphatic side chains in different positions could not be recognized by antibodies. Frequently used immunological melatonin assays like RIA and ELISA are, thus, not suitable for detecting MIs. Even conventional HPLC procedures are not sufficiently sophisticated to identify unknown MIs or to distinguish melatonin from any isomer that may have existed, as long as separation and detection methods have not been adapted to this specific problem.

Using HPLC with mass/mass spectrometry (HPLC-ESI-MS/MS), Rodriguez-Naranjo et al. [53] were the first to report on the existence of a naturally occurring MI in wine. This MI shares the same main fragment ions from m/z 233 (174 and 216) as the melatonin standard when estimated by MS; however, the minor fragments of 196, 161, and 141 do not coincide with those of melatonin. This isomer also has a distinguishable HPLC retention time from the melatonin standard. Interestingly, Rodriguez-Naranjo et al. [53] noticed that wines from different producers had different patterns regarding the content of melatonin and the isomer; that is, some wines only contained melatonin, some contained the MI alone, and others contained both melatonin and its isomer.

Recently, using the same HPLC-ESI-MS/MS method, Gomez et al. [54] found a second potential MI in an extract of wine produced from the grape Vitis vinifera cv. Malbec. Compared with the MS fingerprint of melatonin of which the most stable abundant fragment is 174, the most abundant fragment of the isomer extracted from the wine is 216. On the basis of the data of Diamantini et al. [55], the structure of this isomer was tentatively identified as 1-(2-alkanamidoethyl)-6-methoxyindole. To date, at least two naturally occurring MI have been uncovered in wine. It is predicted that as the methods of identification improve, more naturally occurring MI will be subsequently identified in wine, beer, probably in plants and, perhaps, even in animals. To avoid any potential confusion of different MIs that may be identified and to aid the researchers, herein we propose a nomenclature of all potential naturally occurring MIs that may be characterized.

Proposed nomenclatures of potential naturally occurring melatonin isomers

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

The structure of melatonin has two distinguishable side chains, which are added to the indole ring; these are the methoxy group at position 5 (5-methoxy group) and N-acetylaminoethyl group at position 3 [3-(N-acetylaminoethyl) group] (Fig. 2). Theoretically, either one of these two side chains can be potentially relocated to any one of the seven positions in the indole ring to form MIs. On the basis of this, it is calculated that 42 combinations of these two side chains in seven different positions could be formed. Thus, there are 42 potential MIs. As other MIs will likely be identified, it seems important to introduce a logical nomenclature to identify these naturally occurring MIs. Herein, we propose that the N-acetylaminoethyl group be designated as side chain A (A) and the methoxy group be designated as side chain M (M). A MI can then be denoted according to the positions of A and M side chains at the seven positions on the indole ring (Fig. 3). For example, if the side chain A is at position 1 and side chain M is at position 6 of the indole ring, respectively, this molecule would be designated as melatonin isomer A1/M6 or MI A1/M6. Here, the capital letters A and M represent each of the two side chains, respectively, and the A always precedes M, and they are separated with a forward slash (/). The subscript numbers following the A or M represent one of the seven positions of the indole ring.

image

Figure 2.  Chemical structure of melatonin. The numbers identify the positions on the indole ring.

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image

Figure 3.  Chemical structures of proposed melatonin isomers. The numbers identify the positions on the indole ring and the A or M represents the side chain A and side chain M, respectively.

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Table 1 lists all 42 potential naturally occurring MIs according to the principles of nomenclature described here.

Table 1.   A list of the proposed nomenclature of melatonin isomers which may be present in organisms
NomenclatureSubstituent position in indole ring
Chain A (-CH2CH2NHCOCH3)Chain M (-OCH3)
Melatonin (M)35
Melatonin isomer (MI)A1/M212
MI A1/M313
MI A1/M414
MI A1/M515
MI A1/M616
MI A1/M717
MI A2/M121
MI A2/M323
MI A2/M424
MI A2/M525
MI A2/M626
MI A2/M727
MI A3/M131
MI A3/M233
MI A3/M434
MI A3/M636
MI A3/M737
MI A4/M141
MI A4/M242
MI A4/M343
MI A4/M545
MI A4/M646
MI A4/M747
MI A5/M151
MI A5/M252
MI A5/M353
MI A5/M454
MI A5/M656
MI A5/M757
MI A6/M161
MI A6/M262
MI A6/M363
MI A6/M464
MI A6/M565
MI A6/M767
MI A7/M171
MI A7/M272
MI A7/M373
MI A7/M474
MI A7/M575
MI A7/M676

Origin of melatonin and its isomers in wine

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

Melatonin was first identified in grapes by Iriti et al. [56]. The levels of melatonin in grapes reported by this group are in the range of pg/g tissue. Subsequently, several other groups also found melatonin in grapes with widely different concentrations (pg to μg/g tissue), depending on the cultivars of grape plants, harvest time, degree of ripeness, and environmental conditions such as light intensity [57–61]. Generally, the skin and the seeds contain more melatonin than does of flesh of grapes. Melatonin was also identified in wine by different research groups [53, 58, 62–65]. It seems that melatonin in grapes and in wine is an universal phenomenon. It has also been speculated that the health benefits associated with the consumption of grapes or the consumption of wine may be partially attributed to the melatonin they contain [66, 67]. The data from animal studies also generally support this idea when the protective effects of melatonin were compared with those of resveratrol in an ischemia/reperfusion heart injury model in mice, melatonin proved to be more protective [68]. Resveratrol, which is present in red wine, is known to have health benefits [69, 70].

It was assumed that the origin of melatonin in wine is attributed to the grapes from which the wine was produced. Some indirect evidence supports this suggestion. For example, one study showed that grape plants treated with agrochemicals, which increase their resistance to environmental insults and their melatonin levels, also enhanced the melatonin contents of the red wine produced from these grapes [58]. Red wine contains more melatonin than does white wine, and red grapes were reported to have much higher levels of melatonin than do white grapes [66]. However, differences in the fermentation and postfermentation processes have to be considered, in addition to grape cultivars. In red wine fermentation, skins are included and higher temperatures are used than in white or rosé wines. These differences may be tentatively tested by comparing red and rosé wines, both of which are produced from red grapes.

Recent studies, however, have provided evidence that melatonin in wine is not exclusively from the grapes but is mainly generated during the process of fermentation during wine brewing; specifically, melatonin in wine is a consequence of its synthesis by yeast (EC-1118) [54, 62]. It should be noted that grapes are also covered by yeasts different from Saccharomyces, such as Pichia, Kloeckera, and Torulopsis, which may be particularly important in red wine fermentation, during which skins are floating for an extended time on the top of the must. In addition to yeasts, several bacteria may also inadvertently participate in the process of malolactic fermentation of wine brewing at different stages [71, 72]. The potential contributions of bacteria to the melatonin content of wine cannot be ignored [62]. Interestingly, considerable amounts of the MI(s) is (are) also generated during the process of wine brewing [53]. In some cases, the reported levels of MIs in wine are approximately 20-fold higher than that of melatonin itself (Fig. 4) [54]. It is interesting that the production of melatonin and the MI(s) during the process of wine fermentation is independent of external tryptophan; that is, without tryptophan in the medium, the yeasts can still synthesize these indolamines. However, the precursor tryptophan may have been synthesized intracellularly by either yeast or bacteria, both of which possess the shikimic acid pathway of aromatic biosynthesis [73]. Therefore, a conclusion relative to a tryptophan-independent pathway cannot be drawn from the absence of the amino acid in the medium. However, this may be assumed because of the position of the N-acetylethylamine group different from ring atom 3 in an isomer [54], although secondary isomerizations of precursors should not be ruled out at the present stage of knowledge. Nevertheless, it seems important to clarify this issue, because a completely new pathway of melatonin biosynthesis different from the classic one starting with tryptophan would considerably change our understanding of indoleamine metabolism in nonvertebrate organisms.

image

Figure 4.  Development of melatonin and its isomer during the process of wine making. Data are expressed as means ± S.E.M. of three independent determinations. This figure is modified from Gomez et al. [54].

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The data also show that the amount of the MI(s) formed during the process of wine fermentation has little relation to the melatonin concentrations, which were intentionally added to the medium. The MI(s) is/are probably not modified from the melatonin molecule itself but rather is/are generated by yet unknown pathways. Current data add new challenges to our understanding, or lack thereof, of melatonin synthesis and metabolism as a whole.

Levels of melatonin and its isomers in wine

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

The actual concentration of melatonin in wine is a subject of continuing debate. The reported values range from pg to ng/mL. There is little doubt that different brands of wine or different strains of yeast in wine brewing may partially explain these great variations in melatonin concentrations. It is likely, however, that such huge differences of melatonin levels in wine may be mainly a result of different methods of assays. Rodriguez-Naranjo et al. [53] estimated the melatonin levels in the same wine using both the ELISA method and HPLC-MS/MS. The results showed that melatonin levels in wine measured by HPLC-MS/MS were 1–3 orders of magnitude higher than those measured by ELISA. For example, the level of melatonin in Tempranillo wine measured with ELISA was around 140 pg/mL while it reached 130 ng/mL when measured by HPLC-MS/MS. Additionally, the latter method also has indicated that this same wine contained another 9.3 ng/mL of a MI. The authors propose that ELISA is not a suitable method for measuring melatonin content in wines and that HPLC methods are valid for this purpose. However, HPLC alone could not detect MI(s) in wines without reference standards of MIs. Considering that in some wines, the levels for the MI(s) is/are much higher than that of melatonin itself, the levels of melatonin plus its isomers in the wine are markedly underestimated in the published literature, as the assays used were incapable of recognizing any MI that may have been present. To obtain accurate levels of melatonin and its isomers in wine requires advanced measuring methods such as HPLC-MS/MS and synthetic MIs as reference standards.

Predictions of high levels of melatonin and its isomers in other fermentation processes

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

The general fermentation processes usually involve the use of yeasts and a variety of bacteria. There is evidence that yeast has the capacity to synthesize high levels (μm) of melatonin in culture [15]. Thus, it is not, perhaps, surprising that melatonin in wine is synthesized by yeast per se as has been previously predicted [28]. What is often of greater interest is that yeast can synthesize MIs during the process of fermentation. There is little published information on naturally occurring MIs even though several synthetic MIs have been studied [55]. The discovery of naturally occurring MIs opens a new investigative area including defining their synthetic/metabolic pathways, their potential biological roles, and the health and nutritional implications of these molecules.

It is our hypothesis that bread, beer, and cider which are fermented with yeast have an increased likelihood of containing relatively high levels of melatonin and its isomers. The indolamines in these products may simply be derived from fungi, or they may also be contained in extracts from wheat, barley, and apples, respectively.

As mentioned earlier, many probiotics are used in the fermentation of wine brewing, and it is likely that their use may explain, at least in part, the naturally occurring MIs in wine. The use of probiotics to synthesize melatonin has been patented in USA, and industrial scale melatonin production has employed the following probiotics: Bifidobacterium species (breve subspec. breve, longum subspec. infantis); Enterococcus species (faecalis TH10); Lactobacillus species (brevis, acidophilus, bulgaricus, casei subspec. sakei, fermentum, helveticus subspec. jogorti, plantarum); Streptococcus (thermophilus). These melatonin products are marketed by Quantum Nutrition Labs (http://healthline.cc). It is also anticipated that high levels of melatonin and MIs will be found in cheese and yogurt owing to several probiotics that are used in their production. Interestingly, cheese and yogurt are major components of classic Mediterranean diets, which reportedly have beneficial effects for human health. Likewise, high levels of melatonin and MIs would be expected in healthy oriental foods, such as in kimchi, soy sauce, and homemade vinegar in which Lactobacillus species are used in their fermentation. If these speculations are valid, the beneficial effects in consumption of these products may, at least partially, be explained by the presence of melatonin and its isomers.

The gut plays host to large number of symbiotic microorganisms, and high levels of melatonin consistently have been reported in gut [8]. Whether melatonin in the gastrointestinal tract is mainly produced by gut tissue per se or primarily a result of gut symbiotic microorganisms is unknown. A recent study shows that in mammals, particularly in rats, gut microbiota modulate host systemic metabolism including a possible compensatory mechanism for melatonin production [74]. Another site which may have high concentrations of melatonin and its isomers is the first compartment of the stomach of ruminants. The rumen microflora contains rich populations of several species of bacteria, protozoa, and sometimes yeasts [75], all of which have the capacity to synthesize melatonin [10] and possibly its isomers.

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References

The discovery of naturally occurring MIs synthesized by yeast and possibly by probiotics raises some fundamental questions. For example:

  • 1
     Could other species including humans have a capacity to synthesize MIs? Currently, there is obviously no conclusive answer to this question because of insufficient information on MIs themselves. It is our current hypothesis that the production of naturally occurring MIs may be induced by environmental stressors. During the process of fermentation in wine making, the concentration of alcohol in wine gradually increases until it kills the yeasts. To increase the tolerance for the elevated levels of alcohol in the environment, one of the biological responses of yeast may be to upregulate the generation of members of the melatonin family, especially MI(s). This hypothesis is supported by the observation on development of melatonin and its isomer(s) during the process of wine brewing. During the process of fermentation (which is associated with gradually increased alcohol levels in the wine), melatonin levels only rise gradually. However, the levels of MI(s) increase markedly with time to the end of the fermentative process (Fig. 4). This indicates that the inducibility of MI(s) owing to environmental stressors, particularly in yeast, is much greater than that of melatonin per se.

For bacteria, such as the Lactobacillus species, it may also be essential for them to generate increased amounts of melatonin and/or MIs to tolerate the gradual drop in the environmental pH which is the result of elevated lactic acid production by these bacteria. It has been well documented in other species that melatonin production is also inducible as a result of environmental stress, especially oxidative stress. This has been observed in algae [14, 43, 76], plants [47, 52, 77], and animals including humans [78–83]. Dramatic rises in melatonin levels are induced by moderate temperature decreases in the dinoflagellates L. polyedrum and Alexandrium lusitanicum [23, 84, 85] and by heat stress in a tropical strain of Amphidinium carterae [23], treatments that may also be associated with oxidative stress [43].

From an evolutionary point of review, if MIs provide benefit against unfavorable environmental factors, other organisms such as those mentioned earlier should also have the capacity to produce MIs, especially under stressful conditions. This could be proven/disproven with advanced detection methods such as the HPLC-MS/MS and the availability of reference standards for the MIs.

  • 2
     What are the biological functions of MIs? Melatonin exhibits pleiotropic biological functions [86, 87] including regulation of seasonal reproductive physiology [88], circadian rhythms [89], immunoreactivity [34, 90], lipid metabolism [91], sleep [92] and the antioxidant capacity in species from bacteria to mammals [86, 93, 94]. The pleiotropic functions of melatonin involve melatonin receptor-dependent and receptor-independent mechanisms [95–98]. Tarzia et al. [99] have tested a synthetic MI (MI A1/M6) regarding its ability to bind to melatonin membrane receptors as well as its biological activity. The results indicate that this isomer has a similar affinity binding with melatonin membrane receptors as melatonin itself and it also exhibits full agonistic activity in lowering intracellular cAMP levels. In a follow-up study [100], this group observed that the synthetic MIs induced completely different biological consequences after binding to melatonin membrane receptors. They behaved as agonists or as antagonists depending on the position of the chain A in the indole ring. As to the receptor-independent activities such as their antioxidant and cytoprotective activities, MIs have diverse effects. Spadoni et al. [101] reported that changing the positions of either side chain A or side chain M in the indole ring results in marked alterations regarding their antioxidant and cytoprotective capacities. For example, if side chain M is located in the position 4 of the indole ring, this MI possesses the strongest antioxidant capacity while if the side chain A is in position 3, the isomer is most effective as a cytoprotective agent. These diverse functions of receptor-mediated and receptor-independent activities of MIs may partially explain the multifaceted functions of melatonin in organisms, if in fact the MIs are found to be naturally produced in all organisms.
  • 3
     What are the synthetic and metabolic pathways of the MIs? On the basis of information summarized earlier, MIs seem to have a different synthetic pathway from that of melatonin in vertebrates. In at least one category of isomers, the starting material for the synthesis of MIs may not be tryptophan. However, generalizations should be avoided with regard to the present state of knowledge and the low number of isomers identified. Nevertheless, it seems important to distinguish between isomers concerning the position of the methoxy group and the N-acetylaminoethyl side chain. Variants carrying methoxy residues in varying positions at the indolic moiety may easily derive from tryptophan or tryptamine by hydroxylation and subsequent O-methylation. Numerous monooxygenases with different substrate specificities exist in plants, fungi, and bacteria and also broad-spectrum O-methyltransferases. This is already evident from the existence of many indole alkaloids of plant and fungal origin which carry methoxy groups in positions corresponding to ring atoms 4, 6, and 7 of melatonin. In plants, the substrate spectra of aromate hydroxylating enzymes have been poorly characterized, in part because the focus of interest was mostly directed to monooxygenases involved in aliphatic hydroxylation in the course of auxin biosynthesis [102]. Moreover, isomerization of hydroxyindoles is known, for example, in the case of the 5-hydroxylated indoleamine bufotenin, which is converted via bufotenidine to the 4-hydroxylated psilocin [103]. To date, there is no evidence for isomerization of melatonin itself. Nevertheless, this may occur with its precursors. Contrary to the variants carrying methoxy groups in positions different from ring atom 5, isomers that differ from melatonin in the position of the N-acetylaminoethyl side chain cannot be easily attributed to tryptophan metabolism. This suggests that several unknown enzymes which are not required for melatonin synthesis may be involved in the synthetic pathway for MIs. At this point, there is little information on the metabolism of MIs; this is a new area to be explored in the future.
  • 4
     What are the potential applications of these findings? As mentioned earlier, significantly increased levels of MIs during the process of fermentation enhances the capacity of these organisms to tolerate stressful environments such as low pH or high alcohol concentrations. It has been reported that a commercially important lactic acid bacterium (Leuconostoc mesenteriodes), which is used as a starter culture in kimchi fermentation, is sensitive to acid stress. However, when exogenous glutathione (GSH), an antioxidant, was added to the culture medium, this bacterium became more resistant to acid stress, and as a result, it also improved its performance in kimchi fermentation [104]. Melatonin is a stronger antioxidant than GSH [97], and this also applies to some MIs [101]. In addition to its great antioxidant capacity, melatonin also easily penetrates into cells because of its small molecular weight and its characteristic amphiphilicity [105]. It is likely that if MIs were introduced into the kimchi fermentation process, they would improve the performance of lactic acid bacteria and generate higher quality kimchi with great nutritional or medicinal value. This application could also be applied indirectly to beer or wine brewing using yeast strains selected for high isomer levels. This may also enhance the alcohol tolerance of yeasts. A study has identified, in a series of different brands of beer, that higher levels of alcohol are associated with greater concentrations of melatonin [106]. While alcohol tolerance is not the limiting factor for alcohol levels in beer, but rather the amount of original wort used for fermentation, and as further increases of alcohol in wine may not be desirable under enological aspects of balanced taste, more efficient industrial ethanol production may become a field of application for MIs, for example, production of alcohol as a fuel.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Naturally occurring melatonin isomers
  5. Proposed nomenclatures of potential naturally occurring melatonin isomers
  6. Origin of melatonin and its isomers in wine
  7. Levels of melatonin and its isomers in wine
  8. Predictions of high levels of melatonin and its isomers in other fermentation processes
  9. Discussion
  10. References