N. Nicolaides

In Vivo Transillumination Biomicroscopy and Photography of Meibomian Gland Dysfunction: A Clinical Study

Meibomian gland transillumination biomicroscopy and infrared photography were performed on 18 patients with clinically evident meibomian gland dysfunction (MGD) and dermatologic rosacea, 22 patients having MGD without evidence of dermatologic rosacea, and 15 unaffected individuals who served as controls. All patients having clinical signs of MGD demonstrated morphologic abnormalities of their meibomian glands by transilluminated biomicroscopy. Patients without dermatologic rosacea were noted to have varying degrees of gland distortion. Moreover, infrared photography documented a loss of the normal grape-like clusters of dark spots that represent the gland, suggesting a loss of glandular acini. Patients with dermatologic rosacea had more severe alterations, including marked distortion and loss of normal gland anatomy. There were no such abnormalities in clinically unaffected individuals. These data demonstrate that transilluminated biomicroscopy and infrared photography have the ability to identify a spectrum of morphologic alterations of the meibomian glands in MGD patients. The authors suggest that these techniques could be used to classify clinical MGD based upon the presence or absence of identifiable meibomian gland abnormalities.

The fatty acids of wax esters and sterol esters from vernix caseosa and from human skin surface lipid.

Separation of sterol esters from wax esters in the lipids of vernix caseosa and adult human skin surface was accomplished by column chromatography on MgO. The fatty acids of the sterol esters and wax esters of both samples were separated into saturates and monoenes, and examined in detail by gas liquid chromatography (GLC). The saturated fatty acids of the wax esters of vernix caseosa and of adult human skin surface were remarkably similar. They ranged in chain length from at least C11 to C30, six skeletal types being present: straight even, straight odd, iso, anteiso, other monomethyl branched and dimethyl branched. A large number of patterns of monoenes were observed, each pattern consisting of desaturation of a specific chain at Δ6 or Δ9 plus its extension or degradation products. The mole per cent of the total Δ6 and Δ9 patterns of wax ester fatty acid monoenes of vernix caseosa were 87% and 12%, respectively, and 98% and 1%, respectively, for adult human skin surface lipid. The sterol ester fatty acids of vernix caseosa were much different from those of adult human skin surface: vernix caseosa saturates were largely branched and of lengths greater than C18, whereas the saturates of adult human surface lipid resembled the wax ester fatty acids. Of the vernix caseosa monoene patterns, the mole per cent was 30% Δ6 and 70% Δ9, whereas of the adult human skin surface sterol ester fatty acids 89% were Δ6 and 11% Δ9. Chain extension was particularly pronounced in the sterol ester fatty acid monoenes of vernix caseosa amounting to 7–8 C2 units in some cases. The fatty acids of the sterol esters of both vernix caseosa and adult human skin surface appear to be derived from the sebaceous gland and from the keratinizing epidermis, but those of the wax esters are from the sebaceous glands only.

Diester waxes in surface lipids of animal skin

The literature is surveyed on two types of diester lipid that occur on the skin surfaces of animals: Type 1, a hydroxy fatty acid of which the hydroxyl group and the carboxyl group are esterified respectively with another fatty acid and a fatty alcohol, and Type 2, and alkane α,β-diol of which each OH group is esterified with a fatty acid. New data presented here show that the cow, rabbit and cat produce Type 1, whereas the dog, mouse, guinea pig and baboon produce Type 2 diesters. Each occurs as a major component of the surface lipid. The homologue distribution is given for Type 1 diesters of cow, rabbit and cat as well as the Type 2 diesters of dog and mouse. Distribution of long chain fatty acids of Type 1 diesters parallels that of the fatty alcohols suggesting a biogenetic relation between the two types of compounds. GLC of total diesters for the cow suggests that the components are assembled randomly during biosynthesis. Molecular weight of these diesters are in the range of those of natural triglycerides composed mainly of C16 and C18 fatty acids.

The Di- and triesters of the lipids of steer and human meibomian glands

Three groups of diesters have been isolated and identified in the lipids of steer meibomian glands. The first group, designated as α Type I, with the abbreviated formula FA-αOHFA-FA1c, consisted of α-hydroxy fatty acids esterified to fatty acids and fatty alcohols in the approximate molar ratio 1∶1∶1. The second group, designated as ω Type I-St, with the abbreviated formula FA-ωOHF A-St, consisted of ω-hydroxy fatty acids esterified to fatty acids and sterols in the approximate molar ratio 1∶1∶1. The third group, designated as α,ω Type II, with the abbreviated formula FA-α,ωdiol-FA, consisted of α,ω-diols esterified to 2 moles of fatty acids. The sum of the different diesters comprised about 9% of total steer meibomian lipids.Capillary GLC of the fatty acids of αType I diesters showed the fatty acids to be a family with a two-cluster profile, one at C12 to C20 and the other at C21 to C31, with anteiso chains predominating. Fatty acids from ωType I-St and α,ωType II diesters gave mainly a one-cluster profile in the short long chain, C23 to C30, with anteiso chains predominating, while the α-hydroxy fatty acids were short chain C13 to C18 acids with C16 predominating. The sterols in diesters ωType I-St were cholesterol (∼60%), Δ7 cholestenol (∼35%) and an unidentified compound (∼5%) with a GLC retention time slightly longer than Δ7 cholestenol on SE-30 phase. The ω-hydroxy fatty acids and α,ω-diols both were of exceedingly long chain lengths, C29−C38, and showed similar GLC profiles. Two types of triesters comprising approximately 1% of total steer meibomian lipids have been isolated but incompletely characterized. In terms of molar ratios, one group of triesters gave fatty acids:ω-hydroxy fatty acids:α-hydroxy fatty acids:sterols + fatty alcohols as approximately 1∶1∶1∶1. The other contained fatty acids, α-hydroxy fatty acids and α,ω-diols in what appears to be a complex mixture of several triesters. Diesters ωType I and α,ωType II also were found in human meibum. Hitherto these two diesters have not been found in any animal tissue.

Fatty acid composition of the living layer and stratum corneum lipids of human sole skin epidermis.

Lipids from the living layer and stratum corneum of human sole epidermis were extracted, saponified and the fatty acids analyzed. The proportion of fatty acids to unsaponifiables (mainly cholesterol), was higher in the living layer than in the stratum corneum. Fatty acids of the living layer and stratum corneum of human sole epidermis comprise saturates, monoenes, dienes, traces of polyenes and α-hydroxy fatty acids. Homologs of monoenoic and dienoic fatty acids for both living layer and stratum corneum lipids have a similar distribution. C16 and C18 were major components for each type of acids. There appeared to be two clusters, especially for saturates of both living layer and stratum corneum acids. One of these clusters ranged from C12 to C20 with C16 or C18 as a maximum and the other ranged from C21 to C30 with C24 as a maximum. The proportion of saturated acids with chain length C20 and above was much higher in the stratum corneum than in the living layer. Position isomers of the monoenoic fatty acids for both the living layer and stratum corneum show a predominance of ω9 acids, due to the overwhelmingly large amount of oleic acid. Linoleic acid was by far the major component of the dienoic acids. Homolog distribution of α-hydroxy fatty acids for the living layer was similar to that of the stratum corneum and again two clusters of acids below and above C20 with maxima at C16 and C24 were noticeable. Comparison of epidermal acids with those of sebaceous glands showed that each tissue can synthesize the same kind of acids but in widely different amounts. Oxidation of palmitate and stearate could supply the necessary energy for the late stages of keratinization.

The structures of the branched fatty acids in the wax esters of vernix caseosa

By combined gas liquid chromatography-mass spectrometry a series of monomethyl branched fatty acids was found in the fatty acid moiety of the wax esters of vernix caseosa. The methyl branch occurred on the even C-atoms of chains ranging from C11 to C17 (some 43 compounds in all). Except for the iso acids and possibly some of the anteiso acids, these could be formed by replacement of malonyl CoA with a molecule of methyl malonyl CoA at the point of the branch. Smaller amounts of fatty acids also were found with two methyl branches occurring on the even C-atoms of chains ranging from C9 to C15.

Further studies of the saturated methyl branched fatty acids ofVernix caseosa lipid

By the method of capillary gas chromatography-mass spectrometry, we have identified 35 monomethyl and 46 dimethyl branched acids in the saturated acids of vernix caseosa lipid with chain lengths ranging from C11 to C18. Many other mono-, di-, and trimethyl branched acids have been partially identified. All methyl branches were found to be on the even numbered C-atoms except for some terminal iso methyl groups. Three types of dimethyl branched acids were found: those with a terminal iso structure, those with a terminal anteiso structure, and those with neither iso nor anteiso structures. The 4-methyl branch predominated for all types of branched acids. Equivalent chain length data for di- and trimethyl branched acids were determined on a Pentasil (nonpolar) wall coated capillary column and checked by calculation from monomethyl branched acid data. Mass spectral identification was performed with and without the aid of a data system. A possible mode of formation of these acids is discussed.

The monoene and other Wax alcohols of human skin surface lipid and their relation to the fatty acids of this lipid

Abstract1)Wax alcohols (as acetates) were isolated from human skin surface lipid and separated into a saturated and a monoene fraction.2)Four main chain types were found for both saturated and monoene alcohols: normal even, normal odd, iso and anteiso. (“Even” and “odd” refer to the number of C-atoms in the straight chain.)3)The monoene alcohol acetates were separated into homologues of each chain type by preparative gas-liquid chromatography (GLC) and the positions of the double bonds for each homologue were determined by analytical GLC of the original fraction, its hydrogenated derivative, and the products it formed by reductive ozonolysis.4)The fragments formed by reductive ozonolysis of the monoene alcohol acetates were compared to those formed from the total monoenoic fatty acids (as methyl esters), both obtained from the same sample of surface lipid. (Comparisons were best made by ozonolysis of a portion of the entire sample of each ester group.a)The terminal ends of both groups of monoene fatty chains yielded a very similar pattern of aldehydes in terms of types and amounts. This could be explained by the hypothesis that both fatty acid and fatty alcohol chains of lengths ranging mainly from C14 to C18 were first biosynthesized, then desaturated at Δ6.b)The functional group ends gave a distinct pattern of aldehyde esters for the acids and another for the alcohols. Both patterns consisted nearly entirely of members having aneven number of C-atoms from the double bond to the functional group. This suggested that the members of each pattern were formed by chain extensions of an integral number of C2 units beyond the lengths arrived at in 4a). Thus 71% of the fatty acid monoenes were not extended, 25% were extended by 1 C2 unit and the remainder extended from 2 to 5 C2 units, whereas nearly all the fatty alcohols were extended mainly by 2, 3 or 4 C2 units, with decreasing amounts up to 8 C2 units.5)A small amount (∼5%) of odd chain aldehyde esters for both fatty acids and fatty alcohols were found and some unidentified alcohols were detected.

The structure of alkane diols of diesters in vernix caseosa lipids

cles were sedimented as previously described (3). Autoradiograms obtained showed that the major incorporation of radioactivity into triglycerides occurred in the fraction rich in these lipoprotein particles. These results are identical to those obtained by Patton et aL (3) with freshly secreted milk synthesizing systems from cows and goats. Since results presented herein were readily duplicated with sows of different breeds, they suggest that freshly secreted swine milk contains a well organized system for glyceride synthesis. Apparently this system contains all enzymes and cofactors necessary for net synthesis of glycerides. Although much further work is required to determine the relationship of this system to in vivo milk fat synthesis, the ease of obtaining this physiologically available enzyme source should facilitate the task. These data confirm the earlier work of McCarthy and Patton (1) and suggest that this glyceride synthesizing activity is widespread i n freshly secreted milks. T. W. KEENAN

Fatty acids of unusual double-bond positions and chain lengths found in rat skin surface lipids.

The fatty acids of rat skin surface lipids comprise four main skeletal types of chains which occur both as saturates and monoenes and range from C12 to C38: straight even, straight odd, iso and anteiso (the latter two identified by GC retention data only). Two unidentified series of branched monoenes also occur in trace amounts.Reductive ozonolysis of monoenes reveals two characteristic double-bond position patterns, one for the straight even chain series and the other for the straight odd chain series. The straight even chain pattern comprises four series, of which ω7 ≫ω9>ω5>ω11; the straight odd chain series in contrast shows a large number of ω series with irregular distribution. The biosynthesis of the even chain fatty acid monoenes can be thought of as occurring in two stages: synthesis of 14∶Δ9, 16∶Δ9, 18∶Δ9 and 20∶Δ9, with 16∶Δ9 predominating; elongation of these chains mostly by 1, 2, or 3 C2 units but up to the unusually long lengths by 11 C2 units. For the formation of the former, two schemes by known pathways are proposed.Iso and anteiso chains which are nearly all saturated comprised 1/3 the total fatty acids.