Aliphatic beta-nitroalcohols for therapeutic corneoscleral cross-linking: chemical mechanisms and higher order nitroalcohols.
OBJECTIVE
The recent tissue cross-linking studies indicate that aliphatic beta-nitroalcohols (BNAs) may be useful as pharmacologic corneoscleral cross-linking agents. The present study was performed to identify the specific chemistry involved under physiologic conditions, with the intent of identifying more effective agents.


METHODS
The mechanism of chemical cross-linking at pH 7.4 and 37 degrees C was studied using three techniques. The colorimetric Griess assay was used to follow the release of nitrite from three mono-nitroalcohols (2-nitroethanol [2ne], 2-nitro-1-propanol [2nprop]), and 3-nitro-2-pentanol [3n2pent]). Second, the evolution of 2nprop in 0.2 M NaH(2)PO(4)/Na(2)HPO(4)/D(2)O was studied using (1)H-NMR. Third, thermal shrinkage temperature analysis (T(s)), a measure of tissue cross-linking, was used to support information from (1)the H-NMR studies.


RESULTS
A time-dependent release of nitrite was observed for all three mono-nitroalcohols studied. The maximum levels were comparable using either 2ne or 2nprop (approximately 30%). However, much less (approximately 10%) was observed from 3n2pent. Using (1)H-NMR, 2nprop evolved into a unique splitting pattern. No match was observed with reference spectra from three possible products of denitration. In contrast, 2-methyl-2-nitro-1,3-propanediol (MNPD), a nitro-diol, was identified, implying the formation of formaldehyde from a retro-nitroaldol (i.e., reverse Henry) reaction. In support of this mechanism, T(s) shifts induced by the nitro-triol 2-hydroxymethyl-2-nitro-1,3-propanediol (HNPD) were superior to the nitro-diol MNPD which were superior to the mono nitroalcohol 2nprop.


CONCLUSIONS
BNAs function as both formaldehyde and nitrite donors under physiologic conditions to cross-link collagenous tissue. Higher order BNAs are more effective than mono nitroalcohols, raising the possibility of using these agents for therapeutic corneoscleral cross-linking.
Department of Ophthalmology, Columbia University, College of Physicians and Surgeons, New York, New York 10032, USA. dcp14@columbia.edu
David C
DC
Paik
Marissa R
MR
Solomon
Quan
Q
Wen
Nicholas J
NJ
Turro
Stephen L
SL
Trokel
D000818:Animals; D003094:Collagen / Q000378:metabolism*; D003124:Colorimetry; D003315:Cornea / Q000378:metabolism*; D003432:Cross-Linking Reagents; D000431:Ethanol / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D009573:Nitrites / Q000378:metabolism; D009574:Nitro Compounds / Q000737:chemistry*; D000439:Pentanols / Q000737:chemistry*; D020005:Propanols / Q000737:chemistry*; D012590:Sclera / Q000378:metabolism*; D013552:Swine
C496417:2-nitropropanol; C539264:3-nitro-2-pentanol; D003432:Cross-Linking Reagents; D009573:Nitrites; D009574:Nitro Compounds; D000439:Pentanols; D020005:Propanols; D000431:Ethanol; D003094:Collagen