Interindividual Variability of Enkephalin-Degrading Enzymes in Human Plasma

Marini, Mario; Urbani, Alessandra; Trani, Eugenia; Bongiorno, Lucilla; Roda, L.Giorgio

doi:10.1016/s0196-9781(97)00129-0

Cited by 15 publications

(14 citation statements)

References 30 publications

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“…Finally, the importance of the low molecular weight inhibitors in controlling enkephalin hydrolysis does not seem to be peculiar to the group examined here. On the contrary, in other cases examined, inhibitors appear to play a predominant role in regulating plasma enzyme hydrolysis: these substances are primarily responsible for interindividual [26] as well as intraspecific [27] differences in the hydrolysis of enkephalin and in the formation of its hydrolysis byproducts, and play an essential role in inducing the hydrolysis modifications associated with the allergy-induced activation of the immune system (data submitted).…”

Section: Discussionmentioning

confidence: 99%

“…Sheets were cut according to internal standards, and transferred to scintillation vials. Radioactive label was extracted, and samples counted as reported elsewhere [26].…”

Section: Chromatographic Proceduresmentioning

confidence: 99%

“…Plasma enzymes were fractionated by a 5-Ìm 7.8 ! 300 mm TSK G3000SW column (Toyo Soda Ltd., Tokyo, Japan) under the conditions described elsewhere [26]. One-milliliter fractions were collected and assayed for enzyme activity as described above.…”

Section: Chromatographic Proceduresmentioning

confidence: 99%

“…Since the increase in hydrolysis attributable to aminopeptidases exceeds the decrease attributable to dipeptidylpeptidases, especially to dipeptidylcarboxypeptidases, the result is an overall increase in substrate hydrolysis. Finally, the differences in the shape of the curves representing the formation of hydrolysis byproducts, evident at the shorter hydrolysis times and especially noticeable in the case of dipeptidylaminopeptidases, may suggest the existence of ageinduced differences in the relative activity of the multiple enzymes within each of the three enzyme classes [20,26].…”

Section: Hydrolysis In Whole Plasmamentioning

confidence: 99%

“…Comparing leu-enkephalin hydrolysis in whole and in K) 0-0.6 plasma ( fig. 3) indicates that removing inhibitors, which increases substrate hydrolysis only modestly, perceptibly modifies the relative ratio of the hydrolysis byproducts formed [26]: the activity of aminopeptidases is increased (at 600 s to 226% of the value measured in whole plasma), while that of the dipeptidylaminopeptidases and dipeptidylcarboxypeptidases is decreased (at 600 s to 29 and 23%, respectively, of the activity in whole plasma). These data can be interpreted as resulting from considerable inhibition of aminopeptidases, paired with much lower inhibition of dipeptidylpeptidases.…”

Section: Low Molecular Weight Inhibitorsmentioning

confidence: 99%

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Age Increases Leucine Enkephalin Hydrolysis in Human Plasma

Bongiorno

Marzano²,

Marini³

et al. 1998

Gerontology

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Background: The existence of age-associated alterations in immune functions and neuropeptides capable of modulating these functions prompted us to advance the hypothesis that the degradation of plasma neuropeptides, specifically opioid peptides, may be altered by aging. Objective: To verify the possible existence of age-induced variations in neuropeptide hydrolysis in human plasma, using leucine enkephalin as the model substrate. Methods: The hydrolysis of leucine enkephalin and the formation of its hydrolysis byproducts in the presence of plasma enzymes were studied by kinetic and chromatographic techniques in a group of elderly individuals and a control group. Results: The results obtained indicate that in elderly individuals the activity of enkephalin-degrading plasma enzymes is greater than in controls. ANOVA analysis of these data indicates that the dependency of the variation of hydrolysis upon the 2 age groups is statistically significant. Increased substrate hydrolysis, and a modified hydrolysis pattern, appear to be associated with increased activity of the enzymes involved, and with different distribution of the individual enzymes within each class, as well as with severely reduced activity of the low molecular weight plasma inhibitors. Conclusion: The combination of the above-mentioned factors appears to define a characteristic hydrolysis pattern for elderly individuals which is different from that found in controls.

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Section: Discussionmentioning

confidence: 99%

“…Sheets were cut according to internal standards, and transferred to scintillation vials. Radioactive label was extracted, and samples counted as reported elsewhere [26].…”

Section: Chromatographic Proceduresmentioning

confidence: 99%

Section: Chromatographic Proceduresmentioning

confidence: 99%

Section: Hydrolysis In Whole Plasmamentioning

confidence: 99%

Section: Low Molecular Weight Inhibitorsmentioning

confidence: 99%

See 3 more Smart Citations

Age Increases Leucine Enkephalin Hydrolysis in Human Plasma

Bongiorno

Marzano²,

Marini³

et al. 1998

Gerontology

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Detection of neuropeptides using on-capillary copper complexation and capillary electrophoresis with electrochemical detection

Gawron

Lunte

2000

Electrophoresis

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Capillary electrophoresis with electrochemical detection using a carbon fiber electrode in conjunction with on-capillary copper complexation was evaluated for the determination of peptides in standard and biological matrices. Peptides composed of 2-10 amino acids were investigated. A comparison was made between the responses obtained for peptides containing the oxidizable residue tyrosine and those obtained for their respective copper complexes. Electrochemical detection of non-tyrosine-containing peptides and a cyclic peptide was also demonstrated. A separation of leucine (Leu)-enkephalin and five metabolites was developed and then used for the investigation of Leu-enkephalin metabolism in plasma. The appearance of the des-tyrosine (des-Tyr) Leu-enkephalin, which cannot be detected directly at a carbon electrode, was monitored using the on-capillary complexation technique. Direct injection of the plasma sample was possible using this methodology.

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On‐line coupling of size exclusion and capillary zone electrophoresis via a reversed‐phase C18 trapping column for the analysis of structurally related enkephalins in cerebrospinal fluid

et al. 2003

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On-line coupled analytical techniques can be advantageous in the assay of smaller peptides in complex biological matrices such as plasma, cerebrospinal fluid (CSF) and tissues. The present study shows the feasibility of a recently developed system, consisting of a size-exclusion chromatographic (SEC) separation followed by a trapping procedure on an RP18 microcolumn with subsequent elution of the trapped fraction and separation by capillary zone electrophoresis (CZE) for the quantification of structural-related peptides in biological matrices, as demonstrated for a number of enkephalins in CSF. After SEC separation of the enkephalins from large proteins present in CSF a heart-cut of 200 nuL, containing the enkephalin peak, is taken, concentrated on the RP18 microcolumn and, after elution of the enkephalins with 80% acetonitrile, a fraction of the eluate is electrokinetically injected into the CZE system, where stacking and separation is achieved. The degradation of the peptides, caused by endogenous peptidases in the matrix, is sufficiently inhibited with imipramine HCl. The assay has a satisfactory linearity and intraday (9.70-16.3%) precision considering the complexity of this multidimensional separation system. The sensitivity of the method, with a concentration limit of quantification of 2.5 nug/mL, is comparable with other CZE assays for peptides and sufficient for the quantification of peptide drugs in biological matrices.

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Interindividual Variability of Enkephalin-Degrading Enzymes in Human Plasma

Cited by 15 publications

References 30 publications

Age Increases Leucine Enkephalin Hydrolysis in Human Plasma

Age Increases Leucine Enkephalin Hydrolysis in Human Plasma

Detection of neuropeptides using on-capillary copper complexation and capillary electrophoresis with electrochemical detection

On‐line coupling of size exclusion and capillary zone electrophoresis via a reversed‐phase C18 trapping column for the analysis of structurally related enkephalins in cerebrospinal fluid

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