Prevalence of oedema of the lower limbs in multiple sclerosis patients: a vascular and lymphoscintigraphic study.

The aim of the study was to evaluate the frequency of oedema of the lower limbs in multiple sclerosis (MS) patients utilizing a multidisciplinary approach. A total of 205 patients with definite MS were included in the study. Seventy-five were male and 130 female, with a mean age of 50.53, mean Expanded Disability Status Scale (EDSS) score of 5.27 and mean disease duration of 16.6 years. Seventy-one patients had a relapsing-remitting (RR) disease course, 85 were secondary progressive (SP) and 49 were primary progressive (PP). Ninety-three patients (45%) showed oedema at the examination. EDSS, disease duration and disease course, but not gender, were statistically different between oedema and non-oedema patients. Out of 93 patients with oedema, 69 agreed to undergo a vascular examination. Of 69 patients, 45 (65.2%) had a CEAP score (specific rating scale for oedema) of 3 (presence of oedema) and 24 (34.8%) had a score of 4 (presence of a trophic disorder). Out of 69 subjects, 33 agreed to undergo a lymphoscintigraphy, which was normal in only 29 extremities out of 66. Lower limb oedema is common in MS patients, especially in those with reduced mobility. Early screening is advised in patients with an elevated EDSS.

Biochemical aspects and functional role of the copper-containing amine oxidases.

The copper-containing amine oxidases of the class EC 1.4.3.6 share many biochemical similarities. They contain cupric copper and catalyse the same general reaction. In mammals, diamine oxidase has a role in the metabolism of histamine, some other diamines and spermine oxidase, involved in the metabolism of polyamines. However, the functional role of benzylamine oxidase (BAO) and the tissue semicarbazide-sensitive amine oxidases (SSAO) is still under investigation. Circulating BAO is derived from the tissue SSAO. It has a high affinity for benzylamine and its role in the extra-cellular matrix includes the maturation of pro-elastin and control of the inflammatory process.

Technical issues and pathologic implications of sentinel lymph node biopsy in early-stage breast cancer patients.

BACKGROUND AND OBJECTIVES: Recent studies have demonstrated that the sentinel lymph node (sN) can be considered a reliable predictor of axillary lymph node status in breast cancer patients. However, some important issues, such as optimization of the technique for the intraoperative identification of the sN, and the clinical implications of sN metastasis as regards the surgical management of the axilla still require further elucidation. The objectives of this study was to assess (1) the feasibility of sN identification with a combined approach (vital blue dye lymphatic mapping and radioguided surgery, RGS) and the specific contribution of either techniques to the detection of the sN, and (2) the correlation between the size of sN metastasis (micrometastasis < or = 2 mm; macrometastasis > 2), primary tumour size, and the status of nonsentinel nodes (nsN) in the axilla. METHODS: Between October of 1997 and December of 1999, 212 patients with breast cancer (average age: 61 years; range, 40-79 years) underwent sN biopsy before performing standard axillary dissection. In a subset of 153 patients, both vital blue dye (Patent Blue-V) lymphatic mapping and RGS were used to identify the sN, and the relative contribution of each of the two techniques was assessed. RESULTS: Overall, the sN was identified in 206 of 212 patients (97.1%); at histologic examination of all dissected nodes, 77 of 206 patients had positive nodes (37.3%). The false-negative rate was 6.5% (5/77), the negative predictive value was 96.3% (129/134), and accuracy was 97.6% (201/206). Among 72 patients with positive sN, micrometastases were detected in 21 cases and macrometastases in 51. When micrometastases only were observed, the sN was the exclusive site of nodal metastasis in 17 of 21 cases (80.9%); in the remaining 4 cases (19.1%), nsN metastases were detected in 3 of 14 pT1c patients (21.5%), and 1 of 5 pT2 patients (20%). Macrometastases were detected in patients with tumors classified as pT1b or larger: the sN was the exclusive site of metastasis in 3 of 4 pT1b patients (75%), in 14 of 29 pT1c patients (48.2%), and in 3 of 18 pT2 patients (16.6%). The specific contribution of the two different techniques used in the identification of the sN was evaluated; the detection rate was 73.8% (113 of 153) with Patent Blue-V alone, 94.1% (144 of 153) with RGS alone, and 98.7% (151 of 153) with Patent Blue-V combined with RGS (P < 0.001). Noteworthy, whenever the sN was identified, the prediction of axillary lymph node status was remarkably similar (93-95% sensitivity; 100% specificity; 95-97% negative predictive value, and 97-98% accuracy) with each of the three procedures (Patent Blue-V alone, RGS alone, or combined Patent Blue-V and RGS). CONCLUSIONS: Sentinel lymphadenectomy can better be accomplished when both procedures (lymphatic mapping with vital blue dye and RGS) are used, due to the significantly higher sN detection rate, although the prediction of axillary lymph node status remains remarkably similar with each one of the methods assessed. That patients with small tumours (<1 cm) and sN micrometastasis are very unlikely to harbour metastasis in nsN should be considered when planning randomised clinical trials aimed at defining the effectiveness of sN guided-axillary dissection.

Regional cerebral blood flow and prognostic evaluation in Alzheimer's disease.

The present investigation reports the application of regional cerebral blood flow (rCBF; (133)Xe method) to prognostic purposes in a consecutive series of 76 patients (mean age 68.4 +/- 8.7 years) with probable Alzheimer's disease (AD; NINCDS-ADRDA criteria). The likelihood that rCBF from a posterior temporal-inferior parietal area in each hemisphere at the first visit may predict timing of achievement of three endpoints (i.e. loss of activity of daily living, ADL, incontinence and death due to end-stage AD) was tested by the 'lifereg' procedure of the Statistical Analysis System package. With respect to baseline evaluation, 32 patients lost ADL 20.6 +/- 17.4 months later, 31 developed incontinence 27.1 +/- 19.0 months later, and 16 patients died after 40.9 +/- 23.8 months of follow-up. Baseline rCBF significantly predicted all end-points: the loss of ADL (left hemisphere: p = 0.04; right hemisphere: p = 0.02), incontinence (p = 0.02 in both hemispheres) and death (p = 0.01 in both hemispheres). Statistical significance was maintained for the loss of ADL and incontinence both in a subgroup of mildly demented patients, in whom death was not considered due to the low number of patients who died, and in a multivariate analysis including patient age, age at onset, sex, duration of illness, Mini-Mental State Examination score and presence of extrapyramidal signs and psychotic symptoms at the first visit. This study shows that rCBF measurement in a posterior temporal-inferior parietal area may give prognostic information on timing of evolution of AD, whenever performed during the course of the disease, and may be utilized both in clinical practice and for social planning.

Localization of the sentinel lymph node in breast cancer by combined lymphoscintigraphy, blue dye and intraoperative gamma probe.

Axillary lymph node status represents the most important prognostic factor in patients with operable breast cancer. A severe limitation of this technique is the relatively high rate of false negative sentinel lymph nodes (>5%). We studied 284 patients suffering from breast cancer; 264 had T1 tumors (16 T1a, 37 T1b and 211 T1c), while 20 had T2 tumors. All patients underwent lymphoscintigraphy 18-h before surgery. At surgery, 0.5 mL of patent blue violet was injected subdermally, and the sentinel lymph node (SN) was searched by gamma probe and by the dye method. The surgically isolated SN was processed for intraoperative and delayed examinations. The SN was successfully identified by the combined radioisotopic procedure and patent blue dye technique in 278/284 cases (97.9%). Analysis of the predictive value of the SN in relation to the status of the axillary lymph nodes was limited to 191 patients undergoing standard axillary dissection irrespective of the SN status. Overall, 63/191 (33%) identified SNs were metastatic, the SN alone being involved in 37/63 (58.7%) patients; a positive axillary status with negative SN was found in 10/73 (13.7%) patients with metastatic involvement. In T1a-T1b patients the SN turned out to be metastatic in 9/53 patients (17.0%). In 7/9 patients the SN was the only site of metastasis, while in 2/9 patients other axillary lymph nodes were found to be metastatic in addition to the SN. None of the 44 patients in whom the SN proved to be non-metastatic showed any metastatic involvement of other axillary lymph nodes. Our results demonstrate a good predictive value of SN biopsy in patients with breast cancer; the predictive value was excellent in those subjects with nodules smaller than 1 cm.

Different sites and modes of tracer injection for mapping the sentinel lymph node in patients with breast cancer.

Several studies have been published describing the techniques of identification of the "sentinel lymph node" (SN). There are marked differences in the techniques used by different investigators. Although agreement exists about the tracer particle size and the volume of injection, it is unknown what is the best site where to inject the tracer or the vital dye. The aim of the present study was to define the influence of different sites of injection on imaging of the lymphatic ducts and their SNs. We performed a pilot study in 30 consecutive patients with breast cancer who underwent SN biopsy by means of radioguided surgery and vital blue dye mapping. The patients were divided into six groups of five patients each; each patient was given a subdermal (ID) or peritumoral (IP) injection of radiotracer (300 microCi in 150 mL of 99mTc-HSA microcolloids; Albures, Amersham Sorin) above the tumor site in order to localize the SN. After the identification of the SN, a second injection of radiotracer was performed, which was different in each patient subset. In some cases more than one lymph node appeared on the lymphoscintigraphic scans after the second injection of radiotracer. When the peritumoral route was used it took longer to visualize the lymphatic pathways. For the ID route, injection at the exact skin projection over the tumor is optimal. Internal mammary lymph nodes were identified by both IP (2) and ID (1) injection, irrespective of the quadrant in which the tracer was injected. Our findings support the hypothesis of a precise topographic correspondence between the primary tumor and its specific SN. The subdermal route is more accurate than the intraparenchymal route, as it allows faster identification of the lymphatic vessels and SN. We believe these observations should be taken into account for the proper selection of the injection site of either vital dye or radiopharmaceuticals.

Mapping the sentinel lymph node in malignant melanoma by blue dye, lymphoscintigraphy and intraoperative gamma probe.

Eighty-eight consecutive patients (48 men and 40 women; mean age, 58.9 years; range, 16-84 years) with clinically localized cutaneous melanoma involving the trunk, extremities or head and neck underwent lymphatic mapping at our institution. The primary melanoma had a mean thickness of 2.74 mm (range, 0.95 to 9 mm). Patients were divided into two groups: group A (39 patients) underwent only vital blue dye (VBD) mapping, while group B (49 patients) underwent lymphatic mapping with VBD and radio-guided surgery (RGS) combined. In all patients 1-1.5 mL of VBD was injected subdermally around the biopsy scar 10-20 min before surgery. In group B 37 MBq in 150 microL of 99mTc-HSA nanocolloid was additionally injected intradermally 18 h before surgery (3-6 aliquots injected perilesionally). In all lymphatic basins where drainage was noted the sentinel lymph nodes (SNs) were identified and marked with a cutaneous marker. Final identification of the SN was then performed externally by a hand-held gamma probe. After the induction of anesthesia 0.5-1-0 mL of patent blue V dye was injected intradermally with a 25-gauge needle around the site of the primary melanoma. SNs were examined by routine hematoxylin and eosin (H&E) staining and immunohistochemistry. Patients with histologically positive SN(s) underwent standard lymph node dissection (SLND) in the involved lymph node basin. The SN was identified in 37/39 patients (94.9%) of group A and in 48/49 patients (98.0%) of group B. Blue dye mapping failed to identify the SN in 5 of the 88 patients (5.8%), while the radioisotope method failed in only 1 of 49 patients (2.0%). Similar results were obtained with the combined use of the two probes. The average number of SNs harvested was 1.9 per basin sampled, which does not differ significantly from the numbers reported by other authors. The SN was histologically positive in 18 patients (20.5%). None of the 12 patients with a Breslow thickness less than 1.5 mm had positive SNs, whereas 18 of the 77 patients (23.4%) with a Breslow index exceeding 1.5 mm showed metastatic SNs with H&E or immunohistochemistry. The latter all underwent SLND of the affected basin. In 10 patients (55.6%) the SN was the only site of tumor invasion; eight patients (44.4%) with positive SNs had one or more metastatic lymph nodes in the draining basin.

Semicarbazide-sensitive amine oxidases in heart and bovine serum.

In guinea pig dorsal skin the semicarbazide-sensitive amine oxidase (SSAO) is localised in fibroblasts. Fibroblasts in culture lose the ability to express this enzymatic activity with doublings, thus suggesting that the SSAO expression needs some factors which are not present in the 10% bovine serum culture medium. Fresh bovine serum of adult animals contains two SSAO activities, one with high affinity for benzylamine and one with lower affinity. The enzyme with lower affinity for benzylamine was identified as spermine oxidase, the oxidation of [14C]-benzylamine was inhibited by semicarbazide, alpha-aminoguanidine and B24, a specific inhibitor of benzylamine oxidase and spermine oxidase, both SSAO enzymes. The enzymatic activity of bovine serum was partially purified, the kinetic properties and sensitivity to inhibitors studied. A mathematical procedure for the analysis of the kinetics resulting from the activity of two enzymes acting on the same substrate seems to give better results than the methods previously described.

Mapping sentinel lymph node in breast cancer by combined lymphoscintigraphy, blue-dye, and intraoperative gamma-probe.

The purpose of the present work was two-fold: 1) to evaluate the predictive value of the sentinel lymph node (sLN) versus the axillary-node status in patients with T1-T2 breast cancer, and 2) to form an experimental basis for a randomized trial in which one group of patients with non-metastatic sLN will not have axillary dissection. Of a group of 284 patients considered for this analysis, 264 had a T1 cancer (16 T1a, 37 T1b and 211 T1c), while 20 had a T2 cancer; 243 patients were in clinical stage N0 and 41 were N1. All patients underwent lymphoscintigraphy 18 hr before surgery: 10 MBq in 0.15 mL of 99mTc-human albumin nanocolloids (particle size between 50-80 nm) was injected subdermally at the cutaneous projection of the tumor. Static gamma-camera images were acquired every 10-15 minutes until scintigraphic identification of the sLN. At surgery, 1-2 mL of Patent-Blue Violet was injected subdermally, and the sLN was searched by gamma-probe and by the dye method. The surgically isolated sLN was processed for intraoperative Hematoxylin & Eosin (H&E) histology, then for delayed histological and immunohistochemical examinations. The sLN was successfully identified by the combined radioisotopic procedure and Patent-Blue dye technique in 278/284 cases (97.9%). The Patent-Blue dye technique alone identified fewer sLNs than the radioisotopic procedure alone (56.3% versus 97.2%). Analysis of the predictive value of the sLN as to the status of axillary lymph nodes was limited to 197 patients undergoing standard axillary dissection irrespective of the sLN status. Overall, 63/191 (33%) identified sLNs were metastatic, the sLN alone being involved in 37/63 (58.7%) patients; a positive axilla status with negative sLN was found in 10/73 patients with metastatic involvement (13.7% false-negative rate). In conclusion, subdermal lymphoscintigraphy was confirmed to be an effective technique for sLN mapping; the addition of Patent-Blue dye minimally improved intra-surgical identification of the sLN. There was a high, but not absolute, correlation between a negative sLN and a negative axilla.

Pattern of lymphatic drainage to the sentinel lymph node in breast cancer patients.

BACKGROUND AND OBJECTIVE: We performed a pilot study on 30 consecutive patients undergoing sentinel node (sN) biopsy by radioguided surgery and vital blue dye mapping to determine whether there is a single sN for each breast independent of tumor site or an sN specifically related to the site of the breast neoplasm. METHODS: There were 6 groups of 5 patients; each patient had a subdermal injection of radiotracer on the tumor site plus a second injection of radiotracer that was changed in every subset of patients to test whether modifying the site or the route of injection could have impaired the proper detection of the sN. RESULTS: "False" sN were detected only in patients who had a second injection of radiotracer away from the tumor site; this occurred in 2 of 5 patients (40%) in group I, in 3 of 5 patients (60%) in group II, in all patients in group III, and in 3 of 5 patients (60%) in group IV. The different route of injection (peritumoral or subdermal) always on the tumor site that was tested in groups V and VI did not impair the proper detection of the sN. CONCLUSIONS: Our findings support the hypothesis of a precise topographic correspondence between the primary tumor and its specific sN more than the existence of a first sN in the axillary basin, which indiscriminately drains all quadrants of the breast, like "a neck of a bottle." This should be considered for the proper selection of the injection site of either vital blue dye or radiopharmaceuticals.

Purification of pig heart benzylamine oxidase.

A benzylamine oxidase (E.C. 1.4.3.6) has been purified from pig heart. Western blot analysis showed that the enzyme cross-reacts with a polyclonal antibody raised against homogeneous, crystalline pig plasma benzylamine oxidase (BAO). A subunit molecular mass of 97 kDa obtained by SDS electrophoresis is identical to the plasma enzyme. The purification procedure consisted of sequential DEAE-cellulose, DEAE-Sephadex, Con A-Sepharose, Sephadex G 200 and hydroxyapatite columns. The specific activity of the purified enzyme was 0.037 µmol min-1mg-1 at 37ûC and the Km for benzylamine was estimated to be 29 µM. The enzyme was inhibited by carbonyl reagents such as semicarbazide and á-aminoguanidine. Phenylhydrazine reacts mole to mole with the enzyme giving a peak at 425 nm. The copper content was 2 g-atom/mole of enzyme.

Sentinel lymph node mapping opens a new perspective in the surgical management of early-stage breast cancer: a combined approach with vital blue dye lymphatic mapping and radioguided surgery.

Recent studies have demonstrated the possibility of identifying the sentinel lymph node (sN) as a reliable predictor of axillary lymph node status in both cutaneous melanoma and breast cancer. However, some important issues need further definition: (1) optimization of the technique for intraoperative detection of the sN; (2) predictive value of the sN as regards axillary lymph node status, and (3) reliability of intraoperative histology of the sN. We report our experience in sN mapping in patients with Stage I-II breast cancer, with the aim of assessing: (1) the feasibility of lymphatic mapping with a combined approach (vital blue dye lymphatic mapping and radioguided surgery); (2) the agreement of the intraoperative histologic examination of the sN, by means of hematoxylin and eosin staining with final histology, and (3) the accuracy of sN histology as a predictor of axillary lymph node status.

The mechanism of inhibition of benzylamine oxidase by 3,5-diethoxy-4-aminomethylpyridine (B24).

B24, 3,5-diethoxy-4-aminomethylpyridine, is a specific inhibitor of the semicarbazide-sensitive amine oxidase with high affinity for benzylamine (BnNH2.SSAO). It is a site-directed inhibitor of pig plasma benzylamine oxidase (BAO) with an affinity for the enzyme much higher than that for benzylamine. B24 inhibition is dependent on the molar ratio B24/BAO because the inhibitor reacts mole to mole with the enzyme and benzylamine appears to be ineffective in removing the inhibitor from the adduct [EI]. B24 is a weak substrate of BAO and for this reason the degree of inhibition (when the molar ratio B24/BAO is lower than 1) decreases with the incubation time as well as with the preincubation time. This decrease is dependent on the gradual release of free enzyme which reacts with the substrate, giving [ES] without any interfering free B24. When the B24/BAO molar ratio is higher than 1, the free enzyme released by the oxidative deamination of B24 reacts with the substrate, but the free B24 present competitively inhibits the formation of [ES] and the affinity of benzylamine is therefore reduced. This is the reason why B24, in the kinetic experiments in which the inhibitor is not pre-incubated with the enzyme, may appear to be a competitive inhibitor or a mixed inhibitor, mainly competitive. When B24 is preincubated with the enzyme and the initial rate of benzylamine oxidation is measured, it appears as a non-competitive inhibitor becoming a mixed one only when the B24/BAO molar ratio is high and the incubation time is long.

Semicarbazide-sensitive amine oxidase activity in the human heart.

A semicarbazide-sensitive amine oxidase with affinity for benzylamine (Bz.SSAO) was found in the human heart. This enzymatic activity has a K(m) of 278 +/- 35.3 microM and a V(m) of 114.7 +/- 14.7 nmol mg-1 min-1 (mean +/- SE of eight hearts) for benzylamine and is strongly inhibited by 1 mM histamine and by B24, a specific inhibitor of Bz.SSAO. No diamine oxidase activity was found in the human heart. The levels of MAo and B were assayed: MAO A showed a K(m) of 137.1 +/- 16.2 microM and a V(m) of 10.4 +/- 2.5 nmol mg-1 min-1 for serotonin; MAo B had a K(m) of 9.9 +/- 1.6 microM and V(m) of 4.3 +/- 1.1 nmol mg-1 min-1 for beta-phenylethylamine (mean +/- SE of seven hearts). The human heart has high MAO B activity and Bz.SSAO with histaminase activity.

Semicarbazide-sensitive amine oxidase in pig heart.

A semicarbazide-sensitive amine oxidase (SSAO) (E.C.1.4.3.6) has been purified from pig heart. Western blot analysis showed that the enzyme reacts with a polyclonal antibody raised against homogeneous crystalline pig plasma benzylamine oxidase (BAO). A subunit molecular mass of 97 KDa obtained by SDS electrophoresis is identical to the plasma enzyme. The purification procedure consisted of sequential DEAE cellulose, octyl-Sepharose, Con A-Sepharose and hydroxyapatite columns. Two peaks of activity were obtained on octyl-Sepharose which were found to be kinetically and immunologically indistinguishable. The specific activity of the purified enzyme was 0.045 mumol/min/mg of protein at 37 degrees C and the Km for benzylamine was estimated to be 63 microM. The enzyme was inhibited by carbonyl reagents such as semicarbazide but was insensitive to the effect of pargyline.

Monoamine oxidase and semicarbazide-sensitive amine oxidase activities in isolated cardiomyocytes of spontaneously hypertensive rats.

In the isolated cardiomyocytes of spontaneously hypertensive rats (SHR, 3 months old) MAO A and B activities were significantly increased compared to the myocytes in the hearts of age-matched Wistar-Kyoto rats. This increase was not associated with cardiac hypertrophy in these young animals, but might represent an early event in the development of hypertrophy. A semicarbazide-sensitive amine oxidase (SSAO) activity was found in cardiomyocytes. This activity showed a high affinity for benzylamine (Km 5-6 microM) and was not inhibited by 10(-4) M pargyline and 10(-5) M deprenyl, but was largely inhibited by 10(-4) M B24 (3,5-diethoxy-4-aminomethylpyridine), a specific inhibitor of semicarbazide-sensitive amino oxidase with high affinity for benzylamine. The SSAO enzyme of rat cardiomyocytes is a copper-amine oxidase and has a crossreactivity with the antibodies raised against pure pig plasma benzylamine oxidase. In the cardiomyocytes of 3-month old SHR rats the level of this enzymic activity is not significantly increased.

Evidence of semicarbazide-sensitive amine oxidase activity in guinea pig tissues.

Lung, heart tissues and blood plasma of guinea pigs were investigated to see if tissue-bound semicarbazide-sensitive amine oxidase activities with a high affinity for benzylamine (Bz.SSAO) were present in this species as well as in others. This paper shows that these enzymic activities are present in guinea pig lung and heart where they are mainly localized in the cytosol and in microsomal fraction. These activities have a high affinity for benzylamine and appear unable to oxidize histamine at an appreciable rate in agreement with the observation that the purified Bz.SSAO of guinea pig skin shows weak histaminase activity. These guinea pig Bz.SSAO activities show some homology with the pure pig plasma benzylamine oxidase. They crossreact with the antibodies raised in the rabbit against the pig plasma enzyme. Benzylamine oxidase activity was also found in guinea pig blood plasma.

Effect of tripeptide-copper complexes on the process of skin wound healing and on cultured fibroblasts.

The effects of Gly-His-Lys-Cu and of three synthetic analogues (I, II and III) on wound healing of the guinea-pig dorsal skin, as well as on cultured fibroblasts, were examined. Gly-His-Lys-Cu and peptide I-Cu were tested in vivo. Hydroxyproline, proteins, DNA and semicarbazide-sensitive amine oxidase, with a high affinity for benzylamine, were measured, and the histology of the wounds was observed after staining with hematoxylin/eosin. Another set of wounds was treated in parallel with equivalent amounts of copper acetate. Gly-His-Lys-Cu and the analogues caused a decrease of the activity of semicarbazide-sensitive amine oxidase, with a high affinity for benzylamine, 4-8 days after surgery, followed by an increase on day 11 that was higher than in the control group. No significant difference was found between the two peptides. A slower reorganization of the skin and a delayed activation of fibroblasts are the main effects observed with these peptides-Cu complexes. Preliminary studies on cultured fibroblasts were monitored to see whether these peptides had a direct effect on fibroblasts. The products studied at a concentration of 10(-7) M, decreased cell reproduction and increased collagen expression.

Identification by gas chromatography-mass spectrometry of an adduct between pure pig plasma benzylamine oxidase and the inhibitor 3,5-diethoxy-4-aminomethylpyridine.

3,5-Diethoxy-4-aminomethylpyridine (B24) interacts with pure pig plasma benzylamine oxidase (BAO), giving a Schiff base with the carbonyl active site. This Schiff base was reduced, isolated by chemical hydrolysis of the enzyme, purified by HPLC and identified by gas chromatography-mass spectrometry (GC-MS) after derivatization. The isolated B24 adduct had the same absorption spectrum, retention time on HPLC and GC and the same mass spectrum as B24-pyridoxamine. B24, which is a reversible enzyme inhibitor, is also a weak substrate and competes with benzylamine, which is the best substrate, for the active site. These results further indicate the presence of pyridoxal-phosphate covalently linked to the pig plasma benzylamine oxidase and involved in the active site of this enzyme.