Short communication: Effects of different whey concentrations on physicochemical characteristics and viable counts of starter bacteria in dairy beverage supplemented with probiotics.

Fermented dairy beverages supplemented with the probiotics Lactobacillus acidophilus and Bifidobacterium lactis containing different concentrations of whey in their formulas (0, 20, 35, 50, 65, and 80%, vol/vol) were processed and checked for pH; proteolysis; levels of glucose, lactose, ethanol, acetic acid, lactic acid, diacetyl, and acetaldehyde; and lactic bacteria and probiotic counts. The results allowed the effect of whey concentration on the dairy beverages to be observed for each of the different parameters analyzed. The degree to which the whey concentration was useful for the microbial cultures, particularly probiotic cultures, appeared to have a limit. In general, dairy beverages processed with different levels of whey in their formulation exhibited good potential as a food matrix for supplementation with probiotic bacteria, with production of characteristic compounds of fermented milk products, such as volatiles and organic acids.

A role for the perlecan protein core in the activation of the keratinocyte growth factor receptor.

Perlecan, a widespread heparan sulphate (HS) proteoglycan, is directly involved in the storing of angiogenic growth factors, mostly members of the fibroblast growth factor (FGF) gene family. We have previously shown that antisense targeting of the perlecan gene causes a reduced growth and responsiveness to FGF7 [also known as keratinocyte growth factor (KGF)] in human cancer cells, and that the perlecan protein core interacts specifically with FGF7. In the present paper, we have investigated human colon carcinoma cells in which the perlecan gene was disrupted by targeted homologous recombination. After screening over 1000 clones, we obtained two clones heterozygous for the null mutation with no detectable perlecan, indicating that the other allele was non-functioning. The perlecan-deficient cells grew more slowly, did not respond to FGF7 with or without the addition of heparin, and were less tumorigenic than control cells. Paradoxically, the perlecan-deficient cells displayed increased FGF7 surface binding. However, the perlecan protein core was required for functional activation of the KGF receptor and downstream signalling. Because heparin could not substitute for perlecan, the HS chains are not critical for FGF7-mediated signalling in this cell system. These results provide the first genetic evidence that the perlecan protein core is a molecular entity implicated in FGF7 binding and activation of its receptor.

Acute bucket-handle tear of the medial meniscus in a golfer.

This is the first case report of an acute bucket-handle tear in an amateur golfer. The injury occurred during the follow-through phase of the golf swing. The rate of incidence of knee injuries in golfers is only 10%. There is a difference in the injury pattern between amateur and professional golfers; amateurs are more likely to sustain injuries related to poor swing technique. Professional golfers, on the other hand, report more injuries related to overuse. Although forces in the knee are relatively minor in this sport, acute injuries can happen when poor technique is used.

Incidence of adverse reactions in HIV patients treated with protease inhibitors: a cohort study. Coordinamento Italiano Studio Allergia e Infezione da HIV (CISAI) Group.

OBJECTIVE: To assess the probability that protease inhibitor (PI) therapy might be discontinued because of adverse events (AE) and to evaluate the incidence rate of adverse reactions during PI treatment. DESIGN: A prospective cohort, multicenter study on HIV-positive patients starting treatment with at least one PI. SETTING: Ten departments of infectious diseases in Northern Italy. PATIENTS: A total of 1207 patients who started PI therapy in September 1997 and were consecutively observed up to April 1999. MAIN OUTCOME MEASURES: Adverse reactions following initiation of PI therapy, and time to therapy discontinuation due to AE. RESULTS: During the study period, 35.9% patients presented adverse reactions of any grade, whereas 9.7% presented at least one serious AE. After 12 months of treatment, the percentage of patients who had interrupted treatment was 36% of ritonavir-treated patients, 14.2% of those treated with indinavir, 13.6% of ritonavir-saquinavir hard gel capsules (HGC)-treated patients, and 8.5% and 2.1%, respectively, for those treated with nelfinavir and saquinavir HGC. Women and patients with hepatitis experienced a significantly greater number of adverse events compared with other categories. Gastrointestinal events were more frequently observed in patients treated with either ritonavir alone or in combination with saquinavir HGC, as well as in patients receiving nelfinavir, although in this group serious events were rare. Here again, neurologic, metabolic, and hepatic toxicity occurred more frequently in ritonavir and ritonavir-saquinavir HGC treated patients. Allergic reactions were more often observed in patients receiving nelfinavir. Indinavir-treated patients presented the highest incidence of renal toxicity. CONCLUSION: Ritonavir is the drug associated with the largest number of reactions, which appear during the first few months of treatment. Saquinavir HGC and nelfinavir are the best tolerated drugs in a clinical setting.

Enhanced macrophage uptake of elastase-modified high-density lipoproteins.

Incubation of human HDL (d = 1.063-1.21 g/ml) with monocyte-derived elastase causes selective proteolysis of apoA-II and apoA-I apolipoproteins. We have found that elastase-digested HDL (ED-HDL) bind to J774-A1 murine macrophages with enhanced affinity and are internalized and degraded at a rate threefold higher than that of native HDL. Unlike oxidized LDL and HDL and proteolytically modified LDL, the uptake of ED-HDL lipoproteins does not affect the cellular lipid biosynthesis nor modify the cell lipid content. The cell surface binding of (125)I-ED-HDL can be competed by native HDL but not by acetylated LDL, consistent with the idea that ED-HDL are recognized by the class B type I scavenger receptor. The liberation of elastase by lipid-engorging macrophages is regarded as an important event during atherogenesis. By enhancing the cellular uptake of HDL this process can lead to a local decrease of antiatherogenic HDL particles.

Overexpression of bamacan/SMC3 causes transformation.

Bamacan can occur in certain cell types as either a secreted proteoglycan assembled into basement membranes or as an intracellular protein known as structural maintenance of chromosome 3 (SMC3). To assess the role of this protein in tumorigenesis, we investigated whether induced overexpression of bamacan/SMC3 could transform normal fibroblasts. We generated a full-length cDNA encoding the entire mouse bamacan/SMC3 and demonstrated appropriate transcription and translation into a 146-kDa protein. All the NIH and Balb/c 3T3 murine fibroblasts overexpressing this bamacan/SMC3 transgene generated foci of transformation and acquired anchorage-independent growth. The increased levels of bamacan/SMC3 expression achieved in the transfected fibroblasts were the same as those detected in a series of spontaneously transformed murine and human colon carcinoma cells. Moreover, a 3-4-fold overexpression of bamacan/SMC3 was detected in approximately 70% of human colon carcinoma specimens from matched pairs (n = 19, p < 0.0002) and in a cohort of intestinal tumors from Apc-deficient Min/+ mice. These results support the concept that deregulated expression of bamacan/SMC3 is involved in cell transformation.

[Causality relationship of drug adverse reactions: experience in the use of HIV-protease inhibitors]. / Relazione di causalità delle reazioni avverse a farmaci: un'esperienza nell'utilizzo degli inhibitori della proteasi di HIV.

PURPOSE: To establish the exact cause and effect relationship between protease inhibitors (PIs) and adverse events. MATERIALS AND METHOD: Prospective, cohort, multicenter study on HIV-positive patients who are beginning treatment with a PI. Causal relationships are evaluated using the RUCAM algorithm. RESULTS: Since the beginning of the study 1207 patients have been enrolled. Average time of observation is 10.7 months. To date, 784 adverse events have been observed, distributed as follows: excluded 3.8%, improbable 18.5%, possible 41.3%, probable 30.1%, and highly probable 6.3%. Saquinavir shows a statistically significant difference in the rate of non-correlated events with respect to other groups. CONCLUSIONS: Over 20% of adverse events during PI treatment are shown to be non-correlated to these drugs. Saquinavir shows the highest rate of non-correlated events.

Complete cDNA cloning, genomic organization, chromosomal assignment, functional characterization of the promoter, and expression of the murine Bamacan gene.

Bamacan is a chondroitin sulfate proteoglycan that abounds in basement membranes. To gain insights into the bamacan gene regulation and transcriptional control, we examined the genomic organization and identified the promoter region of the mouse bamacan gene. Secondary structure analysis of the protein reveals a sequential organization of three globular regions interconnected by two alpha-helix coiled-coils. The N- and the C-terminal ends carry a P-loop and a DA box motif that can act cooperatively to bind ATP. These features as well as the high sequence homology with members of the SMC (structural maintenance of chromosome) protein family led us to conclude that bamacan is a member of this protein family. The gene comprises 31 exons and is driven by a promoter that is highly enriched in GC sequences and lacks TATA and CAAT boxes. The promoter is highly functional in transient cell transfection assays, and step-wise 5' deletions identify a strong enhancer element between -659 and -481 base pairs that includes Jun/Fos proto-oncogene-binding elements. Using backcrossing experiments we mapped the Bam gene to distal chromosome 19, a locus syntenic to human chromosome 10q25. Bamacan is differentially expressed in mouse tissues with the highest levels in testes and brain. Notably, bamacan mRNA levels are low in normal cells and markedly reduced during quiescence but are highly increased when cells resume growth upon serum stimulation. In contrast, in all transformed cells tested, bamacan is constitutively overexpressed, and its levels do not change with cell cycle progression. These results suggest that bamacan is involved in the control of cell growth and transformation.

Foam cell conversion of macrophages alters the biosynthesis of heparan sulfate.

Heparan sulfate is thought to regulate the biological activities of several proteins implicated in the pathogenesis of atherosclerosis. While the interactions of heparan sulfate with lipoprotein lipase and various growth factors have been actively studied, little is known of the cellular regulation of heparan sulfate biosynthesis in response to lipid accumulation. We have investigated heparan sulfate biosynthesis during conversion of murine J774 macrophages into lipid-laden foam cells. Such conversion is shown to accelerate the rate of glycosaminoglycan synthesis and the transport of newly synthesized proteoglycans into the medium. Moreover, the structure of heparan sulfate is specifically altered due to an approximately 30% increase in the 6-O-sulfation of glucosamine residues within the N-sulfated heparan sulfate domains, whereas the sulfation of chondroitin sulfate remains unaffected. These results suggest a selective effect of foam cell conversion on the biosynthesis of heparan sulfate.

Factors influencing the outcome of hyperopic lamellar keratoplasty.

PURPOSE: To determine which factors affect the refractive outcome of hyperopic lamellar keratoplasty. SETTING: Jules Stein Eye Institute and Department of Ophthalmology, UCLA, Los Angeles, California, USA. METHODS: This retrospective study comprised 38 consecutive eyes of 25 patients with naturally occurring hyperopia who had automated lamellar keratoplasty by one surgeon. Mean attempted correction was 3.80 diopters (D) (range of 1.50 to 6.00 D); 34 eyes were followed for 3 months. The effect of applanation lens diameter, keratometry, age, corneal thickness, absolute flap thickness in microns, thickness of the posterior lamellae in microns, and flap thickness as a percentage of corneal thickness were determined using multivariate linear regression. RESULTS: With current nomograms, mean undercorrection 3 months after hyperopic lamellar keratoplasty was 1.26 D +/- 0.91 (SD); 14 of 34 eyes were within +/- 1.00 D of the attempted correction. The best predictive factors of achieved correction were applanation lens diameter and absolute flap thickness in microns, which accounted for 54% of the variability in outcome. Keratometry, corneal thickness, and flap thickness as a percentage of corneal thickness had no significant additional predictive value. CONCLUSION: The refractive outcome of hyperopic lamellar keratoplasty was primarily determined by applanation lens diameter and absolute flap thickness in microns. Current nomograms suggest that flap thickness as a percentage of corneal thickness is a major determinant of effect, but this factor appears unimportant to the refractive effect of hyperopic lamellar keratoplasty. We hypothesize that it is slight swelling of the anterior corneal stroma rather than the bulging of the posterior cornea that causes the hyperopic correction in hyperopic lamellar keratoplasty.

Intrasplenic posttraumatic pseudoaneurysm: echo color doppler diagnosis.

Nonsurgical conservative treatment of blunt splenic trauma has gained widespread consensus in the last few years. It has been demonstrated that 60% of patients with blunt abdominal trauma with spleen lesion achieve the best therapy by using conservative therapy. Despite the accuracy of ultrasonography (US) and computed tomography in detecting and grading the spleen lesions, the evolution of the lesion is often unexpected. In 15 to 30% of patients, a two-stage splenic rupture may be expected within 2 weeks. Delayed complications, such as splenic abscesses and pseudoaneurysms of the splenic artery and its branches, have been observed. To prevent complications, a short follow-up has been scheduled for these patients by using US and US color Doppler. The authors propose routine echo Doppler evaluation for all patients affected by intraparenchymal hematoma after blunt abdominal trauma.

A prospective multicenter trial of excimer laser phototherapeutic keratectomy for corneal vision loss. The Summit Phototherapeutic Keratectomy Study Group.

PURPOSE: The 193-nm argon fluoride excimer laser can remove corneal scars and smooth corneal irregularities, obviating corneal transplantation. We conducted a prospective multicenter trial of excimer laser phototherapeutic keratectomy for corneal vision loss as a basis for Food and Drug Administration premarket approval. METHODS: We treated 232 eyes of 211 patients with corneal vision loss. All had corneal pathology in the anterior 100 microns of the stroma. Mean postoperative follow-up was 10 +/- 8 months. The primary outcome variable was change in best spectacle-corrected visual acuity. RESULTS: At postoperative month 12, best spectacle-corrected visual acuity improved in 46 (45%) of 103 eyes and worsened in nine (9%) of 103 eyes by 2 or more Snellen lines. Best spectacle-corrected visual acuity improved by a mean of 1.6 +/- 2.8 Snellen lines (95% confidence interval, 1.1 to 2.1 lines). Every postoperative visit confirmed statistically significant improvement of mean best spectacle-corrected acuity. At month 12, treated eyes had a mean hyperopic shift in refraction of 0.87 diopter and a mean reduction in astigmatism of 0.36 diopter. Treatment appeared most effective in eyes with hereditary corneal dystrophies, Salzmann's nodular degeneration, and corneal scars, and least effective in eyes with calcific band keratopathy. Complications included recurrence of underlying pathology, corneal graft rejection, and bacterial keratitis. CONCLUSIONS: Argon fluoride excimer laser phototherapeutic keratectomy is effective, with relatively few complications, for treating vision loss from corneal opacification or irregularity. Efficacy, however, varies widely depending upon individual eyes and underlying diagnoses.

Correction of dyslipoproteinaemia of casein-fed rabbit by FCE 27677, a potent novel ACAT inhibitor.

Rabbits fed a wheat starch casein diet develop hypercholesterolaemia characterized by the plasma elevation of low density lipoprotein (LDL) that is caused by oversecretion of apoB-100 containing lipoproteins by the liver and by the suppression of the EDTA-sensitive hepatic beta- very low density lipoprotein (VLDL)-LDL receptor. In this study, the effect of FCE 27677 ((-)N-[2,6-bis(1-methylethyl)phenyl]-N'-[(4R,5R)-2-(4-dimethylaminoph eny l)-4,5-dimethyl-dioxolan-2-yl]methylurea) a novel potent systemic acylCoA:cholesterol acetyltransferase (ACAT, EC 2.3.1.26) inhibitor, has been evaluated. When New Zealand White rabbits were fed with casein for 4 weeks, LDL cholesterol increased from 14 +/- 3 mg/dl-1 to 77 +/- 6 mg/dl-1. By contrast the animals receiving FCE 27677 (10 mg kg-1 d-1) mixed with the casein diet maintained a normal LDL concentration (22 +/- 3 mg dl-1). This hypolipidaemic effect was also observed when rabbits previously made hypercholesterolaemic by being fed casein for 4 weeks were then treated for a month with FCE 27677. [125I]LDL plasma turnover studies and [125I]LDL binding studies to liver membranes were carried out with the purpose of investigating the mechanism of action of the drug. The LDL apoB-100 production rate in chow-fed, casein-fed, and casein-fed rabbits receiving FCE 27677, was respectively 10.5, 22.4, and 12.5 mg kg-1 d-1. The turnover rate of [125I]LDL in the animals receiving the drug was not, however, different from that in the rabbits fed the casein diet alone (2.381 vs 2.079 pools d-1). Both values were lower than that in chow-fed animals (3.271 pools d-1). FCE 27677 did not normalize the activity of the hepatic beta-VLDL-LDL EDTA-sensitive receptor which is suppressed by casein feeding. Altogether the results are consistent with the idea that FCE 27677 by acting through inhibition of the cholesterol esterification in the liver normalizes the LDL synthetic rate. ACAT inhibitors may be useful drugs for the treatment of human dyslipoproteinaemia secondary to derangement of the apoB-100 synthetic rate.

Inhibition of acyl-CoA: cholesterol acyltransferase decreases apolipoprotein B-100-containing lipoprotein secretion from HepG2 cells.

There is evidence that the overproduction of apoB-100-containing lipoproteins by the liver is the underlying event in some forms of dyslipoproteinemia. This metabolic status is associated to an increased risk of developing premature coronary artery disease CAD. The conclusions from previous studies suggested that the availability to the hepatocytes of cholesterol that is readily esterified is an important determinant for VLDL and LDL secretion. In the present study, we set out to investigate the effect of the specific stimulation and inhibition of the rate-limiting enzyme of the cholesterol esterification, acyl-CoA:cholesterol acyltransferase (ACAT, E.C. 2.3.1.26), on the lipid and on the apoB-100 secretion rate from a human hepatoma cell line (HepG2). When the specific ACAT inhibitor FCE 27677 (10-5 M) was added to the cultures, a decrease of the cellular cholesteryl ester content and at the same time a significant reduction of the neutral lipids and of the apoB-100 secretion rate were noticed. The stimulation of ACAT by 25-hydroxycholesterol (20 microgram/ml) caused a 4-fold increase of the cellular cholesteryl ester content and a 2-fold increase of the lipoprotein secretion rate. FCE 27677 (10-5 M to 10-7 M) prevented the effects elicited by the oxysterol. On the contrary, lovastatin (10-6 M) and gemfibrozil (10-6 M) had no effect. The analysis of the lipid and of the apolipoprotein composition of the lipoproteins secreted in the medium revealed that ACAT inhibition had the dual effect of both decreasing the number of apoB-100-containing lipoproteins secreted as well as their cholesteryl ester load. Altogether, these data support the idea of a close relationship between ACAT activation, leading to increased cholesteryl ester availability, and apoB-100-containing lipoprotein secretion. It is speculated that ACAT inhibitors may prove useful for the treatment of human dyslipoproteinemias caused by the hepatic overproduction of apoB-100-containing lipoproteins.

FCE 27677: a novel inhibitor of acyl-CoA: cholesterol acyltransferase with potent oral hypolipidemic activity.

FCE 27677 ([(-)N-[2,6-bis(1-methylethyl)phenyl]-N'-[(4R,5R)-2- (4-dimethylaminophenyl)-4,5 dimethyl-dioxolan-2-yl]methylurea) is a new systemically available ACAT inhibitor belonging to the class of ketalic disubstituted ureas. When tested in microsomes from rabbit intestine, aorta and liver, it inhibited the enzyme with IC50 of 9.31, 6.99 and 92.2 nM, respectively. It had no effect on plasma LCAT and intestinal cytosolic cholesterol esterases and, when tested in a tissue culture system, it did not interfere with the synthesis of cholesterol, triglycerides, and phospholipids. Enzyme inhibition kinetics indicated that FCE 27677 is a non-competitive inhibitor of the enzyme with respect to acylCoA and to cholesterol. When administered mixed to a 1.5% cholesterol and 0.5% sodium cholate-enriched diet to rats, it prevented the development of hypercholesterolemia with ED50 of 0.35 mg kg-1 day-1. Given in a single oral dose to hypercholesterolemic rats it significantly reduced both the plasma lipid levels and the hepatic cholesteryl ester content within 6 h from gavage. VLDL and LDL levels and composition were also significantly affected. Similar effects were observed when the drug was given mixed to a regular chow diet for 4 weeks to hypercholesterolemic rabbits. These results are consistent with the idea that systemically available ACAT inhibitors can affect the composition and the metabolism of the atherogenic cholesteryl ester-rich VLDL and LDL. ACAT inhibitors appear promising for the correction of dyslipoproteinemias secondary to lipoprotein overproduction, and in reducing the atherogenic index of apoB-100 containing lipoproteins.

Oxidized lipoproteins induce long-lasting inhibition of nitric oxide synthase from a murine endothelioma cell line (bEnd.4).

BACKGROUND: The vascular endothelium produces nitric oxide, which has vasodilatory properties. It has been postulated that some lipoproteins may increase arterial vascular tone by decreasing the availability of endothelium-derived nitric oxide. The mechanism underlying this effect, however, is still poorly understood. METHODS: We investigated the effect of native and oxidized human low- and high-density lipoproteins on the nitric oxide synthetic activity of an endothelioma cell line (bEnd.4). Oxidized lipoproteins were obtained by incubation with CuSO4. The production of nitric oxide by the cells was monitored by quantifying the nitrite concentration in the medium using Greiss reagent. RESULTS: The synthesis of nitric oxide by the bEnd.4 cell line was calcium-dependent and was abolished by a selective inhibitor of the constitutive nitric oxide synthase. Incubation with oxidized lipoproteins caused a time- and dose-dependent inhibition of nitric oxide synthetic activity. At a concentration of 100 micrograms/ml cholesterol, oxidized low- and high-density lipoproteins inhibited the production of nitric oxide by 27 and 51%, respectively, within 6h. The lipid fraction obtained from the native or the oxidized lipoproteins mimicked the effect of the intact lipoproteins. CONCLUSION: These results support the involvement of oxidized lipoproteins in the modulation of endothelial functions relevant to the pathogenesis of cardiovascular disease.

Regulatory function of glucose and insulin on high-density lipoprotein cholesterol in normolipidemic subjects.

A decreased plasma concentration of high-density lipoprotein (HDL) cholesterol is associated with a higher incidence of coronary artery disease in populations. Therefore, there is intense investigation into the mechanisms responsible for the regulation of HDL cholesterol concentration in plasma. Insulin has a potent effect on HDL cholesterol, but it is unclear whether this is mediated by the primary effect insulin has on plasma triglycerides (TG). In this study, the question of the relationship between glucose, insulin, and HDL cholesterol has been addressed by investigating a cohort of nondiabetic normolipidemic men living in the Venice, Italy, area. One hundred twenty-eight men aged 30 to 69 years were initially recruited. The following parameters were measured: fasting plasma cholesterol, TG, HDL cholesterol, glucose, and insulin. One hundred seventeen of these subjects underwent an oral glucose tolerance test (OGTT), and the glucose and insulin responses were assessed. The final statistical analysis was performed on 98 nondiabetic individuals with plasma lipid levels within the 75th percentile for cholesterol and TG concentrations of the general population of the same age. The insulin response was a positive independent variable for plasma TG (P < .005) and HDL cholesterol (P < .005). On the other hand, HDL cholesterol was negatively associated with plasma TG. This relationship remained significant (P < .0001) also after controlling for age, body mass index (BMI), and glucose- and insulin-related measurements. Consistent with these results, both a stepwise variable selection analysis and a stratification analysis of the data indicated that the plasma TG concentration is the major determinant of HDL cholesterol level.(ABSTRACT TRUNCATED AT 250 WORDS)

The role of ultrasonography in blunt abdominal trauma: results in 250 consecutive cases.

The accuracy of ultrasonography (US) in detecting abdominal lesions and free fluid collections in patients with blunt abdominal trauma was evaluated in 250 patients. Particular attention was paid to the role of associated US-guided paracentesis in doubtful cases and in those referred for nonsurgical therapy. The overall sensitivity of US in detecting free fluid collection was 98% (51 of 52 cases) with a specificity of 99% and a positive predictive value of 100%. The overall sensitivity was 93% in spleen injuries, 80% in liver injuries, and 100% in kidney lesions with a positive predictive value of 93%, 100%, and 100%, and a specificity of 99%, 100%, and 100%, respectively. Three stable patients underwent celiotomy on the basis of the results of US-guided paracentesis. The versatility, sensitivity and, repeatability of US, along with its feasibility at bedside and the possibility of performing a guided paracentesis represent the main characteristics that make US the first diagnostic approach to patients with blunt abdominal trauma.

Imidazolylbenzopyrane derivatives: a new class of acyl-CoA: cholesterol acyltransferase (ACAT) inhibitors.

Inhibitors of the enzyme Acyl-CoA: Cholesterol Acyltransferase are regarded as potentially useful agents in the treatment of hypercholesterolemia and atherosclerosis. We report here a novel series of 2, 6-disubstituted-3-imidazolylbenzopyrane derivatives with significant in vitro ACAT inhibitory activity (IC50 range 0.05-0.5 microM). Compounds of this series such as 26 are examples of a new, structurally distinct class of potent ACAT inhibitors with high specificity for the aortic subtype of the enzyme. The structure-activity relationships of the 3-imidazolylbenzopyrane ACAT inhibitors were investigated by systematic manipulation of two regions of the parent compound 1 and the inhibitory activity resulted linked to the substituent in position 6 of the benzopyrane ring and modulated by the size of lipophilic substituents in position 2. Investigation of the mechanism of the inhibitory effect leads to the conclusion that these compounds act in a non-competitive fashion.