Probing the binding sites of resveratrol, genistein, and curcumin with milk ß-lactoglobulin.

We determined the binding sites of curcumin (cur), resveratrol (res), and genistein (gen) with milk ß-lactoglobulin (ß-LG) at physiological conditions. Fourier transform infrared spectroscopy, circular dichroism, and fluorescence spectroscopic methods as well as molecular modeling were used to determine the binding of polyphenol-protein complexes. Structural analysis showed that polyphenols bind ß-LG via both hydrophilic and hydrophobic contacts with overall binding constants of Kcurcumin-ß-LG = 4.4 (± .4) × 104 M⁻¹, Kresveratrol-ß-LG = 4.2 (± .2) × 104 M⁻¹, and Kgenistein-ß-LG = 1.2 (± .2) × 104 M⁻¹. The number of polyphenol molecules bound per protein (n) was 1 (cur), 1.1 (res), and 1 (gen). Molecular modeling showed the participation of several amino acid residues in polyphenol-protein complexation with the free binding energy of -12.67 (curcumin-ß-LG), -12.60 (resveratrol-ß-LG), and -10.68 kcal/mol (genistein-ß-LG). The order of binding was cur > res > gen. Alteration of the protein conformation was observed in the presence of polyphenol with a major reduction of ß-sheet and an increase in turn structure, causing a partial protein structural destabilization. ß-LG might act as a carrier to transport polyphenol in vitro.

Locating the binding sites of retinol and retinoic acid with milk ß-lactoglobulin.

ß-lactoglobulin (ß-LG) is a member of lipocalin superfamily of transporters for small hydrophobic molecules such as retinoids. We located the binding sites of retinol and retinoic acid on ß-LG in aqueous solution at physiological conditions, using FTIR, CD, fluorescence spectroscopic methods, and molecular modeling. The retinoid-binding sites and the binding constants as well as the effect of retinol and retinoic acid complexation on protein stability and secondary structure were determined. Structural analysis showed that retinoids bind strongly to ß-LG via both hydrophilic and hydrophobic contacts with overall binding constants of K (retinol-) (ß) (-LG )= 6.4 (± .6) × 10(6) M(-1) and K (retinoic acid-) (ß) (-LG )= 3.3 (± .5) × 10(6) M(-1). The number of retinoid molecules bound per protein (n) is 1.1 (± .2) for retinol and 1.5 (± .3) for retinoic acid. Molecular modeling showed the participation of several amino acids in the retinoid-protein complexes with the free binding energy of -8.11 kcal/mol for retinol and -7.62 kcal/mol for retinoic acid. Protein conformation was altered with reduction of ß-sheet from 59 (free protein) to 52-51% and a major increase in turn structure from 13 (free protein) to 24-22%, in the retinoid-ß-LG complexes, indicating a partial protein destabilization.

Milk ß-lactoglobulin complexes with tea polyphenols.

The effect of milk on the antioxidant capacity of tea polyphenols is not fully understood. The complexation of tea polyphenols with milk proteins can alter the antioxidant activity of tea compounds and the protein secondary structure. This study was designed to examine the interaction of ß-lactogolobulin (ß-LG) with tea polyphenols (+)-catechin (C), (-)-epicatechin (EC), (-)-epicatechin gallate (ECG) and (-)-epigallocatechin gallate (EGCG) at molecular level, using FTIR, CD and fluorescence spectroscopic methods as well as molecular modelling. The polyphenol binding mode, the binding constant and the effects of polyphenol complexation on ß-LG stability and secondary structure were determined. Structural analysis showed that polyphenols bind ß-LG via both hydrophilic and hydrophobic interactions with overall binding constants of KC-ß-LG=2.2 (±0.8)×10(3)M(-1), KEC-ß-LG=3.2 (±1)×10(3)M(-1), KECG-ß-LG=1.1 (±0.6)×10(4)M(-1) and KEGCG-ß-LG=1.3 (±0.8)×10(4)M(-1). The number of polyphenols bound per protein molecule (n) was 1.1 (C), 0.9 (EC), 0.9 (ECG) and 1.3 (EGCG). Molecular modelling showed the participation of several amino acid residues in polyphenol-protein complexation with extended H-bonding network. The ß-LG conformation was altered in the presence of polyphenols with an increase in ß-sheet and α-helix suggesting protein structural stabilisation. These data can be used to explain the mechanism by which the antioxidant activity of tea compounds is affected by the addition of milk.

Resveratrol, genistein, and curcumin bind bovine serum albumin.

We report the complexation of bovine serum albumin (BSA) with resveratrol, genistein, and curcumin, at physiological conditions, using constant protein concentration and various polyphenol contents. FTIR, CD, and fluorescence spectroscopic methods were used to analyze the ligand binding mode, the binding constant, and the effects of complexation on BSA stability and conformation. Structural analysis showed that polyphenols bind BSA via hydrophilic and hydrophobic interactions with the number of bound polyphenol (n) being 1.30 for resveratrol-BSA, 1.30 for genistein-BSA, and 1.0 for curcumin-BSA. The polyphenol-BSA binding constants were K(Res-BSA) = 2.52(+/-0.5) x 10(4) M(-1), K(Gen-BSA) = 1.26(+/-0.3) x 10(4) M(-1), and K(Cur-BSA) = 3.33(+/-0.8) x 10(4) M(-1). Polyphenol binding altered BSA conformation with a major reduction of alpha-helix and an increase in beta-sheet and turn structures, indicating a partial protein unfolding.

An overview of structural features of DNA and RNA complexes with saffron compounds: Models and antioxidant activity.

Saffron is the red dried stigmas of Crocus sativus L. flowers and used both as a spice and as a drug in traditional medicine. Its numerous applications as an antioxidant and anticancer agent are due to its secondary metabolites and their derivatives (safranal, crocetin, dimethylcrocetin). In this work we are comparing the spectroscopic results and antioxidant activities of saffron components safranal, crocetin (CRT) and dimethylcrocetin (DMCRT) complexes with calf-thymus DNA (ctDNA) and transfer RNA (tRNA) in aqueous solution at physiological conditions Intercalative and external binding modes of saffron compounds to DNA and RNA were observed with overall binding constants of K(safranal)=1.24x10(3)M(-1), K(CRT)=6.20x10(3)M(-1) and K(DMCRT)=1.85x10(5)M(-1), for DNA adducts and K(safranal)=6.80x10(3)M(-1), K(CRT)=1.40x10(4)M(-1) and K(DMCRT)=3.40x10(4)M(-1) for RNA complexes. A partial B- to A-DNA transition occurred at high ligand concentrations, while tRNA remained in A-conformation in saffron-RNA complexes. The antioxidant activity of CRT, DMCRT and safranal was also tested by the DPPH (2,2-diphenyl-1-picrylhydrazyl) antioxidant activity assay and their IC(50) values were compared to that of well known antioxidants such as Trolox and Butylated Hydroxy Toluene (BHT). The IC(50) values were 95+/-1microg/mL for safranal and 18+/-1microg/mL for crocetin. The inhibition of DMCRT reached a point of 38.8%, which corresponds to a concentration of 40microg/mL.

An overview of DNA and RNA bindings to antioxidant flavonoids.

In this report we are examining how the antioxidant flavonoids can prevent DNA damage and what mechanism of action is involved in the process. Flavonoids are strong antioxidants that prevent DNA damage. The anticancer and antiviral activities of these natural products are implicated in their mechanism of actions. We study the interactions of quercetin (que), kaempferol (kae), and delphinidin (del) with DNA and transfer RNA in aqueous solution at physiological conditions, using constant DNA or RNA concentration 6.25 mmol (phosphate) and various pigment/polynucleotide(phosphate) ratios of 1/65 to 1 (DNA) and 1/48 to 1/8 (tRNA). The structural analysis showed quercetin, kaempferol, and delphinidin intercalate DNA and RNA duplexes with minor external binding to the major or minor groove and the backbone phosphate group with overall binding constants for DNA adducts K(que) = 7.25 (+/-0.65) x 10(4) M(-1), K(kae) = 3.60 (+/-0.33) x 10(4) M(-1), and K(del) = 1.66 (+/-0.25) x 104 (-1) and for tRNA adducts K(que) = 4.80 (+/-0.50) x 10(4) M(-1), K(kae) = 4.65 (+/-0.45) x 10(4) M(-1), and K(del) = 9.47 (+/-0.70) x 10(4) M(-1). The stability of adduct formation is in the order of del>que>kae for tRNA and que>kae>del for DNA. Low flavonoid concentration induces helical stabilization, whereas high pigment content causes helix opening. A partial B to A-DNA transition occurs at high drug concentration, while tRNA remains in A-family structure. The antioxidant activity of flavonoids changes in order delphinidin>quercetin>kaempferol. The results show intercalated flavonoids can make them strong antioxidants to protect DNA from harmful free radical reactions.

DNA interaction with naturally occurring antioxidant flavonoids quercetin, kaempferol, and delphinidin.

Flavonoids are strong antioxidants that prevent DNA damage. The anticancer and antiviral activities of these natural products are implicated in their mechanism of actions. However, there has been no information on the interactions of these antioxidants with individual DNA at molecular level. This study was designed to examine the interaction of quercetin (que), kaempferol (kae), and delphinidin (del) with calf-thymus DNA in aqueous solution at physiological conditions, using constant DNA concentration (6.5 mmol) and various drug/DNA(phosphate) ratios of 1/65 to 1. FTIR and UV-Visible difference spectroscopic methods are used to determine the drug binding sites, the binding constants and the effects of drug complexation on the stability and conformation of DNA duplex. Structural analysis showed quercetin, kaempferol, and delphinidin bind weakly to adenine, guanine (major groove), and thymine (minor groove) bases, as well as to the backbone phosphate group with overall binding constants K(que) = 7.25 x 10(4)M(-1), K(kae) = 3.60 x 10(4)M(-1), and K(del) = 1.66 x 10(4)M(-1). The stability of adduct formation is in the order of que>kae>del. Delphinidin with a positive charge induces more stabilizing effect on DNA duplex than quercetin and kaempferol. A partial B to A-DNA transition occurs at high drug concentrations.

Twenty-four hour continuous ECG recordings in long-distance runners.

Twenty-four hour ambulatory ECG recordings were performed on 20 male long-distance runners, aged 19 to 28 years, during normal activities other than running. Average, maximum, and minimum waking heart rates, respectively, ranged from 58 to 108 (mean +/- SD, 73 +/- 15), 90 to 164 (120 +/- 19), and 34 to 53 (43 +/- 5) beats/min. Longest waking sinus pauses ranged from 1.35 to 2.55 (1.7 +/- 0.3) seconds. Average, maximum, and minimum sleeping heart rates, respectively, ranged from 38 to 58 (47 +/- 6), 69 to 114 (83 +/- 14), and 31 to 43 (36 +/- 3) beats/min. Longest sleeping sinus pauses ranged from 1.60 to 2.81 (2.0 +/- 0.3) seconds. All 20 runners had atrial premature beats, but only one (5 percent) had more than 100/24 hours. Fourteen runners (70 percent) had ventricular premature beats, but only two (10 percent) had more than 50/24 hours, and none had ventricular couplets or ventricular tachycardia. Eight runners (40 percent) had one or more episodes of type 1 second-degree atrioventricular (A-V) block. Compared with untrained males of similar age, the runners had slower heart rates (by approximately 10 beats/min), longer sinus pauses, and a higher prevalence of A-V block. Runners and untrained males did not differ with respect to prevalence of ventricular premature beats, R on T phenomenon, ventricular couplets, or ventricular tachycardia.

Reduced training duration effects on aerobic power, endurance, and cardiac growth.

Thirteen subjects participated in an exercise program of bicycling and running 40 min/day, 6 days/wk. After 10 wk they continued to train either 26 of 13 min/day for an additional 15 wk. Intensity and frequency for the additional 15 wk remained the same as the last 3 wk of training. This study was undertaken to gain further insights into whether the increases in maximum uptake (VO2 max), endurance, and cardiac size can be maintained with reduced training durations. The average increases in VO2 max in response to 10 wk training were between 10 and 20% during the bicycle and treadmill testing. After reduced training, VO2 max continued to remain at the training levels in both groups. Short-term endurance (approx 5 min) was also maintained by both groups. Long-term endurance (2 h or more) remained the same in the 26-min group but decreased significantly (10%, 139-123 min) in the 13-min group. Calculated left ventricular mass increased 15-20% after training and remained elevated after reduced training in both groups. We conclude that it is possible to maintain almost all of the performance increases with up to a two-thirds reduction of training duration. Nevertheless, the data provide initial evidence that all aspects of the endurance-trained state may not be regulated uniformly in reduced training, particularly since VO2 max and short-term endurance were maintained, but long-term endurance decreased in the 13-min group.

Arrhythmias documented by 24-hour continuous ambulatory electrocardiographic monitoring in young women without apparent heart disease.

Results are reported of 24-hour ambulatory ECG recordings in 50 young women without apparent heart disease. During waking periods, maximum (sinus) rates ranged from 122 to 189 beats/min (bpm) (153 +/- 14 mean +/- SD) and minimum rates from 40 to 73 bpm (56 +/- 7). During sleeping periods, maximum and minimum rates ranged from 71 to 128 bpm (105 +/- 13) and from 37 to 59 bpm (48 +/- 6), respectively. Thirty-two subjects (64%) had atrial premature beats, with only one subject (2%) having greater than 100 beats/24 hrs. Twenty-seven subjects (54%) had ventricular premature beats, with only three subjects (6%) having greater than 50 beats/24 hrs. One subject (2%) had one three-beat episode of ventricular tachycardia. Two subjects (4%) had transient type I second-degree atrioventricular block.

Constrictive pericardial disease following mitral valve replacement.

Constrictive pericardial disease developing after open heart surgery is not a well-recognized complication of this procedure. It has been reported only a few times and usually not with good hemodynamic data before and after the subsequent pericardiectomy. We presently report a patient who developed constrictive pericardial disease five years after mitral valve replacement. This was documented with left- and right-sided heart catheterization. The patient underwent pericardiectomy with remarkable clinical improvement. Repeat right- and left-sided heart catheterization done three months postoperatively documented the resolution of the constrictive hemodynamic pattern present before pericardiectomy.

Intraosseous pressure in the lumbar spine.

Intraosseus pressure in the lumbar spine is defined as the hydrostatic pressure of venous blood in the trabecular sinusoids of cancellous bone. Measurements of it in the lumbar vertebrae and the inferior vena cava confirm that the intraosseus pressure in the lumbar spine is identical to, and dependent upon, the pressure in the inferior vena cava. With Valsalva maneuver in the decubitus position, the pressure reaches levels of 90 mm Hg in the inferior vena cava and intraosseus space of lumbar spine.

Myotonia dystrophica and mitral valve prolapse.

Two siblings are described, both afflicted with myotonia dystrophica and mitral valve prolapse. This family supports the recent association of these two familial diseases. One of the siblings had severe conduction disease and recurrent ventricular tachycardia, possibly reflecting potentiation of arrhythmia, because of the association of these two familial diseases.

Left ventricular responses to a program of lower-limb strength training.

Nine healthy male subjects ages 18-27 exercised five days per week. Three days per week they performed five repetitions of squats, leg extensions and leg flexions with maximal resistance for a total of 11 sets. On the other two days each week subjects performed five leg presses and 20 calf raises with maximal resistance. Resting echocardiograms and physiologic evaluations were made prior to starting the strength training and again after ten weeks of training. Resting heart rate +/- SEM before and after training was 65 +/- 2 and 58 +/- 1.7 beats/min (P < .001). Maximal O2 uptake did not change significantly. Left ventricular wall thickness +/- SEM before and after training increased from 0.76 +/- .02 to 0.85 +/- 0.04 cm (P < .05). Left ventricular mass +/- SEM increased from 81.9 +/- 5 to 92.3 +/- 3.7 g (P < .05). The percentage of left ventricular fractional shortening +/- SEM increased from 32 percent +/- 1.2 to 36 percent +/- .9 (P < .001). Lower limb strength training in normal subjects did not increase maximal O2 uptake, but did induce increases in left ventricular wall thickness similar to that seen in champion strength-trained athletes. In addition, improvement in left ventricular performance without significant changes in left ventricular volumes was also observed.

Echocardiographic practice in a large metropolitan area.

A survey performed concerning echocardiography in a metropolitan area. Of 110 hospitals in the area, 62 reported having echocardiographic facilities. Echocardiographic physicians and/or technicians from 41 of these hospitals responded to questionnaires designed to determine the following: (1) educational background and credentials of technicians, (2) average salaries of technicians, (3) role of the physician and technician in the performance and reporting of echocardiograms, (4) volume, cost, and method of storage of echocardiograms, and (5) number and type of echocardiographic units in use. Our data suggest various trends, including a lack of formal training among technicians, the prevalence of cardiologists-internists as directors of echocardiographic facilities, the performance of echocardiograms by cardiology fellows in only 46% of institutions with cardiology training programs, and the widespread projected availability of cross-sectional echocardiographic capability within the next two years.

Echocardiographic demonstration of porcine mitral valve vegetation and dehiscence.

The echocardiographic study of a patient with a malfunctioning porcine valve in the mitral position is presented. Echocardiography of the mitral valve revealed multiple, dense heterogeneous echoes behind and within the valve stent which were suggestive of vegetations. At the aortic valve level, a clear systolic echo in the left atrium was recorded. This echo probably represented the prolapsing anterior valve stent and was caused by a major dehiscence of the valve stent due to endocarditis.

Lack of cardiovascular effects of delta-9-tetrahydrocannabinol in chemically denervated men.

We have previously reported that 25 micrograms/kg of intravenous (i.v.) delta-9-tetrahydrocannabinol (delta-9-THC) produces marked increases in heart rate, prolongation of left ventricular ejection time corrected for heart rate (LVETc), and a shortening of the pre-ejection period in normal volunteers. Beta-adrenergic blockade partially attenuates these responses. To elucidate further the mechanism of action of delta-9-THC, we gave 10 normal volunteers 0.1 mg/kg of i.v. propranolol and 2 mg of i.v. atropine before they received 25 micrograms/kg of i.v. delta-9-THC. Systolic time intervals were compared in the denervated subjects before and after delta-9-THC. Post delta-9-THC responses were measured at a time approximating peak psychologic high. Mean +/- SEM heart rate before and after delta-9-THC was 89 +/- 4 and 87 +/- 3 beats/min (NS); mean +/- SEM pre-ejection period before and after delta-9-TCH was 107 +/- 5 and 109 +/- 4 ms (NS); and mean +/- SEM LVETc before and after delta-9-THC was 433 +/- 6 and 429 +/- 6 ms (NS). Since previous denervation of our subjects with atropine and propranolol totally abolished changes in heart rate and systolic time intervals, the cardiac effects of delta-9-THC appear to be mediated totally via the autonomic nervous system, probably reflecting direct central nervous system stimulation.