Free PAs Research Articles

Protective Effect of Hydrogen Peroxide against Paraquat Toxicity in Young Pea Plants: Possible Role of Endogenous Polyamines

Young pea plants grown as a water culture were sprayed with hydrogen peroxide and 24 h later were treated with the herbicide paraquat. The effects of paraquat and H2O2 treatment were investigated on the endogenous concentrations of free proline, and free and conjugated putrescine, spermidine and spermine. Application of PQ increased the free proline and Put, and decreased conjugated and bound polyamines. The results clearly demonstrate the oxidative damages provoked by PQ application. Single treatment with H2O2 provoked considerable decrease in the free fraction of the three PAs studied 2 h after light exposition, but caused an increase in their respective conjugated fractions. Data suggest that PA changes after H2O2 treatment are due to conversion from free to conjugated form. Three hours later free PAs in H2O2 treated-plants reached control levels; however the conjugated remained higher than the control. No alterations were detected in proline concentration after H2O2 treatment. Pretreatment with Н2О2 activated some component of the plant protection mechanisms by causing an alteration in free/conjugated PAs ratio in plants subsequently subjected to PQ treatment. Data concerning PA and proline concentrations in plants treated with both compounds supposed that H2O2 shows a protective role against PQ and improves the plant tolerance to the oxidative stress generated by paraquat.

Development of a highly biocompatible antituberculosis nanodelivery formulation based on para-aminosalicylic acid-zinc layered hydroxide nanocomposites.

Tuberculosis is a lethal epidemic, difficult to control disease, claiming thousands of lives every year. We have developed a nanodelivery formulation based on para-aminosalicylic acid (PAS) and zinc layered hydroxide using zinc nitrate salt as a precursor. The developed formulation has a fourfold higher efficacy of PAS against mycobacterium tuberculosis with a minimum inhibitory concentration (MIC) found to be at 1.40 μg/mL compared to the free drug PAS with a MIC of 5.0 μg/mL. The newly developed formulation was also found active against Gram-positive bacteria, Gram-negative bacteria, and Candida albicans. The formulation was also found to be biocompatible with human normal lung cells MRC-5 and mouse fibroblast cells-3T3. The in vitro release of PAS from the formulation was found to be sustained in a human body simulated phosphate buffer saline (PBS) solution at pH values of 7.4 and 4.8. Most importantly the nanocomposite prepared using zinc nitrate salt was advantageous in terms of yield and free from toxic zinc oxide contamination and had higher biocompatibility compared to one prepared using a zinc oxide precursor. In summary, these promising in vitro results are highly encouraging for the continued investigation of para-aminosalicylic acid and zinc layered hydroxide nanocomposites in vivo and eventual preclinical studies.

Profiling of Alkaloids and Eremophilanes in Miracle Tea (Packera candidissima and P. bellidifolia) Products

Commercial preparations of the Mexican herbal drug known as “miracle tea” (Packera candidissima and P. bellidifolia) have been profiled qualitatively by HPLC and GC-MS. Pyrrolizidine alkaloids were identified in the alkaloid extracts. The content of free PAs and their N-oxides was determined for a total of 22 samples and the results showed that the amount of these hepatotoxic compounds (0.0005–0.94% free PAs; 0.0004–0.55% N-oxides), through the presence of retrorsine (1) and senesionine (2) as the main constituents, may reach toxic levels. Eremophilanes (3-7) were the major components found in the hexane-soluble fraction. Financial Support: CONACyT (101380-Q).

Effects of hydrogen cyanamide on antioxidant enzymes’ activity, proline and polyamine contents during bud dormancy release in Superior Seedless grapevine buds

The effect of hydrogen cyanamide (HC) on dormancy release, antioxidant enzyme’s activity and proline and free polyamine contents were investigated in ‘Superior Seedless’ grapevine buds. HC application caused a sharp decrease of catalase (CAT, EC 1.11.1.6) activity and a transient stimulation during the 5 days following treatment of peroxidase (POD, EC 1.11.1.7) and ascorbate peroxidase (APX, EC 1.11.1.11) activities. This coincided with an accumulation of total free polyamines, especially putrescine (Put). Proline content increased dramatically. There was a strong correlation between APX and POD activities and total free PAs and Put contents implying a possible stimulating effect of the latter compounds on these enzymes. These observations indicate that HC triggers an oxidative stress leading to bud endodormancy release. Afterward, as budbreak started, we observed a rapid proline and Put degradation; this could be responsible for reactivation of growth. Indeed, the decline in Put to (Spd + Spm) ratio, reported here, may be considered as a reliable biochemical marker of bud growth resumption.

Flow-dependent channel formation in clots by an erythrocyte-bound fibrinolytic agent

AbstractStudies in animal models have shown that plasminogen activators bound to erythrocytes (RBC-PA) have an extended lifetime in the circulation and are safer than free PAs. RBC-PAs incorporate into nascent thrombi, which are focally lysed from within, an attractive thromboprophylactic option. In static systems, RBC-PAs cleave surrounding fibrin fibers, forming pores larger than the cells themselves, and move around the pore edges, enlarging them until eventual clot dissolution. We hypothesized that under flow in blood vessels, RBC-PAs form functional patent channels before clot dissolution. Here we used perfusion chambers to study clot lysis by RBC-PAs under static versus arterial and venous flow conditions. We found that flow decelerates bulk clot lysis but quickly generates patent channels filled with passing RBCs, via pore enlargement and merging in the direction of flow. Formation of such channels by RBC-PAs may help rescue ischemic tissue before bulk dissolution of potentially occlusive clots.

DNA demethylation and decrease on free polyamines is associated with the embryogenic capacity of Pinus nigra Arn. cell culture

Embryogenic cell lines initiated from immature zygotic embryos of Pinus nigra Arn. ssp. Austriaca were characterized in terms of macromorphological traits (colour, bipolar structures formation, germination ability) and their embryogenic potential was defined as high, medium or null. Quantification of global genomic DNA methylation revealed the existence of specific DNA methylation levels for the determinated embryogenic potentials. The line considered as effectively embryogenic, i.e., with the ability of develop the whole embryogenic program and producing plants, showed the lowest methylation levels. There was also proved the existence of an inverse relationship between total contents of free PAs and embryogenic potential, being the highest contents of free putrescine and spermidine in the non-embryogenic line and the lowest in the effectively embryogenic one. Relationships among DNA methylation levels, profiles of free individual polyamine contents and embryogenic potentials based on the ability to produce well-formed somatic embryos with effective plant conversion are discussed.

Polyamines enhance chilling tolerance of cucumber ( Cucumis sativus L.) through modulating antioxidative system

Cucumber chilling-resistant cultivar Changchun mici and -sensitive cultivar Beijing jietou were used in this study to investigate the effects of exogenous PAs on protection against chilling injury as well as on changes of physiological features, and the fluctuation of free PAs content in the leaves under chilling stress. Upon chilling treatment, free spermidine (Spd), spermine (Spm) and putrescine (Put) were remarkably induced in the leaves of cv. Changchun mici 1 day after treatment. The induction of Put declined thereafter, whereas Spd and Spm levels increased steadily. In the leaves of cv. Beijing jietou, Put content was increased only at 1 day after chilling while Spd content decreased significantly upon chilling treatment. Chilling reduced soluble protein content, and decreased the activities of antioxidant enzymes, including superoxidase dismutase (SOD), peroxidase (POD), catalase (CAT) and ascorbate peroxidase (APX) only in Beijing jietou. However, these changes could be renovated by exogenous application of Put and Spd. It was also found that pretreatment with Put and Spd diminished the increased electrolyte leakage and malondialdehyde (MDA) content caused by chilling in the leaves of both cultivars. Pretreatment of methyglyoxal-bis-(guanylhydrazone) (MGBG), the PAs biosynthetic inhibitor cancelled the effects of PAs in most of the treatments. Moreover, histochemical staining and quantitative measurements showed that exogenous application of Put and Spd eliminated but MGBG exaggerated the hydrogen peroxide (H 2O 2) accumulation caused by chilling stress, especially in leaves of Beijing jietou. Interestingly, Changchun mici was found to contain higher endogenous free PAs contents compared to Beijing jietou. While no significant difference of SOD, POD and CAT activities was found between non-chilling Changchun mici and Beijing jietou seedlings, the former exhibited higher APX activity than the latter. These results suggest that PAs play important roles in the tolerance of cucumber against chilling stress, which is most likely achieved by acting as oxidative machinery against chilling injury.

Évolution des teneurs en polyamines libres et conjuguées dans les feuilles de vigne (Vitis vinifera 'Cabernet Sauvignon') saine et atteinte d'eutypiose

Effects of Eutypa lata, the fungal agent of eutypiosis, on polyamine (putrescine (Put), spermidine (Spd), spermine (Spm), diaminopropane (Dap)) and tyramine (Tyr) contents in grapevine leaves were studied to determine the relationship between these components and disease development. The external symptoms of eutypiosis are small and necrotic leaves, which could be observed on only one arm or on two arms of the vine stock. This study was carried out on healthy leaves (S, borne by healthy vine stock), apparently healthy leaves (Aps, borne by one arm without symptoms, whereas the other arm was diseased), and moderately (M. mal.) and strongly (F. mal.) diseased leaves. We analysed free polyamines (free PAs, chlorhydric acid soluble fraction), conjugated polyamines (PAs-fpm, PAs covalently linked to molecules with low molecular mass like hydroxycinnamic acids), and bound polyamines (PAs-mm, covalently linked to macromolecules) in Vitis vinifera L. 'Cabernet sauvignon', a variety sensitive to eutypiosis. In diseased leaves, we noted a decrease of free PAs, PAs-fpm, free Tyr, and Tyr-fpm levels. By contrast, the Dap-fpm level was increased by the disease. Eutypiosis also induced an increase of Spm-mm and Dap-mm levels, whereas the Put-mm and Spd-mm levels were decreased. Free Put, free Spd, PAs-fpm, free Tyr, and Tyr-fpm levels were higher in apparently healthy leaves than in healthy leaves. The PAs-mm levels were slightly affected by eutypiosis in apparently healthy leaves. The specific role of each type of polyamines and tyramine is discussed with regard to the host-parasite relationship.Key words: polyamines, leaf, Eutypa lata, Vitis vinifera.

Effects of eutypiosis on free and conjugated polyamines content in internodes of grapevine (<em>Vitis vinifera</em> l. cv. Cabernet sauvignon) during their active phase of growth

<p>Development of polyamines levels [Putrescine (Put), Spermidine (Spd), and Spermine (Spm)] was investigated in internodes of grapevine infected by eutypiosis fungus, <em>Eutypa lata</em>. The study was performed using (1) internodes from healthy vines, (2) apparently healthy internodes from vines with one arm symptomless and one diseased arm and (3) diseased internodes. Free polyamines (PAs: hydrochloric acid soluble fraction), conjugated polyamines (PAs-fpm: PAs covalently linked to molecules with low molecular weight like hydroxycinnamic acids) and bound polyamines (PAs-mm: covalently linked to macromolecules) were analysed in <em>Vitis vinifera</em> L. cv. Cabernet sauvignon, which is sensitive to eutypiosis.</p><p>The internodes stunting induced by eutypiosis was accompanied by a decrease of free Put and free Spm contents between flower buds separated and fruit set stages. The same results were observed for PAs-mm but more slightly than free PAs. The Put-fpm and Spd-fpm levels increased in the apparently healthy internodes and decreased in diseased organs by <em>Eutypa lata</em> effect. The relationship between free PAs contents in the diseased internodes and their growth reduction, as well as the involvement of PAs-fpm on the grapevine response to <em>Eutypa lata</em> infection are discussed.</p>

Studies on polyamines in Solanum tuberosum in vitro: Effects of DFMO, DFMA, chlorogenic acid and putrescine on the endogenous distribution of polyamines, tuberization and morphology

Summary The effects of inhibitors of putrescine (PUT) biosynthesis, DL-α-difluoromethylornithine (DFMO) and DL-α-difluoromethylarginine (DFMA), chlorogenic acid representing phenolic conjugation partners of PAs, and exogenous putrescine (PUT), on the distribution of free and conjugated PAs were examined in relation to their effects on growth, tuberization and morphology in Solanum tuberosum . Changes in the profile of PAs were consistendy associated with effects on tuberization. DFMO totally prevented, but DFMA only decreased, tuberization. Thus, it is concluded that arginine decarboxylase-activity is low and the PAs required are predominandy generated via the ornithine decarboxylase pathway. DFMO treatment increased the level of free PAs. At 5 mmol/L DFMO the formation of conjugated PAs was strongly reduced, whereas at 1.8 mmol/L conjugated PAs accumulated to a level higher than in untreated controls. Chlorogenic acid promoted tuber formation and tuber growth in a concentration-dependent manner, involving conjugation with PAs. Exogenous PUT, in general, caused improved growth of leafy shoots, rooting and loss of tuber dormancy. Tuberization was either promoted or inhibited, depending on the concentration applied. Added to the tuber-inducing control medium, low concentration (0.2 mmol/L) of PUT accelerated tuberization, establishing the PA-profile typical of tuberization. A high concentration (5 mmol/L) prevented full tuberization and disturbed the typical PA-profile. Neither concentration counterbalanced the impact of DFMO on the distribution of endogenous PAs and tuberization. However, exogenous PUT counteracted the decrease in tuberization due to DFMA treatment and also the delay in tuberization onset caused by chlorogenic acid, which was putatively due to reduced levels of free PAs as a result of conjugation. The results of monitoring the changes in the distribution of endogenous PAs following treatments that perturb tuberization support the conclusion that a specific balance of free and conjugated PAs is required for optimal progress through tuberization.

Polyamines and seed germination

This review examines the presence of free and bound PAs during the life cycle of seeds. Although data are still rather scarce, PAs, present, but with an unknown function during embryogenesis, are accumulated in relatively high quantities in mature dry seeds of some species. PA concentration can be altered by the process of stratification, and PAs are synthesized in growing organs after germination, but it is also possible that the reserve organs synthesize and export some types of PA to the axis. This metabolism and sink-source relationship depend on the germination conditions and on the greater or lesser presence of enzymes related to synthesis (ADC, ODC, LDC and AdoMetDC) and degradation (DAO and PAO) of free PAs. The biochemical mechanism binding the PAs to a series of compounds of low and high molecular weight is currently unknown in seeds. The present study also considers intra- and intercellular compartmentation of free and bound PAs in growing axes and the physiological implications of the ethylene-production pathway in relation to seed behaviour.

Reactions of cloned poplars to air pollution

Ozone-induced changes in ethylene production, ACC oxidase activity and the contents of ACC, MACC and free PAs were studied inPopulus nigra L. cv. Loenen with high ozone sensitivity as judged by the degree of chlorophyll degradation and premature leaf abscission.Ethylene production, ACC oxidase activity, ACC content and MACC levels were induced by the one-, two-, and three-week ozone exposure (36±9 ppb O3 for 11 hours a day). In addition, increases in PA levels, especially in spermidine, were measured in ozone treated plants.The role of free PAs and MACC synthesis as possible antisenescence reactions is discussed.

Receptor-mediated delivery ofp-aminosalicylic acid conjugated to maleylated serum albumin againstmycobacterium tuberculosisinfection in guinea pigs

AbstractWe have previously reported that the scavenger receptor-mediated uptake of para-aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) resulted in elimination of intracellular Mycobacterium tuberculosis (MTB) in mouse peritoneal macrophages in culture (Majumdar, S., and Basu, S. K. (1991) Antimicrob. Agents Chemother. 35, 135–140). In this paper, we report therapeutic efficacy of PASMBSA conjugate in vivo against MTB infection in guinea pigs. Two or 4 intravenous (iv) injections of PAS–MBSA conjugate weekly led to a greater reduction in the number of bacteria in the lung, liver, and spleen of the infected animals than the reduction caused by free PAS. PAS–MBSA treatment also in-creased the survival rate of the infected animals. High and persistent drug activity was observed in the target organs when the drug was delivered as PAS–MBSA conjugate. The superior efficacy of PAS–MBSA is presumably due to selective delivery of the conjugate to the macrophages, resulting in enhance...

Killing of intracellular Mycobacterium tuberculosis by receptor-mediated drug delivery

p-Aminosalicylic acid (PAS) conjugated to maleylated bovine serum albumin (MBSA) was taken up efficiently through high-affinity MBSA-binding sites on macrophages. Binding of the radiolabeled conjugate to cultured mouse peritoneal macrophages at 4 degrees C was competed for by MBSA but not by PAS. At 37 degrees C, the radiolabeled conjugate was rapidly degraded by the macrophages, leading to release of acid-soluble degradation products in the medium. The drug conjugate was nearly 100 times as effective as free PAS in killing the intracellular mycobacteria in mouse peritoneal macrophages infected in culture with Mycobacterium tuberculosis. The killing of intracellular mycobacteria mediated by the drug conjugate was effectively prevented by simultaneous addition of excess MBSA (100 micrograms/ml) or chloroquine (3 microM) to the medium, whereas these agents did not affect the microbicidal action of free PAS. These results suggest that (i) uptake of the PAS-MBSA conjugate was mediated by cell surface receptors on macrophages which recognize MBSA and (ii) lysosomal hydrolysis of the internalized conjugate resulted in intracellular release of a pharmacologically active form of the drug, which led to selective killing of the M. tuberculosis harbored by mouse macrophages infected in culture. This receptor-mediated modality of delivering drugs to macrophages could contribute to greater therapeutic efficacy and minimization of toxic side effects in the management of tuberculosis and other intracellular mycobacterial infections.

A STUDY OF PAS BLOOD CONCENTRATION

PAS blood concentration was studied on 17 Korean patients with pulmonary tuberculosis and 6 healthy Scandinavians. It will be certainly insufficient number to draw any conclusion but some trends were seen as follows through the result. 1) There was no marked difference of free PAS concentration among the preparations we have studied with.(Ca-PAS U.S.P., Ca-PAS J.P., Na-PAS tab., Na-PAS granule) 2) The peak value of single-dose was approximately twice as high as the value of two divided dose, and the peak was 4 hours after medication but the 8 hour value was about the same. 3) With divided dose the blood concentration maintained 2 to 6 mg. %, and with single-dose it maintained 5-15mg. of free PAS. 4) The value in Scandinavian was approximately twice as high as in Korean. 5) The individual difference in Korean was relatively big, particularly with Na-PAS tab, and. Ca-PAS U.S.P. but it was not prominent in Scandinavian. 6) There was no side reaction or difficulty with medication except for the large amount with Ca-PAS J.P. but was not significant.

Renal excretion of para-aminosalicylic acid: a two-way transport system in the dog.

The renal excretion of free para-aminosalicylic acid (PAS) was studied in dogs. With increasing loading and sustaining infusion of the compound alone, a negative T (rate of urinary excretion minus the rate of filtration of PAS) was observed, but a positive T was obtained during added sodium bicarbonate or sodium acetate infusion. While the positive T values were greater during acetate than in bicarbonate infusion, urinary pH was higher in the latter. Studies with intrarenal arterial injection and stop-flow techniques showed evidence of a bidirectional transport of free PAS. The reabsorption of PAS was influenced by the urinary pH. Sodium bicarbonate infusion reduced the reabsorption; lactic acid infusion increased it. Secretion of the free PAS does not require a high urinary pH and is augmented by acetate and blocked by probenecid. This suggests that the secretion of free PAS involves an active process, analogous, though not necessarily identical to the system for PAH and for other related substances.

Prolonged Blood Levels with Sustained Action PAS

1.Sustained action PAS tablets (Parasal-S.A., 1.0 Gm., sodium free) offer a convenient means of PAS therapy with a high degree of toleration. Acceptance of this drug was excellent since a four gram dose of free PAS consisted of only four tablets. Patients who manifested symptoms of gastrointestinal irritation due to standard PAS preparations tolerated sustained action PAS tablets with minimal, if any, gastrointestinal side-effects.2.Sustained action PAS tablets, in dosage of four grams (4 tablets) three times daily at six hour intervals, generally provided sustained therapeutically effective PAS blood concentrations for 24 hours. No other PAS preparation, in equivalent dosage, studied to date, produced such sustained 24 hour PAS blood levels.3.The clinical implications and probable advantages inherent in the use of sustained action PAS, administered concurrently with isoniazid or streptomycin, are believed to be as follows: a) sustained PAS concentrations within the tuberculous lesions; b) sustained and increased biologically active isoniazid levels in the blood and diseased tissues; c) greater inhibition or delay in the emergence of isoniazid or streptomycin resistant bacilli. Sustained action PAS may, therefore, serve to potentiate the therapeutic efficacy of antituberculous combined chemotherapy regimens.

Serum Concentrations of Para-Aminosalicylic Acid (PAS) Produced by Various Forms of PAS

The concentration of free and total PAS in the blood serum of 26 patients was determined after single and multiple oral doses of various forms of free acid PAS and the sodium salt of PAS. Free acid PAS powder and the sodium salt of PAS yielded approximately the same concentrations after single or multiple oral doses. Compressed tablets or enteric-coated granules of free acid PAS gave lower concentrations than either of the first two, apparently due to delayed rather than incomplete absorption of the drug. The concentration was not affected by the concurrent administration of sodium bicarbonate or ammonium chloride. As sodium PAS causes fewer symptoms of gastro-intestinal irritation, it is the preparation of choice for most patients.