Activity of compound G2 isolated from alfalfa roots in experimental dermatophyte infection.

Compound G2 isolated from alfalfa roots was applied topically to skin lesions of guinea pigs experimentally infected with the dermatophyte Trichophyton mentagrophytes var. granulare. After 12 to 15 applications, 80% of the infected lesions were cured, as judged by clinical and microbial criteria, compared with 20% of the untreated lesions which healed spontaneously (P less than 0.01).

Involvement of cysteine residues in the electrophoretic mobility of histone H3 in acid-urea-Triton gels.

Carbamylation of cysteines 96 and 110 in histone H3 increases the electrophoretic mobility of this histone in acetic acid-urea-Triton X-100 polyacrylamide gels but has no effect in gels lacking Triton. Residue 96 appears to be a major determinant in the affinity of histone H3 for the nonionic detergent Triton. Carbamylation and carboxymethylation of cysteine 96 caused a major loss of the gel retardation caused by Triton. Carbamylation of cysteine 110 did not affect Triton binding but prevented ionization of the thiol side-chain moiety in the acetic acid-urea-Triton X-100 gel.

Thiaminase activities and thiamine content of Pteridium aquilinum, Equisetum ramosissimum, Malva parviflora, Pennisetum clandestinum and Medicago sativa.

Thiaminase type 1 and 2 activities and thiamine content of five plants were determined. Of these Pteridium aquilinum and Equisetum ramosissimum were found to have considerably more thiaminase activity and lower thiamine content than Malva parviflora, Pennisetum clandestinum and Medicago sativa.

Digestion, fecal, and blood variables associated with extensive large colon resection in the horse.

Nutritional alterations were evaluated in 9 horses before surgery and 3 weeks, 3 months, and 6 months (4 total trials) after sham operation (group 1; n = 3) or extensive large colon resection (group 2; n = 6). Feed and fecal analyses were performed to determine apparent digestion of dry matter, organic matter, crude protein, calcium, phosphorus, magnesium, potassium, manganese, zinc, copper, and iron, and true digestion of dry matter, organic matter, crude protein, total plant cell wall, hemicellulose, cellulose, and lignin. Additional fecal and metabolic variables included the percentage of fecal water (water in the feces), total fecal water, metabolic organic matter, metabolic crude protein, and metabolic nitrogen. A CBC and standard series of biochemical tests were performed. Large colon resection decreased (P less than 0.05) the true digestion of dietary crude protein and cellulose and apparent digestion of phosphorus, and it increased the fecal metabolic matter and water loss. Total fecal output increased 45% and total fecal water increased 55%. Phosphorus digestion was decreased (P less than 0.05) in group-2 horses, but effects of this were not detected on analysis of blood variables or on physical examination. Nevertheless, after extensive large colon resection, horses can regain body weight lost after surgery and have no overt physical changes when fed an alfalfa pellet diet that meets greater-than-maintenance requirements. Ad libitum water access is suggested, because these horses may have to consume 2 gal/day more than would normal horses.

Use of electrophoretic techniques in determining the composition of seed storage proteins in alfalfa.

Holoprotein molecular weights and polypeptide composition can be determined for complex mixtures of oligomeric proteins using two-dimensional electrophoretic techniques. The variety of two-dimensional analyses presented here is a reflection of the general usefulness of each method for the identification and characterization of the different classes of seed storage proteins in alfalfa. These techniques can be applied to studies of storage proteins in other seeds as well as non-seed storage proteins. The major seed storage proteins in alfalfa are medicagin (a legumin-like globulin), alfin (a vicilin-like globulin) and a family of lower molecular weight albumins (LMW1-3). These comprise 30%, 10%, and 20%, respectively, of the total extractable protein from cotyledons of mature seeds. Alfin is a heterogeneous oligomeric protein (Mr approximately 150,000) composed of polypeptides ranging in size from Mr 14,000 to 50,000 (alpha 1-alpha 6; 50,000, 38,000, 32,000, 20,000, 16,000 and 14,000, respectively). Medicagin is also a high molecular weight oligomeric protein, but requires high concentrations of salt for solubilisation. It is comprised of a family of individually distinct subunits, each composed of an acidic polypeptide (A1-A9; Mr 49,000 to 39,000) linked via disulphide bond(s) to a basic polypeptide (B1, B2, B3; Mr 24,000, 23,000 and 20,000, respectively). This pairing is highly specific and two families are recognizable on the basis of the B polypeptide (B3 or B1/B2). Subunits (Mr approximately 50,000-65,000) are assembled as trimers (8S) or larger oligomers (12S-15S) in mature seeds. The lower molecular weight albumins (LMW1-3) are acidic (pI less than 6), and consist of sets of disulphide-bonded polypeptides (Mr 15,000 and 11,000).

Migration of lead in a glass-lined bottom-unloading silo.

Alfalfa, estimated to contain 25.89 mg lead (Pb)/kg of air dried sample, was ensiled in a glass-lined bottom-unloading silo. Three weeks later air dried haylage from the bottom of the silo contained 118.6 mg of Pb/kg. The concentration of Pb in the haylage was sufficient to induce saturnism in a dairy herd. During the disposal process, the concentration of Pb was determined in each 11.3 m3 load of silage. These data provide evidence that Pb migrated and concentrated in the bottom of the silo.

Simple gas chromatographic method for the determination of medicagenic acid in alfalfa (Medicago sativa).

A gas chromatographic method for the determination of medicagenic acid in alfalfa leaves, stems, entire plant (tops) and roots and also in leaf protein concentrates is described. The method is based on extraction of lucerne saponins followed by hydrolysis of the triterpene glycosides. After derivatization of the liberated sapogenins to silylated products, the trimethylsilylated medicagenic acid was determined by gas chromatography. The sensitivity of the method permits the detection of 50 ng of medicagenic acid.

Extraction of light filth from whole leaves of alfalfa, lemon balm, papaya, and spearmint: collaborative study.

Results are reported for a collaborative study to extend AOAC method 44.A06-44.A08 to extraction of light filth from whole leaves of alfalfa, lemon balm, papaya, and spearmint. A 5 g (spearmint) or 10 g (alfalfa, lemon balm, papaya) test portion is defatted with isopropanol in a simple reflux apparatus. Rat hairs, insect fragments, and whole insects are isolated by wet sieving on a No. 230 sieve, a deaerating boil in 40% isopropanol, and flotation with mineral oil-heptane (85 + 15) from Tween 80-Na4EDTA (1 + 1) and 40% isopropanol in a Wildman trap flask. Each product was spiked at a different level. For rat hairs, recoveries averaged 82.2% from alfalfa, 88.9% from lemon balm, 80.6% from papaya, and 79.6% from spearmint. Recoveries of whole or equivalent insects from these products averaged 66.1, 218.8, 69.4, and 85.4%, respectively; recoveries of insect fragments from these products averaged 89.6, 94.4, 94.1, and 88.1%, respectively. The method has been adopted official first action for extraction of light filth from whole leaves of alfalfa, papaya, and spearmint. The extension of the method to lemon balm was not recommended because of interferences by intrinsic whole insects, which were the same species as the spike material.

Histone variants and acetylated species from the alfalfa plant Medicago sativa.

The histones from the alfalfa plant Medicago sativa have been characterized in terms of type variants and levels of acetylation. Histones were isolated directly from total plant tissue (callus), eliminating the need to develop methods for nuclear isolation. An acid-urea-polyacrylamide gel with a transverse Triton X-100 gradient resolved and identified in a single gel at least one type of histone H4, two variant forms of histone H2B, two variant forms of histone H3, and four variant forms of histone H2A from a crude histone preparation. Histone H4 was present 25% in an unmodified state and 75% as monomodified, presumably as monoacetylated histone. Both histone H3 variants displayed five bands, consistent with up to four internal sites of acetylation. The two H3 variants differed in their steady-state level of acetylation, suggesting that they may reside in different chromatin environments. Several histone H1 species were identified by solubility and cross-reactivity with antiserum raised against the globular part of bovine H1(0), indicating conservation of epitopes between histone H1 of mammals and higher plants.

Effects of varying zinc concentrations on quality of alfalfa for lambs.

Zinc concentrations in alfalfa hay were varied using a N-Zn liquid fertilizer as a foliar applicant (.34 or .68 kg Zn/ha) or as a soil fertilizer (4.07 kg Zn/ha). Mean concentrations of Zn across five cuttings of alfalfa in 2 yr were 18, 27, 41 and 21 mg Zn/kg DM for control, low foliar, high foliar and soil treatments, respectively. Each treatment was fed in ad libitum amounts to eight crossbred wether lambs (20 to 35 kg) in 6-wk growth and intake trials, followed by 2-wk digestibility and balance trials with individual lambs. For one cutting, hays were also fed in an 81-d trial to four ram lambs (30 to 35 kg) and live weight gain and testicular development were measured. Average daily gain (ADG) and intake over 6 wk differed (P less than .01) with cutting but not with Zn treatment. Average daily gain and testes weight of ram lambs also were not affected by treatment. In the metabolism trials, Zn treatment did not alter (P greater than .05) intake or dry matter digestibility (DMD) of alfalfa, but did influence digestibility of neutral detergent fiber (NDF). Digestible NDF (%) was higher (P less than .05) for high foliar than for low foliar treatments. Apparent absorption and retention of Zn was significantly greater for control than for Zn-treated alfalfas and did not differ with cutting. Mean serum Zn concentrations for control, low and high foliar, and soil treatments were .79, .81, .78 and .75 micrograms Zn/ml, respectively, for all cuttings, with no differences due to treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Application of polyelectrolytes to fractionation of alfalfa juice protein (short communication).

The quantities of protein which can be extracted from green plants depend on a number of factors such as the botanical composition of the plant, its growth stage, topdressing and system of extraction. Leaf protein concentrates have been incorporated into food for human consumption, but their green color has limited its application. Thermal precipitation of chloroplastic material is not specific, causes denaturation and requires high energy input. A selective recovery of the green fraction is possible by centrifugation at high centrifugal forces or by the combination of this method with an addition of calcium salt to a green juice. Application of high molecular polyelectrolytes in green juice precipitation allows the separation of pigmented material at room temperature. The objective of this work was to determine the effect of cationic and anionic flocculants on fractionation of alfalfa green juice into two fractions: chloroplastic (green LPC) and cytoplasmatic (white LPC).

T-DNA and opine synthetic loci in tumors incited by Agrobacterium tumefaciens A281 on soybean and alfalfa plants.

We report here the molecular characterization of transferred DNA (T-DNA) in leguminous tumors incited by Agrobacterium tumefaciens A281 harboring the tumor-inducing plasmid pTiBo542. The T-DNA is composed of two regions named TL (left portion)-DNA and TR (right portion)-DNA, in accordance with the nomenclature for the octopine strains. TL-DNA is defined by several internal HindIII restriction fragments totaling 10.8 kilobase pairs (kbp) in uncloned soybean and alfalfa tumors. Alfalfa tumor DNA may contain one more HindIII fragment at the left end of TL-DNA than does soybean tumor DNA. TR-DNA has a 5.8-kbp BamHI-EcoRI internal fragment. All borders other than the left border of TL-DNA appear to be the same within the detection limits of Southern blot hybridization experiments. The two T-DNA regions are separated by 16 to 19 kbp of DNA not stably maintained in tumors. The distance from the left border of TL-DNA to the right border of TR-DNA is approximately 40 kbp. Loci for the mannityl opines are situated in TR-DNA, based on genetic and biochemical criteria.

Characterization of a Rhizobium meliloti fixation gene (fixF) located near the common nodulation region.

Rhizobium meliloti 2011 DNA from pRmSL26, a plasmid which is known to carry genes involved in the early stages of nodulation, was used to construct Tn5 mutations by site-directed Tn5 mutagenesis. Tn5 mutations located within an 8.7 kilobase EcoRI fragment defined two adjacent loci encoding functions for nodulation (nod) and symbiotic N2 fixation (fix). We investigated the organization and regulation of the fix locus and the characteristics of alfalfa nodules induced by these Fix- mutants. By monitoring expression in Escherichia coli minicells, we determined that the fix locus encoded a 36-kilodalton polypeptide. The gene corresponding to this locus was designated fixF. Morphological and ultrastructural studies of the ineffective nodules formed by R. meliloti fixF mutants showed infected host cells similar to those of the wild type. The ineffective nodules were able to accumulate leghemoglobin, but at lower levels than those found in the wild-type nodules. Expression of the nifHDK operon was unaffected by Tn5 insertions in the fixF gene. Expression of the fixF gene was monitored in E. coli by using translational lacZ fusions. It was shown that transcription of the fixF gene in E. coli could be activated by Klebsiella pneumoniae nifA and the R. meliloti nifA-like regulatory gene products. Expression of the fixF gene was also studied in free-living and symbiotic R. meliloti cells. It was found that the fixF gene was transcribed in the symbiotic state.

Residues of five pesticides in field-treated alfalfa seeds and alfalfa sprouts.

Residues of five different pesticides applied to alfalfa seed crops were determined in the harvested seeds and in sprouts grown from these seeds. Although seeds are usually used for future production of alfalfa plants, some of these seeds may be sprouted for human food consumption. The pesticides studied--aldicarb (Temik), chlorothalonil (Bravo), chlorpyrifos (Lorsban), methamidophos (Monitor) and propargite (Comite)--were applied at a normal usage rate and at two to three times that rate. Residues on the seeds and sprouts, if any, were insignificant at rates of application.

Wound-induced trypsin inhibitor in alfalfa leaves: identity as a member of the Bowman-Birk inhibitor family.

The primary structure of the wound-inducible trypsin inhibitor from alfalfa (ATI) establishes it as a member of the Bowman-Birk proteinase inhibitor family. The time course of induction of ATI in alfalfa following wounding is similar to the induction of the nonhomologous proteinase inhibitors I and II in tomato and potato leaves, and, like inhibitors I and II, ATI is induced to accumulate in excised leaves supplied with the proteinase inhibitor inducing factor from tomato leaves. The similarity of the wound induction of ATI to that of inhibitors I and II indicates that wound-regulated systems are present in Solanaceae and Leguminosae plant families that possess a common fundamental recognition system regulating synthesis of proteinase inhibitors in response to pest attacks. ATI is the first Bowman-Birk inhibitor that has been found in leaves and is the only member of this family known to be regulated by wounding.

Purification and characterization of two leaf polypeptide inhibitors of leaf protease from alfalfa (Medicago sativa).

Two polypeptides with antiproteolytic activities have been isolated from alfalfa leaves. Polypeptide I resembles the previously described plant protease inhibitors in both structural and functional features; it has a molecular weight of 15,000, a random coil secondary structure, and inhibits exogenous protease as well as alfalfa leaf protease. Polypeptide II is a novel type of plant inhibitor with a molecular weight of 6300 and a highly organized structure with a high (40-50%) alpha-helix content. It only inhibits endogenous protease with a molar stoichiometry polypeptide/enzyme protein of 1.