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Received: 11 December Accepted: 30 May Among the different toxins that N. In the lolitrem group, lolitrem-B is considered as the most important for livestock, being mainly responsible for the ryegrass staggers syndrome or ryegrass tremor Gallagher et al. This intoxication affects bovines, sheep, and equines that grazed ryegrass pasture infected with the fungus, a neuromuscular alteration becoming apparent with tremors and variable degrees of hypersensibility to external stimuli, uncoordinated movements, rigidity in walking, and ataxia Siegel et al.
Also mentioned is the fact that it can affect animal production Lanuza et al. The concentration of lolitrem-B varies according to climatic conditions Oldenburg,plant tissues Keogh and Tapper, ; Ball et al. Keogh et al. Bearing in mind that the pest L. Therefore, since Chile imports perennial ryegrass mostly from what is relationship marketing pdf country, it is necessary to adequately manage this technology.
This occurs by knowing the endophyte level contained in the seeds and the toxin levels, such as lolitrem-B contained in the plants and plant parts throughout the year. This would allow carrying out adequate management of pasture use and cutting, with the objective of optimizing forage production and decreasing the risk of animal intoxication Lanuza et 3 examples of mutualism in the desert. It is therefore relevant to develop a quantitative method that allows evaluating lolitrem-B in the prairies of southern Chile.
The isolation of lolitrem-B has been described in plant tissues and its identification by mass spectrometry and nuclear magnetic resonance spectroscopy Gallagher et al. Its structure was determined by Gallagher et al. Grancher et al. To determine it, extraction methods with solvents have been described and subsequent quantification by high-performance liquid chromatography HPLCwith ultraviolet and fluorescence detector Gallagher et al. The objective of this study was to develop a low-cost and low solvent consumption analytical method to determine lolitrem-B in ryegrass samples and its quantification by HPLC with fluorescence detector.
Preparation of lolitrem-B standard All the used reagents were suitable can horses get grass staggers residue analyses. With this diluted standard, the curve was prepared and the additions to the ryegrass sample without endophyte were done. Method tests Sample preparation for assay. Diverse consecutive tests were done to optimize can horses get grass staggers obtain the method. For evaporation, to reduce costs, the use of nitrogen versus air in purification was compared, and since it was not indicated in the methods, different quantities of silica gel 60 were studied 0.
Method description For the recovery study carried out in five standard addition levels 0. Diluted lolitrem-B standard was added to five tubes, a sixth was the control without standard addition, and another without a sample was the blank reaction tube. The following procedure was applied to each tube. Extraction: 0. Purification or clean up Preparation of columns for clean-up. Some 3 mL polypropylene syringes were used, a 9 mm diameter paper filter Whatman 41, Maidstone, England was put at the bottom of the syringe, plus 0.
The extraction residue was reconstituted with 1. Lolitrem-B retention time was 2. Due to the toxicity of lolitrem-B, precaution was taken to collect the eluding solvent from can horses get grass staggers chromatograph in a bottle under the solvent extractor hood. One of the precautions that must be taken when preparing the columns is to not compact them too much since this decreases toxin recovery. The calibration curve in the 0. Table 1. Recovery and coefficient of variation of lolitrem-B standard at different addition levels.
The highest CV 5. Gallagher et al. The time required per sample in Can horses get grass staggers with the established conditions was 5 min, unlike the method used by Hovermale and Craig in which the time cycle between injections was 24 min. Figure 1 shows the chromatographs obtained for a perennial ryegrass sample without endophyte, the lolitrem-B comparative standard and the perennial ryegrass sample without endophyte but fortified with 0.
Figure 1. The extraction, purification, and quantification method used allowed determining lolitrem-B in perennial ryegrass samples with no interference. The linear range in which lolitrem-B can be determined in the perennial ryegrass samples was from 0. Furthermore, since it is a micro-method, it has low solvent consumption leading to minor costs and decreased risk for both the operator and the environment. Barker, R. Prestidge, and J. Distribution and accumulation of the mycotoxin lolitrem-B in Neotyphodium lolii-i nfected perennial ryegrass.
Interrelationships between Acremonium lolliperamine, and lolitrem-B in perennial ryegrass. Davies, G. Lane, G. Latch, H. Nott, and B. Effect can horses get grass staggers water deficit on alkaloid concentrations in perennial ryegrass endophyte associations. In Hume, D. Latch, and H. Easton eds. Radostits, J. Arundel, y C. Medicina veterinaria: Libro texto de las enfermedades del ganado vacuno, ovino, porcino, caprino y equino. Lane, D. Baird, and E. Seasonal variations of alkaloid concentrations in two perennial ryegrass B endophyte associations.
In Paul, V. Dapprich eds. Hawkes, and J. Rapid determination of the neurotoxin lolitrem-B in perennial ryegrass by high performance liquid chromatography with fluorescence detection. Hawkes, P. Steyn, and R. Tremorgenic neurotoxins from perennial ryegrass causing ryegrass staggers disorder of livestock: Structure elucidation of lolitrem-B.
White, and P. Ryegrass staggers: isolation of potent neurotoxins lolitrem A and lolitrem-B from staggers-producing pastures. Jaussaud, A. Durix, A. Berthod, B. Fenet, Y. Moulard, et al. Countercurrent chromatographic isolation of lolitrem-B from endophyte-infected ryegrass Lolium perenne L. Cromatography A Correlation of ergovaline and lolitrem-B levels in endophyte-infected perennial ryegrass Lolium perenne.
Acremonium loliilolitrem-B, and peramine concentrations within vegetative tillers of perennial ryegrass. Tapper, and R. Distributions of the fungal endophyte Acremonium loliiand of the alkaloids lolitrem-B and peramine, within perennial ryegrass. Tapper, E. Davies, D. Hume, G. Latch, D. Barker, et al. Effect of growth conditions on alkaloid concentration in perennial ryegrass naturally infected with endophyte.
In Bacon, C. Hill eds. Torres, y E. Antecedentes generales del complejo Listronotus example of causation hypothesis lolii en la producción bovina. In Lanuza, F. Cisternas eds. El gorgojo y el endófito de las ryegrass en la producción de leche y carne en el sur de Chile. Uribe, O. Araya, F. Wittwer, A Torres, E. Cisternas, et al.