Jack P. Goodman

Identification of chemically modified peptide from poly(D,L-lactide-co-glycolide) microspheres under in vitro release conditions

The purpose of this research was to study the chemical reactivity of a somatostatin analogue octreotide acetate, formulated in microspheres with polymers of varying molecular weight and co-monomer ratio under in vitro testing conditions. Poly(D,L-lactide-co-glycolide) (PLGA) and poly(D,L-lactide) (PLA) microspheres were prepared by a solvent extraction/evaporation method. The microspheres were characterized for drug load, impurity content, and particle size. Further, the microspheres were subjected to in vitro release testing in acetate buffer (pH 4.0) and phosphate buffered saline (PBS) (pH 7.2). In acetate buffer, 3 microsphere batches composed of low molecular weight PLGA 50∶50, PLGA 85∶15, and PLA polymers (≤10 kDa) showed 100% release with minimal impurity formation (<10%). The high molecular weight PLGA 50∶50 microspheres (28 kDa) displayed only 70% cumulative release in acetate buffer with significant impurity formation (∼24%). In PBS (pH 7.4), on the other hand, only 50% release was observed with the same low molecular weight batches (PLGA 50∶50, PLGA 85∶15, and PLA) with higher percentages of hydrophobic impurity formation (ie, 40%, 26%, and 10%, respectively). In addition, in PBS, the high molecular weight PLGA 50∶50 microspheres showed only 20% drug release with ∼60% mean impurity content. The chemically modified peptide impurities inside microspheres were structurally confirmed through Fourier transform-mass spectrometry (FT-MS) and liquid chromatography/mass spectrometry (LC-MS/MS) analyses after extraction procedures. The adduct compounds were identified as covalently modified conjugates of octreotide with lactic and glycolic acid monomers within polymeric microspheres. The data suggest that due to steric hindrance factors, polymers with greater lactide content were less amenable to the formation of adduct impurities compared with PLGA 50∶50 copolymers.

2,3-Dihydrofarnesoic acid, a unique terpene from trichomes ofLycopersicon hirsutum, repels spider mites

Lycopersicon hirsutum, a wild relative of the tomato, is highly resistant to arthropod herbivores. Both botanic forms ofL. hirsutum, L. hirsutum f.glabratum (C.H. Mull.) andL. hirsutum f.typicum (Humb. & Bonpl.), are resistant to two-spotted spider mites,Tetranychus urticae Koch. However, leaves and trichome secretions from f.typicum repel mites more so than those from f.glabratum. We have previously demonstrated that trichome secretions from LA 1363 and LA 1927, accessions of f.typicum, repelled mites. In this paper we report the identification of the primary component of trichome secretions responsible for repellency. Leaflet washes having compositions similar to trichome secretions were collected and separated into neutral and acid fractions; repellency was mainly associated with the acid fraction, which, when applied to nonrepellent leaflets of f.glabratum, rendered them repellent. Separation of leaflet washes by HPLC allowed purification and subsequent identification by gas chromatography-mass spectrometry and nuclear magnetic resonance of 2,3-dihydrofamesoic acid (3,7,11-trimethyl-6, 10-dodecadienoic acid) as the primary chemical component responsible for repellency. Application of this acid to leaflets ofL. esculentum rendered them repellent. Other volatile compounds present in minor amounts in the acid fractions were farnesoic acid and 16∶0, 16∶3, 18∶0, 18∶2, and 18∶3 fatty acids. This is the first report of the natural occurrence of 2,3-dihydrofarnesoic acid.

Metabolism of the peptide deformylase inhibitor actinonin in tobacco

Abstract Actinonin is a naturally occurring hydroxamic acid and a potent inhibitor of the essential cotranslational protein processing enzyme peptide deformylase. Actinonin has both pre- and post-emergence herbicidal activity, but it is rapidly metabolized by plants, thus limiting herbicidal efficacy. Studies designed to elucidate the metabolic fate of actinonin revealed that after absorption actinonin was metabolized by tobacco plants with only about 17% of the parent compound remaining 48 h after application. Subcellular fractionation revealed that a microsomal fraction was capable of metabolizing actinonin in vitro. Two actinonin metabolites were isolated by reverse-phase high-performance liquid chromatography and identified by mass spectrometric analyses. The major metabolite was derived from the hydrolysis of the hydroxamate group to its corresponding acid, and a relatively minor metabolite through reduction of the hydroxamate group to the corresponding amide. Both metabolites were functionally inact...

Effects of endophyte infected tall fescue seed and red clover isoflavones on rumen microbial populations, fiber fermentation, and volatile fatty acids in vitro

Negative impacts of endophyte-infected Lolium arundinaceum (Darbyshire) (tall fescue) are responsible for over

Effects of red clover isoflavones on tall fescue seed fermentation and microbial populations in vitro

2 billion in losses to livestock producers annually. While the influence of endophyte-infected tall fescue has been studied for decades, mitigation methods have not been clearly elucidated. Isoflavones found in Trifolium pretense (red clover) have been the subject of recent research regarding tall fescue toxicosis mitigation. Therefore, the aim of this study was to determine the effect of ergovaline and red clover isoflavones on rumen microbial populations, fiber degradation, and volatile fatty acids (VFA) in an in vitro system. Using a dose of 1.10 mg × L−1, endophyte-infected or endophyte-free tall fescue seed was added to ANKOM fiber bags with or without 2.19 mg of isoflavones in the form of a control, powder, or pulverized tablet, resulting in a 2 × 3 factorial arrangements of treatments. Measurements of pH, VFA, bacterial taxa, as well as the disappearance of neutral detergent fiber (aNDF), acid detergent fiber (ADF), and crude protein (CP) were taken after 48 h of incubation. aNDF disappearance values were significantly altered by seed type (P = 0.003) and isoflavone treatment (P = 0.005), and ADF disappearance values were significantly different in a seed x isoflavone treatment interaction (P ≤ 0.05). A seed x isoflavone treatment interaction was also observed with respect to CP disappearance (P ≤ 0.05). Seventeen bacterial taxa were significantly altered by seed x isoflavone treatment interaction groups (P ≤ 0.05), six bacterial taxa were increased by isoflavones (P ≤ 0.05), and eleven bacterial taxa were altered by seed type (P ≤ 0.05). Due to the beneficial effect of isoflavones on tall fescue seed fiber degradation, these compounds may be viable options for mitigating fescue toxicosis. Further research should be conducted to determine physiological implications as well as microbiological changes in vivo.

Metabolites of Nuatigenin ((22S,25S)22,25-Epoxy-3β,26-Dihydroxy-Furost-5-Ene) Accumulate in the Bile of Rabbits Fed Oats

Negative impacts of endophyte-infected Lolium arundinaceum (Darbyshire) (tall fescue) are responsible for over

Insulysin Hydrolyzes Amyloid β Peptides to Products That Are Neither Neurotoxic Nor Deposit on Amyloid Plaques

2 billion in losses to livestock producers annually. While the influence of endophyte-infected tall fescue has been studied for decades, mitigation methods have not been clearly elucidated. Isoflavones found in Trifolium pratense (red clover) have been the subject of recent research regarding tall fescue toxicosis mitigation. Therefore, the aim of this study was to determine the effect of ergovaline and red clover isoflavones on rumen microbial populations, fiber degradation, and volatile fatty acids (VFA) in an in vitro system. Using a dose of 1.10 mg × L-1, endophyte-infected or endophyte-free tall fescue seed was added to ANKOM fiber bags with or without 2.19 mg of isoflavones in the form of a control, powder, or pulverized tablet, resulting in a 2 × 3 factorial arrangements of treatments. Measurements of pH, VFA, bacterial taxa, as well as the disappearance of neutral detergent fiber (aNDF), acid detergent fiber (ADF), and crude protein (CP) were taken after 48 h of incubation. aNDF disappearance values were significantly altered by seed type (P = 0.003) and isoflavone treatment (P = 0.005), and ADF disappearance values were significantly different in a seed × isoflavone treatment interaction (P ≤ 0.05). A seed × isoflavone treatment interaction was also observed with respect to CP disappearance (P ≤ 0.05). Eighteen bacterial taxa were significantly altered by seed × isoflavone treatment interaction groups (P ≤ 0.05), eight bacterial taxa were increased by isoflavones (P ≤ 0.05), and ten bacterial taxa were altered by seed type (P ≤ 0.05). Due to the beneficial effect of isoflavones on tall fescue seed fiber degradation, these compounds may be viable options for mitigating fescue toxicosis. Further research should be conducted to determine physiological implications as well as microbiological changes in vivo.

Analysis of the Subsite Specificity of Rat Insulysin Using Fluorogenic Peptide Substrates

In spite of frequent occurrence of steroidal saponins in nutritionally important plants, little is known about metabolism of the corresponding sapogenols. Only relatively recent data indicate that certain saponins are of a considerable health significance to livestock and this toxicity seems to be related to discrete metabolites of sapogenols identified in the bile. It has been noted that lambs grazing on a kleingrass, Panicum coloratum, develop photosensitization secondary to the hepatic dysfunction with lesions, necrosis of hepatocytes and obstruction of small bile ducts with a crystalline material.1 Similar material, accumulating in sheep fed Agave lecheguilla, has been identified as either smilagenin ((25R)3β-hydroxy-5β-spirostane) la or sarsapogenin ((25S)3β-hydroxy-5β-spirostane) 2a.2 Diosgenin ((25R)3β-hydroxyspirost-5-ene) 3 and yamogenin ((25R)3β-hydroxyspirost-5-ene) 4, released from saponins of P. coloratum upon hydrolysis, may give rise to these insoluble products.3 Two related species, P. dichotomiflorum and P. schinzii, are also hepatotoxic to sheep and their intake causes accumulation of a calcium salt of β-D-glucuronide of epi-smilagenin ((25R)3α-hydroxy-5β-spirostane) lb.4,5,6,7 Subsequently, however, the true sapogenol of P. dichotomiflorum has been identified as (25R)-3β,22α,26-trihydroxy-furost-5-ene 5.8 Thus, in case of Panicum, furostanols and not spirostanols may serve as precursors of these bile-insoluble products. In addition, it has been found that in case of the intoxication of sheep grazing on signal grass Brachiaria decumbens, epi-sarsapogenin ((25S)3α-hydroxy-5β-spirostane) 2b and epi-smilagenin lb accumulate in the rumen content,9,10 whereas feeding furostanol saponins of Trigonella foenum graecum to dogs results in the fecal excretion of smilagenin la, diosgenin 3, and gitogenin ((25R)2α, 3β-dihydroxy-5α-spirostane) 6.11