A. Raymond Miller

Cyanidin 3-rutinoside and cyanidin 3-xylosylrutinoside as primary phenolic antioxidants in black raspberry.

Anthocyanin constituents in black raspberries (Rubus occidentalis L.) were investigated by HPLC-DAD, and their involvement as potent, significant antioxidants in black raspberries was demonstrated by three common antioxidant assays (FRAP, DPPH, ABTS) in this study. Five anthocyanins were present in black raspberries: cyanidin 3-sambubioside, cyanidin 3-glucoside, cyanidin 3-xylosylrutinoside, cyanidin 3-rutinoside, and pelargonidin 3-rutinoside. Their identities and structures, with particular emphasis on cyanidin 3-xylosylrutinoside, were confirmed by NMR spectroscopy. Two of these anthocyanins, cyanidin 3-rutinoside and cyanidin 3-xylosylrutinoside, predominated, comprising 24-40 and 49-58%, respectively, of the total anthocyanins in black raspberries. On the basis of both potency and concentration, cyanidin 3-rutinoside and cyanidin 3-xylosylrutinoside were found to be the significant contributors to the antioxidant systems of black raspberries. These findings indicate that these two anthocyanin compounds may function as the primary phenolic antioxidants in black raspberries. These two compounds exhibit potential biological activities that may be exploited in conjunction with other naturally occurring bioactive compounds in black raspberry fruit-based products used in clinical trials for the treatment of various types of cancer.

Culture medium pH is influenced by basal medium, carbohydrate source, gelling agent, activated charcoal, and medium storage method

SummaryWhen four carbohydrates were tested against six commonly cited inorganic basal media, post-autoclave pH was highest for carbohydrate-free and sucrose containing media, and progressively lower for maltoseglucose and fructose-containing media, respectively. Post-autoclave pH for these media without carbohydrates was related to medium buffering capacity. Addition of gelling agents (10 of 11 tested) increased the postautoclave pH of MS medium containing sucrose. Neutralized and acid-washed activated charcoal also increased the post-autoclave pH of liquid and agarsolidified MS medium, and the pH changed further during 8 weeks of storage. Changes in medium pH caused by gelling agents, but not charcoal, could be alleviated by adjusting the pH after their addition but prior to autoclaving.

Oxidation of cell wall polysaccharides by hydrogen peroxide: A potential mechanism for cell wall breakdown in plants

Incubation of cellulose, sodium carboxylcellulose, pectin, polygalacturonic acid, xylan and arabinogalactan with hydrogen peroxide (0.1-10 mM) resulted in rapid breakdown of the polysaccharides when measured by a reduction of solution viscosity or an increase in reducing groups. When the reaction mixtures were precipitated with ethanol or fractionated on G-25-300 Sephadex, low molecular weight reducing groups increased with incubation time indicating that polymer cleavage was occurring and not simply polymer modification. Oxidation was most rapid at pH 6.5 or 7.5, although secondary optima between pH 3.5 and 5.5 were also observed, depending on the polysaccharide. Purified cell walls isolated from various organs of tomato, cucumber and soybean were similarly degraded and the ethanol-soluble reaction products were partially characterized. The data support the hypothesis that hydrogen peroxide generated by peroxidase from NADH may play a role during cell wall breakdown in plants.

Effects of tomato cultivar, leaf age, and bacterial strain on transformation by Agrobacterium tumefaciens

Differences in transformation of the tomato cultivar (Ohio 7870, Roma, UCD82b) by wild-type Agrobacterium strains (A6, A66, A281) were identified in a leaf disk assay system. Transformation was expressed as the percentage of explants producing callus on hormone-free medium and was confirmed by opine production. Ohio 7870 and Roma were more readily transformed than UCD82b by all three strains of A. tumefaciens. Cotyledons and older true leaves of all three cultivars were more readily transformed than younger leaves. Transformation was biphasic over the bacterial concentrations tested (2×103–7×109 colony forming units ml-1; cfu ml-1) for all cultivars and leaf ages, and was greatest at 5×108 cfu ml-1. Transformation decreased significantly at levels less than 2×107 cfu ml-1 and slightly at concentrations higher than 5×108 cfu ml-1. UCD82b tissue was more necrotic than Ohio 7870 or Roma after incubation with bacteria, which may account for reduced transformation of this cultivar.

Anodic peroxidase isoenzymes and polyphenol oxidase activity from cucumber fruit: Tissue and substrate specificity

Abstract Peroxidase activity in fresh cucumber fruit was highest in the skin; followed by pericarp then carpel tissues, respectively. Polyphenol oxidase activity, by contrast, was present only in the skin. In crude extracts, both activities had pH optima near 7.0, but exhibited different temperature optima and thermostability. Using hydrogen peroxide with either benzidine, guaiacol, p-phenylenediamine, o-phenylenediamine, o-dianisidine, or 3-amino-9-ethylcarbazole as substrates, more anodic peroxidase isoenzymes were observed in the skin than the pericarp or carpel. The reactivity of the peroxidase isoenzymes toward these stains varied qualitatively and quantitatively, e.g. benzidine stained more and different isoenzymes with less relative intensity than did p-phenylenediamine. Further, no staining occurred with the above substrates in the absence of hydrogen peroxide, or with Fast Blue BB base, caffeic acid, syringaldazine or polyphenol oxidase substrates (catechol, hydroquinone, L-DOPA), regardless of hydrogen peroxide availability. These data show that cucumber fruit anodic peroxidase isoenzymes vary widely in substrate specificity. Further, the high thermostability of cucumber peroxidase and polyphenol oxidase activity suggests that these enzymes may play a role in the darkening of processed cucumber products.

Haploid plant regeneration from anther cultures of three north american cultivars of strawberry (Fragaria x ananassa Duch.)

SummaryA study was conducted to maximize plant regeneration frequencies from cultured anthers of ‘Chandler’, ‘Honeoye’, and ‘Redchief’ strawberries (Fragaria x ananassa Duch.). A comparison of auxins (IAA, NAA), cytokinins (BA, BPA, KIN) and carbohydrates (sucrose, glucose, maltose) in MS medium showed that the highest shoot regeneration across cultivars (8%) occurred when using a medium containing 2 mg/l IAA, 1 mg/l BA, and 0.2 M glucose. A comparison of MS, NN, and H1 inorganic medium (a new formulation based on the anther culture literature) solidified with either agar or gellan gum and containing IAA, BA, and glucose, showed the highest shoot regeneration across cultivars (19%) when using H1 and gellan gum. Lastly, media containing Fe-EDTA yielded more shoots than media containing Fe-Metalosate, and anthers cultured on Fe-EDTA media in darkness for 30d followed by 30d in white light produced more shoots (16% average regeneration) than those cultured on Fe-EDTA media under white or yellow light (16h photoperiod) for the initial 30d (0.3% and 5% respectively). Plants were acclimated ex vitro where they flowered and set fruit. Chromosome counts of root tip cells confirmed that haploid plants were obtained from all three cultivars.

NMR-based metabolomic investigation of bioactivity of chemical constituents in black raspberry (Rubus occidentalis L.) fruit extracts.

Black raspberry (Rubus occidentalis L.) (BR) fruit extracts with differing compound profiles have shown variable antiproliferative activities against HT-29 colon cancer cell lines. This study used partial least-squares (PLS) regression analysis to develop a high-resolution (1)H NMR-based multivariate statistical model for discerning the biological activity of BR constituents. This model identified specific bioactive compounds and ascertained their relative contribution against cancer cell proliferation. Cyanidin 3-rutinoside and cyanidin 3-xylosylrutinoside were the predominant contributors to the extract bioactivity, but salicylic acid derivatives (e.g., salicylic acid glucosyl ester), quercetin 3-glucoside, quercetin 3-rutinoside, p-coumaric acid, epicatechin, methyl ellagic acid derivatives (e.g., methyl ellagic acetyl pentose), and citric acid derivatives also contributed significantly to the antiproliferative activity of the berry extracts. This approach enabled the identification of new bioactive components in BR fruits and demonstrates the utility of the method for assessing chemopreventive compounds in foods and food products.

An examination and correction of plant tissue culture basal medium formulations

The inorganic formulations of fourteen common plant tissue culture basal media were examined from the primary literature. Inaccuracies and errors were found for molecular formulae, chemical hydrations, and molar equivalences for iron/EDTA complexation. A comparison with published basal medium formulations from six commercial suppliers uncovered additional inaccuracies, modifications, and errors, thereby emphasizing the need for investigators to examine and describe medium formulations precisely in future publications.

Mechanical stress-induced changes in sugar composition of cell walls from cucumber fruit tissues

Abstract The neutral sugar composition, uronic acid content, esterified methanol content and percent uronic acid methylation of cell walls from the mesocarp and carpel of mechanically stressed and non-stressed (control) 14-day-old cucumber fruit were determined. Forty-eight hr after treatment, mesocarp cell walls from mechanically stressed fruit had significantly less mannose (77% decrease) and galactose (49% decrease) than control fruit stored for the same time, while rhamnose, arabinose, xylose and glucose did not vary appreciably. Concurrently, the per cent uronic acid methylation of mesocarp cell walls did not change following mechanical stress, but increased uronic acid content led to increased uronic acid: neutral pectic sugar ratios. By contrast, carpel cell walls from mechanically stressed fruit 48 hr after treatment exhibited increased uronic acid content, esterified methanol content and percent uronic acid methylation (11, 44 and 33%, respectively) compared to control fruit. The neutral sugar composition of carpel cell walls did not change in response to mechanical stress, although xylose and mannose increased ca two-fold during storage. Thus, mechanical stress caused significant cell wall changes in cucumber fruit tissues, but these changes occurred by different mechanisms in the mesocarp and carpel.