Investigation on the role of nodule cytokinins in regulation of nitrate reductase activity ofPhaseolus mungo (L.)

The effects of some nodular cytokinis, zeatin (Z), zeatin riboside (ZR), N⁶ (Δ²-isopentenyl) adenine (IPA), and N⁶ (Δ²-isopentenyI) adenosine (IPAS) on nitrate reductase (E.C 1.9.6.1) activity of root nodules ofPhaseolus mungo were investigated. The cytokinis were also tested for their effect on nitrate uptake by nodules. The results show that IPAS is the most effective of all the four cytokinins tested. Z and IPA, which caused higherin vivo activity than ZR and IPAS, stimulated uptake of nitrate by nodules. The other two (ZR and IPAS) while inhibiting uptake showed greaterin vitro activity than Z and IPA. It may be concluded that some cytokinins, in addition to their direct effects on the enzyme, may increase the substrate availability to it, whereas others may have only an direct effect on the enzyme activation or degradation.     

On the biogenesis of cytokinins in roots of Phaseolus vulgaris

Roots of intact bean plants were supplied with [¹⁴C]adenine by pulse-chase experiments. The rate of incorporation of radioactivity into tRNA and oligonucleotides of roots as well as the content of radioactive labeled cytokinin nucleotides in these RNA fractions were determined. On the average, 1/70 of the radioactivity incorporated into tRNA was localized in N⁶(²isopentenyl)adenosine. The half life of tRNA was estimated to be 65–70 h. Shortly after the pulse period, oligonucleotides contained zeatin riboside at a ratio of 1:800, on the basis of radioactivity. The half life of these oligonucleotides was determined to be about 8 h. The main free radioactive cytokinin of roots and leaves was zeatin. Comparing the rate of degradation of ¹⁴C-labeled tRNA and the oligonucleotides of roots and the rate of appearance of radioactive cytokinins in roots and leaves, we found strong indications for their dependency. The results contradict the hypothesis of de novo synthesis of cytokinins in roots of intact bean plants.Abbreviations AMP adenosine monophosphate - IPA N⁶(²isopentenyl)adenosine - IPAde N⁶(²isopentenyl)adenosine - Z zeatin - ZR zeatinriboside - TLC thin-layer chromatography - HPLC high performance liquid chromatography Part of the doctoral thesis, Bonn 1980     

Endogenous cytokinins in vegetative shoots of peas

In G2 peas senescence only takes place in long days. In order to determine the role of cytokinins in this process the endogenous cytokinins from vegetative shoots of G2 peas were characterized using gas chromatography-mass spectroscopy following purification by HPLC. Cytokinins were extracted and purified with and without the addition of ¹⁵N labelled internal standards of several cytokinins to estimate cytokin content by isotope dilution in the mass spectra. Samples without internal standards were bioassayed after HPLC. Bioassays showed the presence of zeatin, zeatin riboside and zeatin-0-glucoside. The presence of zeatin was confirmed by its mass spectrum of its permethylated derivative. Tentative identification of zeatin riboside, zeatin-0-glucoside, dihydrozeatin, and dihydrozeatin-0-glucoside was obtained by the coincidence of the major ion for the permethylated natural and ¹⁵N labelled internal standards on GC-MS, and the similar coincidence of ions for permethylated zeatin riboside-0-glucoside by direct probe MS. There was no indication of the presence of significant quantities of zeatin-7-glucoside or zeatin-9-glucoside. The amounts in the tissue ranged from 200–1000 ng/kg fresh weight for each cytokinin and about 2–4 g/kg fresh weight for total cytokinins. There was no apparent difference in the levels in mature but pre-senescent shoots grown in long days and short days indicating that apical senesecence in G2 peas does not appear to be induced by a decline in cytokinin level in the shoots.Cytokinin abbreviations CK Cytokinin - Z trans zeatin - [9R]Z t-zeatin riboside - [9R-5P] Z t-zeatin riboside-5-monophosphate - (OG)Z t-zeatin-0-glucoside - (OG)[9R]Z t-zeatin riboside-0-glucoside - [7Z]G t-zeatin-7-glucoside - [9G]Z t-zeatin-9-glucoside - (diH)Z dihydrozeatin - (diH)[9R]Z dihydrozeatin riboside - iP N6(²-isopentenyl) adenine - [9R]iP N6(²-isopentenyl) adenosine Work performed while PJD was on leave at the University College of Wales at Aberystwyth.     

Mass-spectrometric quantification of cytokinin nucleotides and glycosides in tobacco crown-gall tissue

the cytokinins of tobacco crown-gall tissue have been analysed by quantitative mass spectrometry using ²H₂-labelled cytokinin riboside 5-monophosphates and ¹⁵N₄-labelled cytokinin glycosides as internal standards. The principal endogenous cytokinin of this tissue is zeatin riboside 5-monophosphate. The biologically inactive 7-glucoside of zeatin is the most abundant basic cytokinin in the tissue. These findings expose the limitations of previously reported analyses of similar tissues, which were restricted to biologically active basic cytokinins. The present study demonstrates that the endogenous cytokinins of tobacco crowngall tissue show a clear correspondence to the range of metabolites formed when exogenous cytokinins are supplied to nontumorous tobacco cells.Abbreviations DHZ dihydrozeatin - DHZ7G dihydrozeatin 7-glucoside - DHZMP dihydrozeatin 9-riboside 5-monophosphate - DHZR dihydrozeatin 9-riboside - GC-MS coupled gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - Z7G zeatin 7-glucoside - Z9G zeatin 9-glucoside - ZOG zeatin O-glucoside - ZMP zeatin 9-riboside 5-monophosphate - ZR zeatin 9-riboside - ZROG zeatin 9-riboside O-glucoside     

Identification and quantitative analysis of cytokinins from shoot apices of Mercurialis ambigua by gas chromatography—Mass spectrometry computer system

This paper describes the identification and quantitative analysis of cytokinins from natural sources (150–500 g fresh weight) at the submicrogram level. It summarizes an improved purification procedure with high resolution power that permits the detection of Trimethylsilylderivatives by gas chromatography-mass spectrometry. A comparison of the intensity of a characteristic ion in the mass spectrum of suitable standard (5 g) and theintensity of the same ion in the mass spectrum of the extraction product permitted precise quantitative analysis. The method has been used to determine zeatin, trans- and cis-ribosylzeatin, and ²-isopentenyladenosine concentrations in extracts from female and monoecious Mercurialis ambigua apices. It has been proved that differences appear in the endogenous cytokinin pools of monoecious and female individuals.Abbreviations IPA isopentenyladenosine - Z Zeatin - RZ Ribosylzeatin - GC gas chromatography - TMS Trimethylsilyl     

Cytokinins in maturing and germinatingLupinus luteus L. seeds

³H-labelled zeatin riboside (ZR) was applied to pod walls of intactLupinus luteus L. plants. Metabolites present in mature, dry seeds were zeatin nucleotide (ZNT), zeatin riboside (ZR) and zeatin (Z), zeatin O-glucosides and lupinic acid (LA), and the corresponding dihydro-derivatives of the cytokinins listed. Endogenous cytokinins were rapidly metabolised in germinating seeds. In seeds labelled with [³H]ZR for 90 min following a 2 h period of imbibition in water, ZR was actively converted to ZNT and dihydro-ZNT but the prevailing CTK was Z in cotyledons and ZR in embryo axes (EA); later LA and dihydro-LA, and O-glucoside metabolites accumulated. When [³H] zeatin was introduced into imbibing seeds, it was converted to dihydro-ZNT, ZNT, dihydro-ZR, ZR and dihydro-Z; in EA of the Z-labelled seeds, dihydro-ZR and ZR were the main cytokinins. After incubation of the Z-labelled seeds for 6 h in water, the ratios of dihydro-ZNT: ZNT and dihydro-ZR: ZR were, respectively, 20: 1 and 3.4: 1 in EA, and 3.5: 1 and 1.4: 1 in cotyledons.     

Further studies on the effects of different auxins and cytokinins on flower formation in thin cell layers of Nicotiana tabacum: The effects of zeatin riboside, dihydrozeatin and dihydrozeatin riboside

Organogenesis in thin cell layers of Nicotiana tabacum L. was studied in relation to the effects of natural and synthetic auxins in combination with various cytokinins. All cytokinins tested, benzyladenine (BA), kinetin, zeatin (Z), zeatin riboside (ZR), N⁶-Δ²-isopentenyl) adenine (IPA), dihydrozeatin [(diH)Z] and dihydrozeatin riboside [(diH)ZR], seem to be active in flower bud formation. In addition to the initiation of flower buds, vegetative buds or roots were also formed on the explants in the presence of BA, Z or IPA as exogenous cytokinins. Only dihydrozeatin and its riboside stimulated the initation of flower buds alone (as is known for kinetin), especially if supplemented with indole-3-acetic acid (IAA) as exogenous auxin. A high number of explants with flower buds was also found with high cytokinin/2,4-D ratios. In these conditions the presence of (diH)Z yielded the higest number of flower buds per explant.     

Solid-phase extraction of cytokinins from aqueous solutions with C18 cartridges and their use in a rapid purification procedure

Eleven cytokinins-including bases, ribosides, glucosides, and ribotides-were tested for their retention on C₁₈ cartridges that were washed with 40 mL of water or a dilute acid at pH 3. Cytokinins were then eluted with methanol and analyzed by high performance liquid chromatography (HPLC). All pure cytokinin were well retained when the cartridge was washed with water, but Z and (diH)Z were less well retained at pH 3. The ribotides required 80% methanol for elution. Cotton leaf tissue (500 mg dry wt) was spiked with cytokinins, extracted with 80% methanol, and the extract bulk purified with hexane, insoluble polyvinylpyrrolidone, and minicolumns (strong anion exchange, amino, and C₁₈ cartridges). Ribotides, added to leaf tissue, could not be recovered as ribotides; it was necessary to hydrolyze and purify them as ribosides. The cytokinins were separated and analyzed by HPLC on strong cation exchange and C₁₈ columns. Recoveries through the entire procedure averaged 70%.Cytokinin abbreviations (diH)Z Dihydrozeatin - (diH)Z dihydrozeatin riboside - (diH)[9R]Z trans-zeatin - Z t-zeatin riboside - [9R]Z t-zeatin-O-glucoside - (OG)Z t-zeatin riboside-O-glucoside - (OG)[9R]Z t-zeatin riboside-5-monophosphate - [9R-5P]Z N6(^2-isopentenyl)adenine - iP N6(^2-isopentenyl)adenosine - [9R]iP N6(^2-isopentenyl)adenosine-5-monophosphate-[9R-5P]iP     

Partial auxin deprivation affects endogenous cytokinins in an auxin-dependent,cytokinin-independent tobacco cell strain

The dynamics of individual endogenous cytokinins within the growth cycle (subculture interval) of an auxin-dependent and cytokinin-independent cell suspension culture ofNicotiana tabacum L. (strain VBI-0) were determined using high performance liquid chromatography and radioimmunoassay. In cells grown at an optimum auxin concentration the transient maxima of N⁶-(²-isopentenyl)adenine and N⁶-(²-isopentenyl)-adenosine correlated with the onset of cell division. Cultivation of the cells in a partially auxin-deprived medium resulted in ca. tenfold increase of all endogenous cytokinins. A very distinct maximum of N⁶-(²-sopentenyl) adenine appeared at the beginning of subculture. This indicates that a lack of auxin induced an accumulation of cytokinins predominantly in the form of the free bases, which are physiologically more active than the corresponding ribosides.Abbreviations iP N⁶-(²-isopentenyl)adenine - [9R]iP N⁶-(²-isopentenyl)adenosine - t-Z trans-zeatin - t-[9R]Z trans-zeatin riboside - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - f.w. fresh weight - SBI subculture interval - C complete medium - PAD partially auxin-deprived medium - RP-HPLC reverse phase high performance liquid chromatography - RIA radioimmunoassay - PAL L-phenylalanine ammonia lyase     

Effects of cannabinoids on the activities of mouse brain lipases

Cannabinoids were found to augment phospholipase activities and modify lipid levels of mouse brain synaptosomes, myelin and mitochondria. Delta-1-tetrahydrocannabinol (¹-THC) and several of its metabolites induced a dose-dependent (0.32–16 M) stimulation of phospholipase A₂ (PLA₂) activity resulting in the increased release of free arachidonic acid from exogenous [1-¹⁴C]phosphatidylcholine (PC). The potencies of the cannabinoids in modulating PLA₂ activity were approximately of the order: 7-OH-¹-THC > ¹-THC > 7-oxo-¹-THC > ¹-THC-7oic acid = 6 OH-¹-THC 6-OH-¹-THC. The hydrolysis of phosphatidylinositol (PI) by synaptosomal phospholipase C (PLC) was enhanced significantly by ¹-THC and promoted diacylglyceride levels by greater than 100 percent compared to control values. In contrast, arachidonate was the major product resulting from phospholipase activities of a 20,000g pellet. Synaptosomal diacylglyceride lipase activity was inhibited by ¹-THC. [1-¹⁴C]Arachidonic acid was readily incorporated into subcellular membrane phospholipids and after exposure to cannabinoids led to diminished phosphoglyceride levels and concomitant increases in released neutral lipid products. These data suggest that cannabinoids control phospholipid turnover and metabolism in mouse brain preparations by the activation of phospholipases and, through this mechanism, may exert some of their effects.     

D-Gluconate is an alternative growth substrate for cultivation of Schizosaccharomyces pombe mutants

Schizosaccharomyces pombe cells grow on d-gluconate as the sole carbon and energy source. d-Gluconate is taken up in symport with protons by a specific symporter, pH being the sole driving force. d-Gluconate uptake is independent of the sugar transporting system (e.g. for d-glucose) and of . The carrier is expressed constitutively, and its activity is not subject to glucose repression. Hence, d-gluconate is a suitable carbon and energy source for growth, when d-glucose or other hexoses have to be eliminated e.g. for selection of mutants deficient in hexose transport.Abbreviations 2-DG 2-deoxy-d-glucose - CCCP carbonylcyanide m-chlorophenylhydrazone - pH pH-gradient - electrical potential difference across the plasma membrane - SD standard deviation - SEM standard error of the mean - TPP⁺ tetraphenylphosphonium     

Thylakoid membrane energization and swelling in photoinhibited Chlamydomonas cells is prevented in mutants unable to perform cyclic electron flow

Photoinhibition of Photosystem II in unicellular algae in vivo is accompanied by thylakoid membrane energization and generation of a relatively high pH as demonstrated by ¹⁴C-methylamine uptake in intact cells. Presence of ammonium ions in the medium causes extensive swelling of the thylakoid membranes in photoinhibited Chlamydomonas reinhardtii but not in Scenedesmus obliquus wild type and LF-1 mutant cells. The rise in pH and the related thylakoid swelling do not occur at light intensities which do not induce photoinhibition. The rise in pH and membrane energization are not induced by photoinhibitory light in C. reinhardtii mutant cells possessing an active Photosystem II but lacking cytochrome b₆/f, plastocyanin or Photosystem I activity and thus being unable to perform cyclic electron flow around Photosystem I. In these mutants the light-induced turnover of the D1 protein of Reaction Center II is considerably reduced. The high light-dependent rise in pH is induced in the LF-1 mutant of Scenedesmus which can not oxidize water but otherwise possesses an active Reaction Center II indicating that PS II-linear electron flow activity and reduction of plastoquinone are not required for this process. Based on these results we conclude that photoinhibition of Photosystem II activates cyclic electron flow around Photosystem I which is responsible for the high membrane energization and pH rise in cells exposed to excessive light intensities.Abbreviations cyt b₆/f cytochrome b₆/f - Diuron 3-(3,4-dichlorophenyl)-1 dimethyl urea - Q_B the secondary quinone acceptor of reaction center II - DNP 2,4,Dinitrophenol - FCCP carbonyl cyanide trifluoromethoxy phenylhydrazone - SDS-PAGE sodium dodecylsulfate polyacrylamide gel electrophoresis     

Endogenous hormone concentrations and embryogenic callus development in wheat

Immature zygotic embryos of two wheat (Triticum aestivum L.) genotypes, known for their different ability to generate embryogenic callus, were used as initial explants to establish callus cultures. Embryogenic and non-embryogenic calluses were obtained from the competent genotype (`Combi'), while only non-embryogenic callus was produced by the incompetent one (`Devon'). The morphogenetic competence of each callus type was evaluated by transferring some segments to regeneration conditions. The endogenous hormone concentrations (free indole-3-acetic acid [IAA], abscisic acid [ABA], gibberellins ₁, ₃ and ₂₀ [GAs], zeatin/zeatin riboside [Z/ZR] and N ⁶[²-isopentenyl] adenine/ N ⁶[²-isopentenyl] adenosine; [iP/iPA]) of the initial explants were determined by means of radio-immunoassay and showed that the only difference was the higher concentration of ABA found in the embryos of the most competent genotype; whose embryos showed a reduced rate of precocious germination. When analysing the endogenous hormone concentrations in the various callus types generated in each genotype, it was found that only differences in the free IAA concentrations were associated with variations in the morphogenic properties of the calluses. Higher concentrations of endogenous free IAA were typical of embryogenic callus cultures. It was also observed that a loss in the embryogenic competence of the calluses, due to a prolonged time of culture, occurred concomitantly with a reduction in free IAA concentrations, practically to the concentrations found in the non-embryogenic calluses.     

Sterols in seeds and leaves of oats (Avena sativa L.)

In seeds and leaves of oats (Avena sativa L.) 12 different sterols (cholesterol, cholstanol, ⁷-cholestenol, campesterol, campestanol, stigmasterol, lophenol, sitosterol, stigmastanol, ⁵-avenasterol, ⁷-avenasterol and ⁷-stigmastenol) have been identified. The sterol pattern is qualitatively the same, but the relative composition is different in leaves and in seeds. Leaves contain mainly sitosterol, stigmasterol, cholesterol and campesterol, but only minor portions of avenasterols. Seeds contain sitosterol, ⁵- and ⁷-avenasterol, campesterol, but only minor amounts of stigmasterol and cholesterol. In leaf lipids 1-hexacosanol (2.35 wt % of total lipid) has also been identified.     

Mass-spectrometric quantitation of cytokinins in tobacco crown-gall tumours induced by mutated octopine Ti plasmids of Agrobacterium tumefaciens

The levels of the major cytokinins, zeatin, zeatin riboside, zeatin riboside-5-monophosphate and zeatin-7-glucoside were measured in tobacco (Nicotiana tabacum L.) crown-gall tissues carrying insertion and deletion mutations in the T-DNA. Measurements were made by combined gas chromatography-mass spectrometry using selected ion monitoring with ¹⁵N- and ²H-labelled internal standards. The results demonstrate that, relative to wild-type tumour tissue, cytokinin levels are considerably elevated in tissues lacking functional T-DNA auxin-biosynthetic genes. From a detailed analysis of the major cytokinin metabolites it is concluded that a reduction in the extent of cytokinin degradation via N⁶-side-chain cleavage is an important factor leading to increased cytokinin levels in these tissues.Abbreviations IAA indole-3-acetic acid - SIM selected ion monitoring - Z zeatin - [7G]Z zeatin-7-glucoside - [9R]Z zeatin-9-riboside - [9R-5P]Z zeatin riboside-5-monophosphate     

Effects of competition and N and P supply on carbon isotope discrimination and <Superscript>15</Superscript>N-natural abundance in four grassland species

The effect of interspecific competition and element additions (N and P) on four grassland species (Poa pratensis, Lolium perenne, Festuca valida, Taraxacum officinale) grown under field conditions was studied. Two grasses (L. perenne, F. valida) grown in monoculture (absence of competition) showed lower carbon isotope discrimination (¹³C) and enriched ¹⁵N values. Nitrogen addition (as urea) had inconsistent effects on species ¹³C while caused enrichment of ¹⁵N of P. pratensis and F. valida but strong depletion of ¹⁵N of T. officinale. Phosphorous had no significant effect on ¹³C but depleted ¹⁵N of all species.     

A study on Na+-coupled oxidative phosphorylation: ATP formation supported by artificially imposed ΔpNa and ΔpK inVibrio alginolyticus cells

Addition of Na⁺ to the K⁺-loadedVibrio alginolyticus cells, creating a 250-fold Na⁺ gradient, is shown to induce a transient increase in the intracellular ATP concentration, which is abolished by the Na⁺/H⁺ antiporter, monensin. The pNa-supported ATP synthesis requires an additional driving force supplied by endogenous respiration or, alternatively, by a K⁺ gradient (high [K⁺] inside). In the former case, ATP formation is resistant to the protonophorous uncoupler. Dicyclohexylcarbodiimide and diethylstilbestrol, but not vanadate, completely inhibit Na⁺ pulse-induced ATP formation. The data agree with the assumption that Na⁺-ATP-synthase is involved in oxidative phosphorylation inV. alginolyticus. Interrelation of H⁺ and Na⁺ cycles in bacteria is discussed.Abbreviations and electrochemical gradients of H⁺ and Na⁺, respectively - transmembrane electric potential difference - pH, pNa, and pK concentration gradients of H⁺, Na⁺, and K⁺, respectively - CCCP carbonyl cyanidem-chlorophenylhydrazone - DCCD N,N-dicyclohexylcarbodiimide - DES diesthylstilbestrol - HQNO 2-heptyl-4-hydroxyquinolineN-oxide - Tricine N[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine     

The stability and biological activity of cytokinin metabolites in soybean callus tissue

The activity, uptake and metabolism of cytokinin metabolites was determined in soybean (Glycine max (L.) Merr.) callus tissue. The following activity sequence was established: zeatin riboside (ZR)>zeatin (Z)>O-glucosides of Z, ZR and their dihydro derivatives>lupinic acid (an alanine conjugate of Z)>7- and 9-glucosides of Z which were almost inactive. The 7- and 9-glucosides and lupinic acid were taken up very slowly by the callus tissue and showed great metabolic stability, but some degradation to 7-glucosyladenine, 9-glucosyladenine and the 9-alanine conjugate of adenine occurred. Compared with its aglycone, O-glucosyl-ZR exhibited slow uptake and greatly enhanced stability but gas chromatographic-mass spectrometric analysis showed that appreciable amounts were hydrolyzed to ZR in the tissue. Both ZR and O-glucosyl-ZR were metabolised extensively, with adenine, adenosine, and adenine nucleotide(s) as the major metabolites. A diversity of minor metabolites of ZR were identified, including O-glucosides, lupinic acid and dihydrolupinic acid. The metabolism of ZR was suppressed by 3-isobutyl-1-methylxanthine. When compared with the soybean callus line normally used for cytokinin bioassays (cv. Acme, cotyledonary callus), related callus lines exhibited greatly differing growth responses to cytokinin: however, these were not reflected in marked differences in metabolism.Abbreviations GC-MS gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - LA lupinic acid - OGZR O--D-glucopyranosylzeatin riboside - TLC thin-layer chromatography - IMX 3-isobutyl-1-methylxanthine - Z zeatin - ZR zeatin riboside     

Determination of plant growth substances in liquid endosperm of immature walnut (Juglans nigra) nuts by an ELISA technique

Plant growth substances (PGSs) were analysed in liquid endosperm of black walnut using HPLC and an ELISA procedure. Of all the PGSs studied, we show no GA₃, low levels of cytokinins (io⁶A, i⁶Ade, i⁶Ado) and ABA, and very high level of IAAAbbreviations ABA Abscisic acid: - Ade Adenine: - GA₃ Gibberellic acid: - IAA Indole-3-acetic acid: - i⁶Ade N⁶(²-1) adenine: - i⁶Ado N⁶(²-isopentenyl adenosine: - io⁶A Zeatin riboside:     

The ability of acidic pH,growth inhibitors,and glucose to increase the proton motive force and energy spilling of amino acid-fermenting <Emphasis Type="Italic">Clostridium sporogenes</Emphasis> MD1 cultures

Clostridium sporogenes MD1 grew rapidly with peptides and amino acids as an energy source at pH 6.7. However, the proton motive force (p) was only –25 mV, and protonophores did not inhibit growth. When extracellular pH was decreased with HCl, the chemical gradient of protons (ZpH) and the electrical membrane potential () increased. The p was –125 mV at pH 4.7, even though growth was not observed. At pH 6.7, glucose addition did not cause an increase in growth rate, but increased to –70 mV. Protein synthesis inhibitors also significantly increased . Non-growing, arginine-energized cells had a of –80 mV at pH 6.7 or pH 4.7, but was not detected if the F₁F₀ ATPase was inhibited. Arginine-energized cells initiated growth if other amino acids were added at pH 6.7, and and ATP declined. At pH 4.7, ATP production remained high. However, growth could not be initiated, and neither nor the intracellular ATP concentration declined. Based on these results, it appears that C. sporogenes MD1 does not need a large p to grow, and p appears to serve as a mechanism of ATP dissipation or energy spilling.Mandatory disclaimer: Proprietary or brand names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product, and exclusion of others that may be suitable.