A new method for electrophysiological identification of antennal pH receptor cells in ground beetles: the example of Pterostichus aethiops (Panzer, 1796) (Coleoptera, Carabidae)

The responses of antennal taste sensilla of the ground beetle Pterostichus aethiops to 100mM Na(+)-salts and their mixtures with 1 and 10mM NaOH were compared. An increase in pH by 0.3-0.6 units in 100mM Na(+)-salt solutions, caused by the content of 1mM NaOH, was too small, except for alkaline Na(2)HPO(4), to influence the firing rate of the cation cell and pH cell significantly. However, different sensitivity of the two cells to increased pH was clearly demonstrated when the concentration of NaOH in 100mM stimulating salt solutions was increased to 10mM. Increasing pH by 1.2-2 units caused the 1st s firing rate to increase by 140-1050% and 0-26% in the pH cell and cation cell, respectively. Compared to the buffer series method used for identification of the pH receptors in ground beetles earlier, considerably stronger responses of the pH cell to a similar increase in pH were observed when the NaOH method was used for testing. At the same time, undesirable changes in salt ions concentration that occur when stimulating solutions differing by 1-2 pH units are prepared were much smaller using the latter method. Behavioural and ecological relevance of the results is discussed.     

Electrophysiological identification of the sugar cell in antennal taste sensilla of the predatory ground beetle Pterostichus aethiops

By single sensillum tip recording technique, in addition to the salt and pH cells found in antennal taste sensilla of some ground beetles earlier, the third chemosensory cell of four innervating these large sensilla was electrophysiologically identified as a sugar cell in the ground beetle Pterostichus aethiops. This cell generated action potentials of considerably smaller amplitude than those of the salt and pH cells, and phasic-tonically responded to sucrose and glucose over the range of 1-1000 mM tested. Responses were concentration dependent, with sucrose generating more spikes than glucose. During the first second of the response, maximum rates of firing of the sugar cell reached up to 19 and 37 imp/s when stimulated with 1000 mM glucose and sucrose, respectively. Three to four seconds later, the responses decreased close to zero. Both sugars are important in plant carbohydrate metabolism. These ground dwelling insects may come into contact with live and decayed plant material everywhere in their habitat including their preferred overwintering sites in brown-rot decayed wood. In conclusion, we hypothesize that high content of soluble sugars in their overwintering sites and refugia is unfavourable for these ground beetles, most probably to avoid contact with dangerous fungi.     

Electrophysiological responses of the chemoreceptor neurones in the antennal taste sensilla to plant alkaloids and glucosides in a granivorous ground beetle

The electrophysiological response of chemoreceptor neurones from the antennal chaetoid taste sensilla of the omnivorous ground beetle Pterostichus oblongopunctatus to several plant alkaloids and glucosides is investigated. A quinine‐sensitive neurone responding to quinine and quinine hydrochloride is found, most probably related to the granivorous feeding habit of P. oblongopunctatus. The response to quinine hydrochloride is concentration‐dependent at 0.001–50 mm , with the response threshold at 0.01 mm and a maximum rate of firing of 67 spikes/s at 50 mm . The stimulatory effect of caffeine is very weak, where the firing rate increases by only 1.4 spikes/s at a concentration of 10 mm compared with that evoked by a control stimulus. In addition, both quinine and quinine hydrochloride strongly inhibit spike production by the salt‐ and pH‐sensitive neurones when presented in mixtures with 10 mm NaCl. Several tested plant secondary compounds (i.e. salicin, sinigrin, caffeine and nicotine), which have only little or no effect on the firing rate of the quinine‐sensitive neurone, greatly reduce the responses of the salt‐ and pH‐sensitive neurones. The results of the present study suggest that the antennal taste sensilla of P. oblongopunctatus may detect plant defensive compounds both through the activation of a quinine‐sensitive neurone and via peripheral inhibition of other chemoreceptor neurones of the taste sensillum.     

The K+-H+ exchanger, nigericin, modulates taste cell pH and chorda tympani taste nerve responses to acidic stimuli

The relationship between acidic pH, taste cell pH(i), and chorda tympani (CT) nerve responses was investigated before and after incorporating the K(+)-H(+) exchanger, nigericin, in the apical membrane of taste cells. CT responses were recorded in anesthetized rats in vivo, and changes in pH(i) were monitored in polarized fungiform taste cells in vitro. Under control conditions, stimulating the tongue with 0.15 M potassium phosphate (KP) or 0.15 M sodium phosphate (NaP) buffers of pHs between 8.0 and 4.6, KP or NaP buffers did not elicit a CT response. Post-nigericin (500 × 10(-6) M), KP buffers, but not NaP buffers, induced CT responses at pHs ≤ 6.6. The effect of nigericin was reversed by the topical lingual application of carbonyl cyanide 3-chloro-phenylhydrazone, a protonophore. Post-nigericin (150 × 10(-6) M), KP buffers induced a greater decrease in taste cell pH(i) relative to NaP buffers and to NaP and KP buffers under control conditions. A decrease in pH(i) to about 6.9 induced by KP buffers was sufficient to elicit a CT response. The results suggest that facilitating apical H(+) entry via nigericin decreases taste cell pH(i) and demonstrates directly a strong correlation between pH(i) and the magnitude of the CT response.     

Electrophysiological responses to salts from antennal chaetoid taste sensilla of the ground beetle Pterostichus aethiops

Antennal gustatory sensilla of the ground beetle Pterostichus aethiops (Pz., 1797) (Coleoptera, Carabidae) respond to salts, the three sensory cells, A-, B- and C-cells, producing action potentials that are distinguished by differences in their shape, amplitude, duration and polarity of spikes. The B-cell (salt cell) was highly sensitive to both ionic composition and concentration of the tested nine salt solutions showing phasic-tonic type of reaction with a pronounced phasic component. The stimulating effect was dominated by the cations involved, and in most cases, monovalent cations were more effective stimuli than divalent cations. Salt concentration/response relations were tested with NaCl at 1, 10, 100 and 1000 mmol l⁻¹: mean firing rates increased from 0.8 to 44 spikes per first second of the response, respectively. The pH value of the stimulating solutions also influenced the B-cell rate of firing. By contrast, the pH level of stimulus solutions influenced the A-cells’ phasic-tonic response more than the ionic composition or concentration of these solutions. Compared to a standard 100 mmol l⁻¹ salt (NaCl) solution (pH 6.3), alkaline solutions of the salts NaCH₃COO, Na₂HPO₄ and Na₂B₄O₇ (pH 7.9, 8.5 and 9.3, respectively, all 100 mmol l⁻¹) induced remarkably stronger responses in the A-cell. On the other hand, the reaction to an acid solution of NaH₂PO₄ (pH 4.5, 100 mmol l⁻¹) was minimal. A-cell responses to neutral salts like NaCl, KCl, CaCl₂, MgCl₂ and C₅H₁₄NOCl (pH 6.1-6.5) varied largely in strength. Very low or no responses were observed with chlorides of divalent cations, CaCl₂ and MgCl₂, and choline chloride (C₅H₁₄NOCl), indicating that the ionic composition of the solutions also affected A-cell responses. Neural activity of the C-cell was not influenced by the salt solutions tested.     

Resealing to small solutes of white erythrocyte membranes after incubation with EDTA, Ca2+, salt, sucrose, phospholipase C

White, stable erythrocyte ghosts can recover their impermeability to small solutes after storage for several days in low-ionic-strength phosphate buffers at 0 °C. The accessibility, to their substrates, of the inner surface enzymes, glyceraldehyde-3-phosphate dehydrogenase, (G3PD), and NADH cytochrome c oxidoreductase, was used to assess resealing. The data from the two enzymes were confirmatory. None of the conditions used to investigate resealing altered the activity of the outer surface enzyme, acetylcholinesterase. Using G3PD activity, ghosts (freshly prepared by gentle stepwise hemolysis in hypotonic phosphate buffers and stored in 11 mm phosphate buffer, pH 7.4) were shown to be slightly sealed (33%). Incubation at 37 °C in the storage buffer with or without EDTA did not alter their permeability. Ionic strength rather than osmotic pressure appears to influence the sealing process since salt (286 mosm) elicited 91% sealing whereas sucrose (278 mosm) had little effect. Calcium in trace amounts caused resealing to 80%. Phospholipase C (C. welchii) completely abolished Ca²⁺-induced resealing. The data were highly reproducible although these ghosts were found to contain only 10 to 20% of the G3PD activity of the leaky ghosts prepared by shock hemolysis in 5 mm phosphate buffer, pH 8.0. The response to the resealing agents was similar regardless of the level of G3PD present. Neither calcium nor ETDA altered the chemical composition (sialic acid, cholesterol, phospholipid) of the membranes. The small amount (5%) of nonspecific loosely bound protein lost during incubation, could not be attributed to any of the test agents. The results suggest that calcium induced the recovery of impermeability by altering the association, distribution, and/or conformation of the proteins and phospholipids within the membrane.     

Chain length-dependent cooperativity in fatty acid binding and oxidation by cytochrome P450BM3 (CYP102A1)

Fatty acid binding and oxidation kinetics for wild type P450_BM3 (CYP102A1) from Bacillus megaterium have been found to display chain length-dependent homotropic behavior. Laurate and 13-methyl-myristate display Michaelis-Menten behavior while there are slight deviations with myristate at low ionic strengths. Palmitate shows Michaelis-Menten kinetics and hyperbolic binding behavior in 100 mmol/L phosphate, pH 7.4, but sigmoidal kinetics (with an apparent intercept) in low ionic strength buffers and at physiological phosphate concentrations. In low ionic strength buffers both the heme domain and the full-length enzyme show complex palmitate binding behavior that indicates a minimum of four fatty acid binding sites, with high cooperativity for the binding of the fourth palmitate molecule, and the full-length enzyme showing tighter palmitate binding than the heme domain. The first flavin-to-heme electron transfer is faster for laurate, myristate and palmitate in 100 mmol/L phosphate than in 50 mmol/L Tris (pH 7.4), yet each substrate induces similar high-spin heme content. For palmitate in low phosphate buffer concentrations, the rate constant of the first electron transfer is much larger than k_cat. The results suggest that phosphate has a specific effect in promoting the first electron transfer step, and that P450_BM3 could modulate Bacillus membrane morphology and fluidity via palmitate oxidation in response to the external phosphate concentration.     

The Donnan effect in tobacco mosaic virus and its components

Osmotic pressure studies were carried on tobacco mosaic virus (TMV) and its components, protein and RNA, as well as on bis(3,3′-aminopropyl)amine, reported to be present in TMV preparations. Solvents were phosphate and barbital buffers at different values of pH and ionic strength. Measurements were made at room temperature. The Donnan effect was exhibited by TMV protein in phosphate buffer of 0.01 ionic strength at pH values ranging between 5.8 and 7.5. The observed values of the Donnan effect at pH 5.8 and 5.97 were in reasonable agreement with theoretical values calculated from the charge obtained by hydrogen ion titration. TMV-RNA in phosphate buffer at pH 7.5 and ionic strength 0.01 did not exhibit more than 1% of the expected Donnan effect. This is explained tentatively as the result of firm binding of metal ions. Negative values of osmotic pressure were observed with bis(3,3′-aminopropyl)amine. Similar anomalous osmosis was sometimes observed with TMV protein and with TMV. In agreement with earlier observations, TMV did not exhibit the Donnan effect in phosphate buffer of 0.01 ionic strength at pH values ranging from 5.5 to 8.0. However, TMV dialysed extensively in the presence of EDTA at pH 8.5 and TMV produced by reconstitution of purified protein and RNA did exhibit the Donnan effect in both phosphate and barbital buffers. The magnitude was of the same order as that calculated from the net charge determined by hydrogen ion titration. When reconstituted TMV, which did exhibit Donnan effect, was treated with calcium ions, the effect was abolished.     

Inactivation of viruses using novel protein A wash buffers

Low pH viral inactivation is typically performed in the eluate pool following the protein A capture step during the manufacturing of monoclonal antibodies and Fc‐fusion proteins. However, exposure to low pH has the potential to alter protein quality. To avoid these difficulties, novel wash buffers capable of inactivating viruses while antibodies or Fc‐fusion proteins were bound to protein A or mixed mode resins were developed. By equilibrating the column in high salt buffer (2 M ammonium sulfate or 3 M sodium chloride) after loading, the hydrophobic interactions between antibodies and protein A ligands were increased enough to prevent elution at pH 3. The ammonium sulfate was also found to cause binding of an antibody to a mixed mode cation exchange and a mixed mode anion exchange resin at pH values that caused elution in conventional cation and anion exchange resins (pH 3.5 for Capto Adhere and pH 8.0 for Capto MMC), indicating that retention was due to enhanced hydrophobic interactions. The potential of the 2 M ammonium sulfate pH 3 buffer, a 1 M arginine buffer, and a buffer containing the detergent LDAO to inactivate XMuLV virus when used as protein A wash buffers with a 1 hour contact time were studied. The high salt and detergent containing wash buffers provided about five logs of removal, determined using PCR, and complete combined removal and inactivation (> 6 logs), determined by measuring infectivity. The novel protein A washes could provide more rapid, automated viral inactivation steps with lower pool conductivities. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:406–413, 2015     

Sensitivity of antennal gustatory receptor neurones to aphid honeydew sugars in the carabid Anchomenus dorsalis

Using electrophysiology, the stimulating effect of 13 sugars and three sugar alcohols (each at a concentration of 100 mm ) to antennal gustatory receptor neurones (GRNs) is tested in the carabid beetle Anchomenus dorsalis (Pontoppidan, 1763) (Coleoptera, Carabidae). Maltose, sucrose, glucose and raffinose are the most stimulating sugars for the sugar‐sensitive neurone (SuN), evoking 6.7–18.6 spikes s^?1 in fed insects, whereas the others had little or no effect. The firing rate of the antennal GRNs is not affected by any of the tested sugar alcohols, dulcitol, inositol and sorbitol. Additionally, concentration/response curves for sucrose and maltose are obtained in the range 0.01–100 mm . The responses of beetles starved for 96 h to this range of sucrose are two‐ to three‐fold higher compared with those of fed beetles. The presence of a terminal α‐glucose unit is an important feature of the molecular structure determining the stimulating properties of the two disaccharides, maltose and sucrose, as well as glucose. The other monosaccharide unit of the molecule is also of great importance in determining the stimulating properties of various disaccharides. The sensitivity of the SuN to the four most prevalent aphid honeydew sugars suggests that A. dorsalis uses these chemicals as sensory cues when searching for aphids as prey.     

Increased visualization of microtubules by an improved fixation procedure.

We have found that when a buffer utilized for in vitro polymerization of microtubules, i.e., 1 mM guanosine triphosphate, 1 mM MgSO4, 2 mM ethylene glycol bis(beta-aminoethyl ether)-N, N'-tetraacetic acid 100 mM piperazine-N,N'-bis(2-ethanesulfonic acid), pH 6.9 polymerization mix, was used in the glutaraldehyde prefixation regimen instead of classical fixative buffers, i.e., isotonic cacodylate or phosphate buffer, the following features were observed in thin-sections of the cytoplasm of interphase HeLa cells: (a) a greater than 2-fold increase in total microtubule contour length, (b) a 2-fold increase in a number of microtubules greater than or equal to 1 mu long, (c) an enhanced association of microtubules with cytoplasmic organelles, and (d) an increased clustering of 100 A filaments located in a perinuclear region of the cell. Furthermore, we found that after we incubated purified chick brain microtubules on a Sephadex G-25 column pre-equilibrated with polymerization mix, cacodylate or phosphate buffer at 37 degrees C, and then eluted the microtubules at 37 degrees C, the exposure to cacodylate or phosphate buffer caused extensive depolymerization, but exposure to polymerization mix buffer allowed reisolation of highly polymerized microtubules. Our results imply that prefixation with cacodylate or phosphate buffered glutaraldenyde destabilizes microtubules leading to the decreased visualization of microtubules.     

Ion-dependence of Z-line and M-line response to calcium in striated muscle fibres in rigor.

The calcium-dependent contraction of vertebrate skeletal muscle is thought to be primarily controlled through the interaction of the thick and thin filaments. Through measurement of the Donnan potential, we have shown that an electrical switching mechanism (sensitive to both anions and cations) is present in both A- and I-bands [1]. Here we show that this mechanism is not confined to the contractile apparatus and report for the first time the presence of M-line potentials. The Z-line responds to Ca2+ ions in a similar manner to the A-band under the same solution conditions (phosphate-chloride and imidazole buffers), even though it has no reported Ca2+ binding sites. Z-line potentials were not observed in tris-acetate buffer. The M-line has a markedly different response to any of the other subsarcomeric regions, however, and can only be detected in the phosphate-chloride buffer. Preliminary observations of the M-line potential in creatine kinase-deficient mouse muscle (phosphate-chloride buffer) reveal significant differences in the calcium-induced transitions between two of the genotypes and demonstrate definitively that it is the M-line potential that is being recorded. From these results, it seems likely that the charge response of the Z-line and M-line is being mediated by titin in an anion-dependent manner. Our evidence comes from several observations. First, the similarity between the response of the Z-line potentials to the A-band potentials, where titin is the only link between these structures and second, the differential observation of M-line and Z-line potentials in a range of buffers containing different anion(s). Both Z-line and M-line potentials were seen in phosphate-chloride buffer, but only the Z-line potentials could be detected in chloride-only (imidazole) buffer and neither was observed in the acetate buffer. The latter observations can be attributed to two sources. The first is the effect of acetate buffer on the conformation of myosin [2]; the second is the absence of binding of the M-line protein, myomesin, to titin in the absence of phosphate ions [3].     

Coating efficiency of a 1 -acid glycoprotein in competitive enzyme immunosorbent assays with antigen immobilized on the solid phase

Coating efficiency of rat and human serum a 1 -acid glycoprotein (a 1 -AGP) was investigated for competitive enzyme immunosorbent assays with antigen immobilized on the solid phase by using different pHs and buffers. Blocking materials and pH of coating buffer had a marked influence on the amount of a 1 -AGP that binds to plate. Usually, carbonate buffer is used at pH 9.6 or 9.0, but phosphate buffered saline (PBS) at pH 7.2 can be used for an effective coating. At pH 7.2, coating of a 1 -AGP in Tris buffered saline was five - tenfold as effective as in PBS and phosphate buffer. Blocking of uncoated surface with casein was ten - twenty times as effective as with fetal bovine serum albumin for coating of a 1 -AGP.     

Effect of ethanesulfonic acid buffers and pH on the accumulation of a nervous system-specific protein (S-100) and a glial-enriched enzyme in a clonal line of rat astrocytes (C6).

Stationary phase cultures of a clonal line of rat astrocytes (C6) were maintained at pH values ranging from 6.0 to 8.4 using media buffered with various combinations of organic buffers or graded concentrations of bicarbonate ion at a constant CO2 tension. The accumulation of a soluble acidic protein unique to the nervous system (S-100) in media buffered with organic buffers was optimal in the pH range 6.4 to 6.8, significantly more acid than that optimal for cell growth (pH 7.0 to 7.8). Cells maintained in CO2-bicarbonate-buffered media exhibited a higher and less marked pH optimum for S-100 protein accumulation and a lower efficiency of accumulation of the protein. These data suggest that the organic buffer ions themselves, apart from their function as buffers, are influencing the accumulation of S-100. The specific activity (assayed at the enzymatic pH optimum) of a membrane-bound enzyme enriched in glial cells and myelin, 2',3'-cyclic nucleotide 3'-phosphohydrolase, was markedly pH-dependent. The optimal pH range was 6.4 to 6.7 in organic buffer controlled media. In CO2-bicarbonate controlled media the optimal pH range was only slightly higher (pH 6.6 to 7.0), but the specific activities were reduced relative to organic buffer-grown cells. The structural relationship of some of the aminoethanesulfonic acid buffers used in these experiments to certain compounds of neurochemical interest (such as taurine and alpha-flupenthixol) is noted.     

Excitability of single firing human motoneurones to single and repetitive stimulation (experiment and model)

The activity of single motoneurones of m. flexor carpi ulnaris (FCU) was investigated by recording their motor unit (MU) action potentials during weak and moderate voluntary muscle contractions. The MU firing rate range was 4.5-15 imp/s. The excitability of motoneurones was tested with a number of single stimuli eliciting a monosynaptic H-reflex of low amplitude. Two different indices were defined which relate to motoneuronal excitability: the response index--the ratio of the number of responses of a motoneurone to the total number of stimuli, and the response time--the time after the last background MU discharge at which motoneurone is ready to respond to the excitatory volley. Both the response index and the response time were determined for single motoneurones at different levels of background activity. In the lower range of firing rates, the response index for all motoneurones decreased when increasing the firing rate, but it remained constant in the higher rate range. This kind of response seems to be a typical motoneuronal response to the stimulation with single stimuli. The data on the response time were used to study the excitability of the same single motoneurones to computer simulated repetitive stimulation (stimulation rate 40-100 imp/s). In this case, the excitability of each motoneurone was determined as an increment of its firing rate in response to the stimulation. For the lower firing rate range, the excitability for all motoneurones also decreased when the firing rates increased whereas a variety of slopes was obtained in the higher rate range.(ABSTRACT TRUNCATED AT 250 WORDS)     

Factors modulating the blood feeding behavior and the electrophysiological responses of labral apical chemoreceptors to adenine nucleotides in the mosquito Aedes aegypti (Culicidae)

The feeding of Aedes aegypti (L.) on blood is induced by the presence of phagostimulants: adenine nucleotides. Three chemoreceptive cells in the labral apical sensilla can distinguish the presence of adenine nucleotides depending on the other stimulus components. This work aims at correlating the sensory information arising from the labral apical sensilla with the feeding behavior in response to the same stimuli. The saline stimulating solution, containing adenine nucleotides, is modulated by changing one of the following components: salt concentration, buffer or pH. Cell 3 that responds to NaCl in a dose dependent manner seems to have another unique modality. The response of this cell is unaffected by ATP when the stimulating solution is NaCl buffered by NaHCO(3). It responds at a higher spike frequency to the presence of ATP in a NaCl solution without NaHCO(3). Thus in the presence of ATP Cell 3 detects whether the NaCl solution is buffered by NaHCO(3). Both the blood feeding response and the sensory information from Cell 2 (which responds at high spike frequencies to the presence of ATP) are modulated by pH in a similar way. Both responses present a bi-modal response, with a major peak at pH 4.0 and a moderate peak at the most alkaline pH value tested.     

Selective removal of histone H1 from nucleosomes at low ionic strength

The method for removal of histone H 1 from chromatin by treatment with ion-exchange resin AG 50 WX 2 in the presence of 100 mM NaCl and 50 mM phosphate buffer (Thoma and Koller, 1977, Cell, 12, 101–107) results in production not only of H1-depleted chromatin but also free DNA. We have now modified this procedure so that the nucleosome is treated with the cation exchange resin in two steps, first in 50 mM sodium phosphate buffer and then in 50 mM sodium phosphate and 50 mM NaCl whereby histone H 1 is selectively removed without a release of free DNA at low resin concentrations.Abbreviations NaP Sodium phosphate buffer of molarities and pH as stated in the text - SDS Sodium dodecyl sulfate     

Baroresistant buffer mixtures for biochemical analyses

Hydrostatic pressure is a useful tool in the study of varied fields such as protein aggregation, association, folding, ligand binding, and allostery. Application of pressure can have a significant effect on the pK(a) values of buffers commonly used for biochemical analysis. Consequently, cationic buffers, rather than neutral ones, are generally used to minimize pH effects; however, even with these buffers, the change in pH over 3 kbar may be consequential in highly pH-sensitive biochemical systems. Using fluorescence-based assays, we have systematically examined the effects of pressure on various buffers in the neutral pH range. We show that many commonly used cationic and Good's buffers increase in pH with pressure on the order of 0.1 to 0.3 pH units/kbar, in agreement with other published values. Carboxylates and phosphate decrease in pH to a similar extent. Buffer mixtures, composed of both cationic and carboxylate or phosphate components, are shown to be an order of magnitude less pressure sensitive than the individual component buffers. Using various relative concentrations of Tris and either phosphate, tricarballylate (1,2,3-propanetricarboxylate), or CDA (1,1-cyclohexane diacetate) at pH values between 7 and 8 yields baroresistant buffer mixtures. Buffer mixtures can be optimized for a specific pH, and a list of mixtures is presented for general laboratory use.     

Effect of hydrogen ion buffers on photosynthetic oxygen evolution in the blue-green alga, Agmenellum quadruplicatum.

The photosynthetic oxygen evolution capacity of Agmenelium quadruplication suspended in four hydrogen ion buffers (pH 7.4, 0.05 M) and its synthetic marine growth medium was measured with an oxygen electrode. High rates of oxygen evolution were obtained in the growth medium and N-tris(hydroxymethyl)-methylglycine (Tricine) buffer. Compared to oxygen evolution in the growth medium, rates in phosphate buffer and N-tris(hydroxymethyl)-2-aminoethanesulphonic acid (TES) buffer were sometimes reduced by up to 30% and rates in tris (hydroxymethyl) amino-methane (Tris) were consistently reduced by 50%. An incubation-rinsing procedure caused inhibition of oxygen evolution in TES, phosphate, and Tris by 50 to 100%. Oxygen evolution could be restored to cells rinsed in TES or phosphate by resuspension in growth medium or in buffer plus magnesium and calcium ions. Bezoquinone-supported oxygen evolution was not affected by rinsing with any buffer tested except Tris. Ferricyanide was photoreduced at a low rate by cells rinsed in Tes but at a high rate in TES plus magnesium and calcium ions. We interpreted our results to mean that, in Agmenellum quadruplicatum, inhibition of photosynthetic oxygen evolution by Tris occurs at the level of photosystem 2 while the effects of TES and phosphate are on electron-transport occurring after the rate-limiting reaction.     

Effect of temperature and pH on absorption of carbon dioxide by a free level of mixed solutions of some buffers

The rate of absorption of carbon dioxide by solutions of NaHCO₃, KH₂PO₄, hydrogencarbonate, phosphate and borate buffers at 20, 30 and 40°C was determined manometrically. The absorption rate increases for all buffers tested with increasing pH. The CO₂ absorption rate by KH₂PO₄ and by the phosphate buffer at low pH is lower than that of water. For other buffers tested it is equal to or higher than that of water, especially at higher temperatures.