Ascarishaemoglobin as an indicator of the oxygen produced by isolated chloroplasts

Abstract

A technique for using haemoglobin as an indicator of oxygen produced by the photosynthetic activity of green leaves was first described by Hoppe-Seyler 1879). The method has been used with notable success by Hill (1937, 1939) and by Hill & Scarisbrick (1940) in their observations on the sub-cellular activity of isolated phloroplasts. The quantities of oxygen evolved in their experiments were small, and since the sensitivity of measurement was dependent upon the affinity of the haemoglobin for oxygen, it was desirable to select a haemoglobin having as high an oxygen affinity as possible. Hill (1936), however, had shown that muscle haemoglobin in dilute solution has a greater affinity than the blood haemoglobin of the same mammal. Thus at pH 8 and 19°C he found ox muscle haemoglobin to be half saturated at an O₂pressure of 0.7 mm. compared with 1.8 mm. for ox blood haemoglobin under the same conditions. Using muscle haemoglobin as indicator, Hill (1939) was able to show that illuminated chloroplasts in presence of leaf extracts evolve oxygen to a pressure of 1 mm. Hg and in presence of ferric oxalate to a pressure of 4 mm. Hg. Haemoglobins are now known which have oxygen affinities higher than that of mammalian muscle haemoglobin. In particular it has been shown thatAscaris lumbricoides,a nematode parasitic in the pig, contains two haemoglobins re­markable for their extreme resistance to deoxygenation when they are equilibratedin vacuo(Davenport 1949). Because of this property the standard methods for the determination of the oxygen equilibrium curves could not be used. Hill (1939) had used a haemoglobin of known oxygen affinity to measure the tensions of oxygen produced by illuminated chloroplast systems. The object of the present paper is to compare the response of ox muscle haemoglobin and theAscarishaemoglobins to oxygen produced photochemically under standard conditions and thus to obtain an indication of the relative affinities of the haemoglobins.