From protein engineering to artificial enzymes - biological and biomimetic approaches towards sustainable hydrogen production.

Hydrogen gas is used extensively in industry today and is often put forward as a suitable energy carrier due its high energy density. Currently, the main source of molecular hydrogen is fossil fuels via steam reforming. Consequently, novel production methods are required to improve the sustainability of hydrogen gas for industrial processes, as well as paving the way for its implementation as a future solar fuel. Nature has already developed an elaborate hydrogen economy, where the production and consumption of hydrogen gas is catalysed by hydrogenase enzymes. In this review we summarize efforts on engineering and optimizing these enzymes for biological hydrogen gas production, with an emphasis on their inorganic cofactors. Moreover, we will describe how our understanding of these enzymes has been applied for the preparation of bio-inspired/-mimetic systems for efficient and sustainable hydrogen production.

Characterization of a monocyanide model of FeFe hydrogenases - highlighting the importance of the bridgehead nitrogen for catalysis.

An azadithiolate bridged monocyanide derivative [Fe2(adt)(CO)5(CN)]- of [Fe2(adt)(CO)4(CN)2]2- has been prepared and extensively characterized as a model of the [FeFe]-hydrogenase active site, using a combination of FTIR spectroscopy, electrochemical methods and catalytic assays with chemical reductants. The presence of two basic nitrogen sites opens up multiple protonation pathways, enabling catalytic proton reduction. To our knowledge [Fe2(adt)(CO)5(CN)]- represents the first example of a cyanide containing [FeFe]-hydrogenase active site mimic capable of catalytic H2 formation in aqueous media.

Biomimetic assembly and activation of [FeFe]-hydrogenases.

Hydrogenases are the most active molecular catalysts for hydrogen production and uptake, and could therefore facilitate the development of new types of fuel cell. In [FeFe]-hydrogenases, catalysis takes place at a unique di-iron centre (the [2Fe] subsite), which contains a bridging dithiolate ligand, three CO ligands and two CN(-) ligands. Through a complex multienzymatic biosynthetic process, this [2Fe] subsite is first assembled on a maturation enzyme, HydF, and then delivered to the apo-hydrogenase for activation. Synthetic chemistry has been used to prepare remarkably similar mimics of that subsite, but it has failed to reproduce the natural enzymatic activities thus far. Here we show that three synthetic mimics (containing different bridging dithiolate ligands) can be loaded onto bacterial Thermotoga maritima HydF and then transferred to apo-HydA1, one of the hydrogenases of Chlamydomonas reinhardtii algae. Full activation of HydA1 was achieved only when using the HydF hybrid protein containing the mimic with an azadithiolate bridge, confirming the presence of this ligand in the active site of native [FeFe]-hydrogenases. This is an example of controlled metalloenzyme activation using the combination of a specific protein scaffold and active-site synthetic analogues. This simple methodology provides both new mechanistic and structural insight into hydrogenase maturation and a unique tool for producing recombinant wild-type and variant [FeFe]-hydrogenases, with no requirement for the complete maturation machinery.

Dynamic neck strength training effect on pain and function.

This study examined whether neck resistance training could increase strength and reduce pain in workers with a high prevalence of neck disorders. Middle-aged women workers (n = 17) exercised twice weekly for 8 weeks. Each session (12min) consisted of three sets of 12 repetitions of resisted rotation, flexion, and extension muscles using hydraulic dampers. Resistance was set individually and progressively increased every second week. Angle-specific maximal isometric rotator, extensor, and flexor torque was measured before and after training. Torque during rotation, flexion and extension increased (p < .05) on average 35%, 27%, and 19%, respectively, after training. Perceived pain, assessed using a four-graded scale, was reduced (p < .05) after training. It is suggested that 12 minutes of specific neck strength training twice weekly for 8 weeks increases strength of rotator, extensor, and flexor muscles of the neck. Also all individuals who had pain reported reduced perceived neck pain after training.

Development of Heterodera glycines on Soybean Damaged by Soybean Looper and Stem Canker.

Short-term greenhouse studies with soybean (Glycine max cv. Bragg) were used to examine interactions between the soybean cyst nematode (Heterodera glycines) and two other common pests of soybean, the stem canker fungus (Diaporthe phaseolorum var. caulivora) and the soybean looper (Pseudoplusia includens), a lepidopterous defoliator. Numbers of cyst nematode juveniles in roots and numbers of cysts in soil and roots were reduced on plants with stem cankers. Defoliation by soybean looper larvae had the opposite effect; defoliation levels of 22 and 64% caused stepwise increases in numbers of juveniles and cysts in both roots and soil, whereas numbers of females in roots decreased. In two experiments, stem canker length was reduced 40 and 45% when root systems were colonized by the soybean cyst nematode. The absence of significant interactions among these pests indicates that the effects of soybean cyst nematode, stem canker, and soybean looper on plant growth and each other primarily were additive.

Presentation of growth velocities of rural Haitian children using smoothing spline techniques.

The examination of monthly (or quarterly) increments in weight or length is important for assessing the nutritional and health status of children. Growth velocities are widely thought to be more important than actual weight or length measurements per se. However, there are no standards by which clinicians, researchers, or parents can gauge a child's growth. This paper describes a method for computing growth velocities (monthly increments) for physical growth measurements with substantial measurement error and irregular spacing over time. These features are characteristic of data collected in the field where conditions are less than ideal. The technique of smoothing by splines provides a powerful tool to deal with the variability and irregularity of the measurements. The technique consists of approximating the observed data by a smooth curve as a clinician might have drawn on the child's growth chart. Spline functions are particularly appropriate to describe bio-physical processes such as growth, for which no model can be postulated a priori. This paper describes how the technique was used for the analysis of a large data base collected on pre-school aged children in rural Haiti. The sex-specific length and weight velocities derived from the spline-smoothed data are presented as reference data for researchers and others interested in longitudinal growth of children in the Third World.

Correlation between quantitative in vivo and in vitro responses in nickel-allergic patients.

Forty-three nickel-allergic patients (40 females and 3 males) participated in this study which was designed to elucidate the correlation between quantitative in vivo and in vitro responses. All were patch-tested with various concentrations of nickel-sulphate serial-diluted in water. The lowest concentration eliciting a cutaneous reaction (indicating the severity of the allergy) was recorded for each individual and compared with the in vitro lymphocyte response, using the lymphocyte transformation test. The best correlation coefficients obtained were 0.42 and 0.46, tested for linear and logarithmic correlation respectively. However, there were individuals who showed a weak cutaneous response and high lymphocyte reactivity. The converse situation was also found, however, indicating that other mechanisms than those mediated via lymphocytes may be of significance in the patch test response.

Comparison of Haitian children in a nutrition intervention programme with children in the Haitian national nutrition survey.

PIP: Weight-for-height and height-for-age data were compared for preschool-age Haitian children enrolled in a community health and nutrition intervention program and children measured in the Haiti national nutrition survey of 1978. Cross-sections of the longitudinal data of the intervention program corresponding to the season when the national survey was conducted (May-September) were chosen for the 3 years of available program data (1969-71). Significantly less stunting was found in children in the 1970 and 1971 intervention group than in the children covered by the national survey. Tests of trend also showed that the height (or length) status of the children in the intervention program improved from 1969-71. Wasting, or low weight status, was in general not significantly different in any of the comparisons. Nevertheless, the data were more favorable to children in the intervention groups, even in 1970, a year of food shortages. The results of the comparison are consistent with a positive program effect. (author's)^ieng

Population Development and Influence of Bursaphelenchus xylophilus on Gliocladium virens.

Gliocladium virens was isolated from slash pine trees symptomatic and asymptomatic for pine wilt disease with frequencies of 24% and 10%, respectively. Populations of Bursaphelenchus xylophilus, the nematode incitant of this disease, reproduced on this fungus and inhibited its growth. Growth inhibition of the fungus was characterized by an absence of sporulation and by the formation of chains of dark, thick-walled, chlamydospore-like cells. Population increase during a 12-day period following infestation of cultures of the fungus with 10,000 nematodes averaged 3-fold at 16 C, 9-fold at 20 C, and 24-fold at 24 C. In greenhouse studies, nematode recovery from slash pine seedlings coinoculated with both organisms was significantly greater than that obtained from seedlings inoculated with the nematode alone.

Traditional midwives, tetanus immunization, and infant mortality in rural Haiti.

PIP: Lessons learned from Haiti's integration of a training program for traditional birth attendants with the maternal and child health and family planning program are reported. The available data on illness and deaths reveal that Haiti has continuing problems of gastroenteritis, malnutrition, tuberculosis, malaria, and tetanus. The latter is of particular interest since neonatal tetanus derived from umbilical cord contamination continues to affect up to 10-20% of Haitian newborns in rural areas lacking health programs. Neonatal tetanus has largely disappeared in the Artibonite Valley due to a mass immunization program for the entire population, including young women, against tetanus. In the Albert Schweitzer Hospital program for indigenous midwives in Artibonite Valley, at least 36 midwives were reached on a regular basis in 1968 -- less than 1/3 of the midwives operating in the Artibonite Valley. There was a rapid decline in neonatal tetanus admissions during the period following 1968. This decline has been attributed to the use of rural health auxiliaries in immunizing the women in the hospital district, but indigenous midwives may have played a role. By 1970, the Albert Schweitzer Hospital program had grown from 36 midwives regularly attending midwife classes to 175 registered with the program during 1970. Although direct supervision proved difficult due to lack of communication and transport to the scene of delivery, some deliveries were observed and indirect supervision by the community became evident. An important finding of the traditional midwife training program of the Albert Schweitzer Hospital was the amount of time required for an indigenous midwife to have referred 50 newborns to the hospital for BCG vaccination. At the end of the 1st year of this program, only 2 midwives reached this goal. Another surprise was the increase in demand for "cord cut" services at the outpatient clinic rather than increased use of the nearby maternity unit. The elimination of neonatal tetanus as a cause of infant mortality was the most important outcome of the maternal and child health component of the community health program.^ieng

Preinvasive carcinoma of the cervix uteri and smoking.

The occurrence of smoking among 609 women with preinvasive cervical uterine cancer has been examined. Of these women, 66.3% were smokers. For 6 090 controls the figure was 41.9%. The difference is statistically significant (p less than 0.001). In all the age groups the proportion of smokers was greater among the women with cancer. These also started smoking at an earlier age, and the number of cigarettes smoked daily was greater than for the controls.

The influence of renal function on plasma levels and urinary excretion of acebutolol and its main N-acetyl metabolite.

The pharmacokinetics of acebutolol and its major N-acetyl metabolite diacetolol were determined following acute intravenous and chronic oral administration to 22 subjects with glomerular filtration rate (GFR) between 3 and 127 ml/min. Following chronic oral administration the mean terminal elimination half-life of unchanged acebutolol was about 10 hours independent of renal function, whereas the hall-life of the N-acetyl metabolite increased from 12.8 hr in subjects with normal renal function (GFR greater than 90 ml/min) to 24.0 hr in preuremic patients (GFR less than 10 ml/min). The mean area of the plasma level-time curve (AUC) of the metabolite was 14.2 mg 1-1 hr in patients with normal renal function and rose to a value of 81.4 mg 1-1 hr in preuremic patients. The mean AUC of the parent drug was not influenced by changes in renal function. The considerable accumulation of the acetyl metabolite, which has about the same beta blocking activity as the unchanged drug, necessitates dose reduction of acebutolol in the presence of different degrees of renal impairment (reduction by half of the normal daily dose in patients with GFR between 30 and 10 ml/min and by three quarters when GFR less than 10 ml/min). In subjects with normal renal function, the ratio between the AUC of the parent drug and that of its major N-acetyl metabolite was 3:1 after intravenous injection of the drug, while it was 1:2.5 following chronic oral administration, indicating a marked first pass metabolism of acebutolol.

Reduction of mortality in rural Haiti through a primary-health-care program.

Deaths and their causes in a rural Haitian population of 8820 were studied through hospital records, death registration, a disease survey, and health surveillance. The results were used in selecting eight diseases for the delivery of health services by village-level health workers. The impact of the services was measured by monitoring annual age-specific and disease-specific mortality rates and by comparing them with officially estimated national mortality rates. Mortality rates fell progressively during five years, to levels only one fourth as high as the national estimates. The fall in mortality was associated principally with services that prevented deaths due to tetanus, malnutrition, diarrhea, and tuberculosis. The total program of hospital and village health services saved 495 years of potential life per thousand population per year. Most of the saving was attributable to preventive services. The program eventually served more than 115,000 persons, and it has been replicated by other agencies for an additional 135,000 Haitians.

PIP: Deaths and their causes in a rural Haitian population of 8820 were studied through hospital records, death registration, a disease survey, and health surveillance. The results were used in selecting 8 diseases for the delivery of health services by village-level health workers. The impact of the services was measured by monitoring annual age-specific and disease-specific mortality rates and by comparing them with officially estimated national mortality rates. Mortality rates fell progressively during 5 years, to levels only 1/4 as high as the national estimates. The fall in mortality was associated principally with services that prevented deaths due to tetanus, malnutrition, diarrhea, and tuberculosis. The total program of hospital and village health services saved 495 years of potential life/thousand population/year. Most of the saving was attributable to preventive services. The program eventually served more than 115,000 persons, and it has been replicated by other agencies for an additional 135,000 Haitians.

Acute water shortage and health problems in Haiti.

During a severe drought Port-au-Prince, Haiti, lost hydroelectric power for 10 weeks. This led to water shortages in areas of the city dependent on water supplied from electrically driven pumps. In a study of the impact of water restriction on disease, 400 families were randomly selected from two urban areas differentially affected by the water shortage. Disease in children was found to be related to quantity of water used, socioeconomic status, employment of head of household, and family size. The methods used in this study are recommended for the investigation of the relationship between water quantity and health.

Physical work capacity of young men in Sweden.

Each year between 1961 and 1975 all male students in a secondary school were submaximally tested on several occasions on bicycle ergometers with regard to estimated maximal aerobic power. The school is situated in central Sweden (Falun). The students were between 16 and 20 years of age. Data from the tests in Falun and data compiled from other sources cannot confirm the hypothesis that young men of today have a lower maximal aerobic power than young men of the 1960s. The results indicate that Swedish boys aged 16 to 20 have, on average, a maximal aerobic power of about 45 ml O2 X (kg x min)-1. The standard deviation is about 8 ml O2.

Preventive and therapeutic benefits in relation to cost: performance over 10 years of Mothercraft Centers in Haiti.

Evaluation of the effectiveness of Mothercraft Centers operating in Haiti over the period 1964 to 1975 as a public health measure in private and public health services is reported. For the most part, the data are derived from centers operating routinely in the health services rather than from pilot demonstrations. Their impact in both therapy and prevention of severe malnutrition in preschool children is considered, and it is found that numerically the greater benefit is in prevention. At a total annual cost of $4034 a typical center accomplishes successful therapy and prevents relapse in 105 children. In addition, it provides protection against severe malnutrition to their 306 younger siblings. It is thus estimated that, considering only these 411 children demonstrably benefited from a year's operation, the cost of providing essentially lifetime protection from severe malnutrition is approximately $10 per child.