Using videoconsultations to deliver dietary advice to children with chronic kidney disease: a qualitative study of parent and child perspectives.

BACKGROUND: Children with chronic kidney disease require specialist renal paediatric dietetic care, regardless of disease severity or geographical location; however, under-resourcing makes this challenging. Videoconsultation may offer a solution but research exploring its acceptability is limited. The present study explored parent/carer and child perspectives of videoconsultation as an alternative or supplement to existing regional dietetic care. METHODS: Children and families using a regional paediatric nephrology service were recruited through purposeful sampling techniques. Renal paediatric dietitians used existing hospital software to host videoconsultations with families. Perspectives were subsequently explored in telephone interviews with the children, their parents and separately with the renal dietitians. Data were transcribed verbatim and an inductive framework analysis conducted. RESULTS: Twelve families took part in the study, comprising 13 parents and five children (aged 9 months to 14 years). Two renal dietitians were also interviewed. Six themes emerged which were 'Logistics', 'Understanding Information', 'Family Engagement', 'Establishing Trust', 'Willingness to Change' and 'Preferences'. Satisfaction with the videoconsultations was high, with no data security fears and only minor privacy concerns. Parents reported that screen-sharing software enhanced their understanding, generating greater discussion and engagement compared to clinic and telephone contacts. Parents praised efficiencies and  improved access to specialist advice, requesting that videoconsultations supplement care. Children preferred videoconsultations outright. CONCLUSIONS: Dietetic videoconsultations were acceptable to families and perceived to be a feasible, high-quality complement to regional specialist dietetic care. Enhanced understanding and engagement might improve self-care in adolescents. The acceptability and feasibility of videoconsultations could address inequitable regional service provision.

Improving the nutritional intake of hospital patients: how far have we come? A re-audit.

BACKGROUND: Malnutrition affects up to 33.6% of hospitalised patients, with consequences that are detrimental for both patients and healthcare providers. In 2015, an audit demonstrated inadequate nutritional provision and consumption by hospitalised patients, comprising a major risk factor for malnutrition. This re-audit evaluates whether patients are meeting recommended energy and protein standards and estimated individual requirements, subsequent to food service improvements since 2015. METHODS: Patients (n = 111) were included from a South West hospital, and Malnutrition Universal Screening Tool scores (MUST) categorised patients as 'nutritionally well' (MUST 0) or 'nutritionally vulnerable' (MUST ≥ 1). Individual energy and protein requirements were estimated using weight-based equations. Nutritional intakes were assessed via 24-h dietary recall and compared against the British Dietetic Association's Nutrition and Hydration Digest standards, as well as estimated individual requirements. RESULTS: In total, the Digest standards for energy and protein were met by 35% and 63% of patients respectively, which is an increase of 19% and 36% since 2015. 'Nutritionally well' patients were more likely to meet nutrient standards for protein (62%) than estimated individual requirements (30%) (P ≤ 0.001). 'Nutritionally vulnerable' patients were more likely to meet estimated individual requirements for energy (60%) than the Digest standards (30%) (P = 0.047). CONCLUSIONS: The proportion of patients meeting the Digest standards has increased considerably following numerous food service changes. Nutritional training for housekeepers, energy/protein-dense snacks and drinks, and fortified dietary items may further increase nutritional intakes. Additionally, as a result of discrepancies between the Digest standards and individual estimated requirements, more research is required to identify the most appropriate auditing standards that reflect best practice.

Future Dietitian 2025: informing the development of a workforce strategy for dietetics.

BACKGROUND: Healthcare is changing and the professions that deliver it need to adapt and change too. The aim of this research was to inform the development of a workforce strategy for Dietetics for 2020-2030. This included an understanding of the drivers for change, the views of stakeholders and recommendations to prepare the profession for the future. METHODS: The research included three phases: (i) establishing the context which included a literature and document review (environmental scan); (ii) discovering the profession and professional issues using crowd-sourcing technology; and (iii) articulating the vision for the future using appreciative inquiry. RESULTS: The environmental scan described the current status of the dietetic profession, the changing healthcare environment, the context in which dietitians work and what future opportunities exist for the profession. The online conversation facilitated by crowd-sourcing technology asked the question: 'How can dietitians strengthen their future role, influence and impact?' Dietitians and interested stakeholders (726 and 109, respectively) made 6130 contributions. Seven priorities were identified and fed into the appreciative inquiry event. The event bought together 54 dietitians and analysis of the discussions generated five themes: (i) professional identity; (ii) strong foundations-creating structure and direction for the profession; (iii) amplifying visibility and influence; (iv) embracing advances in science and technology; and (v) career advancement and emerging opportunities. CONCLUSIONS: A series of recommendations were made for the next steps in moving the workforce to a new future. The future for dietetics looks bright, embracing technology, as well as exploring different ways of working and new opportunities, as this dynamic profession continues to evolve.

Are energy and protein requirements met in hospital?

BACKGROUND: Malnutrition is a problem within hospitals, which impacts upon clinical outcomes. The present audit assesses whether a hospital menu meets the energy and protein standards recommended by the British Dietetic Association's (BDA) Nutrition and Hydration Digest and determines the contribution of oral nutrition supplements (ONS) and additional snacks. METHODS: Patients in a UK South West hospital were categorised as 'nutritionally well' or 'nutritionally vulnerable' in accordance with their Malnutrition Universal Screening Tool score. Energy and protein content of food selected from the menu ('menu choice'), menu food consumed ('hospital intake') and total food consumed including snacks ('overall intake') were calculated and compared with the standards. RESULTS: In total, 93 patients were included. For 'nutritionally well' patients (n = 81), energy and protein standards were met by 11.1% and 33.3% ('menu choice'); 7.4% and 22.2% ('hospital intake'); and 14.8% and 28.4% ('overall intake'). For 'nutritionally vulnerable' patients (n = 12), energy and protein standards were met by 0% and 8.3% ('menu choice'); 0% and 8.3% ('hospital intake'); and 8.3% and 16.7% ('overall intake'). Ten percent of patients consumed ONS. Patients who consumed hospital snacks (34%) were more likely to meet the nutrient standards (P ≤ 0.001). CONCLUSIONS: The present audit demonstrated that most patients are not meeting the nutrient standards recommended by the BDA Nutrition and Hydration Digest. Recommendations include the provision of energy/protein-dense snacks, as well as menu, offering ONS where clinically indicated, in addition to training for staff. A food services dietitian is ideally placed to lead this, forming a vital link between patients, caterers and clinical teams.

An evaluation of an Internet-based approach to weight loss with low glycaemic load principles.

BACKGROUND: The prevalence of overweight and obesity is increasing worldwide at an alarming rate. An Internet-based weight-loss programme has the potential to reach larger numbers of people than traditional face-to-face programmes. A growing body of evidence supports the use of low glycaemic load (GL) diets for weight loss. The present study aimed to investigate the efficacy of an Internet-based weight-loss programme that included foods with a low GL. METHODS: One hundred and three volunteers, with a body mass index (BMI) ≥28 kg m(-2) , enrolled into an Internet weight-loss programme. A dietitian counselled participants over the Internet via weekly interactive chat rooms and monthly e-mails. Participants self-recorded body weight and food intake directly on to the Internet site. Weight, BMI and waist circumference were measured, and dietary data collected, at baseline and 6 months. RESULTS: Seventy participants completed the 6-month weight-loss programme. Among these, mean weight, BMI and waist circumference significantly decreased by 3.5 kg (95% CI = 2.3-4.7), 1.2 kg m(-2) (95% CI = 0.8-1.7) and 4.8 cm (95% CI = 2.8-6.8), of baseline values respectively (P < 0.001). Twenty-five (36%) of the 70 participants lost a clinically significant amount of weight (>5% of initial body weight). CONCLUSIONS: This descriptive study has shown that an Internet-based weight-loss programme with low GL principles can promote weight loss. This type of intervention and approach could be used to enhance other weight-loss strategies.

Early-life nutritional and environmental determinants of thymic size in infants born in rural Bangladesh.

AIM: The aim was to assess the impact of nutritional status and environmental exposures on infant thymic development in the rural Matlab region of Bangladesh. METHODS: In a cohort of N(max) 2094 infants born during a randomized study of combined interventions to improve maternal and infant health, thymic volume (thymic index, TI) was assessed by ultrasonography at birth and at 8, 24 and 52 weeks of age. Data on birth weight, infant anthropometry and feeding status were also collected. RESULTS: At all ages, TI was positively associated with infant weight and strongly associated with the month of measurement. Longer duration of exclusive breastfeeding resulted in a larger TI at 52 weeks. TI at birth and at 8 weeks correlated positively with birth weight, but by 24 and 52 weeks and when adjusted for infant weight this effect was no longer present. Thymic size was not affected by pre-natal maternal supplementation or by socioeconomic status but was correlated to arsenic exposure during pregnancy. CONCLUSION: In this population of rural Bangladeshi infants, thymic development is influenced by both nutritional and environmental exposures early in life. The long-term functional implications of these findings warrant further investigation.

An immunotoxin prepared with blocked ricin: a natural plant toxin adapted for therapeutic use.

Ricin, the cytotoxic protein isolated from castor beans, is composed of two subunits, A-chain and B-chain. Ricin intoxicates cells by binding through its B-chain to galactose-terminated oligosaccharides found on the surface of all eukaryotic cells and then transferring its A-chain to the cytosol where it disrupts protein synthesis by inactivating ribosomes. In addition to binding, the B-chain plays an important, but not yet understood, role in the translocation of the A-chain through a cellular membrane to the cytosol. Blocking the two galactose-binding sites of native ricin by chemical modification with affinity ligands created an altered toxin, called blocked ricin, that has at least a 3500-fold lower binding affinity and is more than 1000-fold less cytotoxic than native ricin for Namalwa cells (a Burkitt's lymphoma line) but that has maintained the translocation function of the B-chain and the catalytic activity of the A-chain. Conjugation of blocked ricin to monoclonal antibodies that bind to cell surface antigens creates new cytotoxins that approach the potency of native ricin. These cytotoxins incorporate the three essential functions of natural toxins, i.e., binding to cells, transport through a membrane, and catalytic inactivation of an essential cellular process; but in addition they possess a defined cellular target specificity. Such potent immunotoxins may play an important therapeutic role in cancer treatment. Clinical trials with an anti-CD19-blocked ricin and an anti-CD33-blocked ricin conjugate against B-cell cancers and acute myeloblastic leukemia have begun.

Immunotoxin therapy of small-cell lung cancer: a phase I study of N901-blocked ricin.

PURPOSE: Immunotoxins could improve outcome in small-cell lung cancer (SCLC) by targeting tumor cells that are resistant to chemotherapy and radiation. N901 is a murine monoclonal antibody that binds to the CD56 (neural cell adhesion molecule [NCAM]) antigen found on cells of neuroendocrine origin, including SCLC. N901-bR is an immunoconjugate of N901 antibody with blocked ricin (bR) as the cytotoxic effector moiety. N901-bR has more than 700-fold greater selectivity in vitro for killing the CD56+ SCLC cell line SW-2 than for an antigen-negative lymphoma cell line. Preclinical studies suggested the potential for clinically significant cardiac and neurologic toxicity. We present a phase I study of N901-bR in relapsed SCLC. PATIENTS AND METHODS: Twenty-one patients (18 relapsed, three primary refractory) with SCLC were entered onto this study. Successive cohorts of at least three patients were treated at doses from 5 to 40 microg/kg/d for 7 days. The initial three cohorts received the first day's dose (one seventh of planned dose) as a bolus infusion before they began the continuous infusion on the second day to observe acute toxicity and determine bolus pharmacokinetics. Toxicity assessment included nerve-conduction studies (NCS) and radionuclide assessment of left ventricular ejection fraction (LVEF) before and after N901-bR administration to fully assess potential neurologic and cardiac toxicity. RESULTS: The dose-limiting toxicity (DLT) of N901-bR given by 7-day continuous infusion is capillary leak syndrome, which occurred in two of three patients at the dose of 40 microg/kg (lean body weight [LBW])/d. Detectable serum drug levels equivalent to effective in vitro drug levels were achieved at the 20-, 30-, and 40-microg/kg(LBW)/d dose levels. Specific binding of the immunotoxin to tumor cells in bone marrow, liver, and lung was observed. Cardiac function remained normal in 15 of 16 patients. No patient developed clinically significant neuropathy. However, a trend was noted for amplitude decline in serial NCS of both sensory and motor neurons. One patient with refractory SCLC achieved a partial response. CONCLUSION: N901-bR is an immunotoxin with potential clinical activity in SCLC. N901-bR is well tolerated when given by 7-day continuous infusion at the dose of 30 microg/kg(LBW)/d. Neurologic and cardiac toxicity were acceptable when given to patients with refractory SCLC. A second study to evaluate this agent after induction chemoradiotherapy in both limited- and extensive-stage disease was started following completion of this study.

Immune function in rural Gambian children is not related to season of birth, birth size, or maternal supplementation status.

BACKGROUND: We previously showed that mortality from infectious diseases among young adults in rural Gambia is strongly correlated with the season of their birth. This suggests that early life insults that involve fetal malnutrition, exposure to natural toxins, or highly seasonal infections affecting the infant or pregnant mother cause permanent damage to the immune system. Excess mortality begins after puberty and has a maximal odds ratio of >10 for deaths between ages 25 and 50 y. OBJECTIVE: We investigated the immune function of children according to birth weight, season of birth, and exposure to maternal dietary supplementation during pregnancy. DESIGN: Immune function was measured in 472 prepubertal children aged 6.5-9.5 y from 28 villages in rural Gambia. The mothers of these children had been randomly assigned to a high-energy prenatal supplementation program, which significantly increased birth weight. This permitted supplementation status, birth weight, and season of birth to be investigated as exposure variables. The outcome variables tested were naive responses to rabies and pneumococcus vaccines, delayed-type hypersensitivity skin reactions, and mucosal defense (secretory immunoglobulin A and dual-sugar permeability). RESULTS: Immune responses were strongly related to current age and sex, suggesting a high level of sensitivity, but were not consistently related to birth weight, season of birth, or maternal supplementation (control compared with intervention). CONCLUSION: Events in early life did not predict a measurable defect in immune response within this cohort of rural Gambian children. It is possible that the early programming of immune function may be mediated through a defect in immunologic memory or early senescence rather than through impairment of early responses.

Prenatal or early postnatal events predict infectious deaths in young adulthood in rural Africa.

BACKGROUND: Research over the past decade has suggested that prenatal and early postnatal nutrition influence the risk of developing chronic degenerative diseases up to 60 years later. We now present evidence that risk of death from infectious diseases in young adulthood is similarly programmed by early life events. METHODS: In three rural Gambian villages, affected by a marked annual seasonality in diet and disease, we have kept detailed demographic, anthropometric and health records since 1949. Fate was known with certainty for 3,162 individuals (2,059 alive/1,103 dead, most dying in childhood). For this case-control analysis of antecedent predictors of premature mortality, all adult deaths (n = 61) were paired with two randomly selected controls matched for sex and year of birth. RESULTS: Mean age at death was 25 (SD: 8) years. Adult death was associated with a profound bias in month of birth with 49 cases born in the nutritionally-debilitating hungry season (Jul-Dec) versus 12 in the harvest season (Jan-Jun). Relative to harvest season the hazard ratio for early death in hungry-season births rose from 3.7 (for deaths >14.5 years, P = 0.000013) to 10.3 (for deaths >25 years, P = 0.00002). Anthropometric and haematological status at 18 months of age was identical in cases and controls, indicating an earlier origin to the defect. Most deaths for which cause was known had a definite or possible infectious aetiology; none were from degenerative diseases of affluence. CONCLUSIONS: Early life exposures, correlated with season of birth, strongly influence susceptibility to fatal infections in young adulthood. The evidence suggests that nutritionally-mediated intrauterine growth retardation may permanently impair the development of immune function.

PIP: This paper presents the influence of prenatal and early postnatal nutrition on the risk of developing chronic degenerative diseases during late adulthood. Investigation of the thrifty phenotype hypothesis using 1949 birth records in three rural Gambian villages illustrates the severity of seasonality in diet and disease patterns. A database of 3162 individuals--2059 survivors and 1103 fatalities--was examined. Using a case-control analysis of antecedent predictors of premature mortality, all deaths were paired with two randomly selected controls matched for sex and year. Results revealed that the mean age at death was 25 years, with 49 adult death cases occurring in the nutritionally debilitating hungry season compared to 12 cases occurring during the harvest season. In relation to the harvest season, the hazard ratio rose from 3.7 (for deaths at age 14.5 years; p = 0.000013) to 10.3 (for deaths at age 25 years; p = 0.00002). Both the anthropometric and hematological status at 18 months was similar in cases and controls, with most deaths caused by a definite or possible infectious etiology without evidence of degenerative disease cause. This study concludes that early life exposures correlated with season of birth significantly influence the susceptibility to fatal infections in young adulthood. This also suggests that nutritionally mediated intrauterine growth retardation could permanently impair the development of immune function.

Regulation of soluble 5'-nucleotidase I from rabbit heart.

Rabbit heart contains two soluble 5'-nucleotidases, termed N-I and N-II, which can be separated using phosphocellulose chromatography. N-I prefers AMP over IMP as substrate, in contrast to N-II which prefers IMP over AMP. Both enzymes require Mg2+, but the optimum Mg2+ concentrations for the two enzymes are different. High concentrations of NaCl inhibit N-I and activate N-II. Purified N-I is activated by ADP but not by ATP. According to Itoh et al. (1986), purified N-II is activated by both ADP and ATP. N-I has been purified approximately 1000-fold to a specific activity of approximately 100 mumol/mg protein/min. The properties of N-I suggest that it is the enzyme responsible for the release of adenosine from AMP under conditions of hypoxia or increased work load.

Managing serum phosphate in haemodialysis patients: time for an innovative approach?

BACKGROUND/OBJECTIVES: Hyperphosphataemia, a common biochemical abnormality in chronic kidney disease, poses significant management challenges. This study aims to determine whether the reasons for this are multifactorial; including poor dietary knowledge, poor adherence to a low phosphate diet and phosphate-binding medications and the impact of age on these parameters. SUBJECTS/METHODS: In order to compare serum phosphate and other associated parameters to the UK Renal Association Clinical Practice Guidelines 2010 an audit and service evaluation questionnaire was carried out in May 2011 on 130 haemodialysis outpatients attending the Plymouth Dialysis Unit. RESULTS: Fifty-three percent of patients had serum phosphate within the target range of 1.1-1.7 mmol/l, 77% and 85% had serum calcium and parathyroid hormone within target ranges, respectively. Younger patients (18-45 years) were significantly less likely to have serum phosphate within range χ(2) (2, n=124)=18.77, P<0.001. Despite better knowledge of their own phosphate levels (P=0.005), phosphorus-rich foods (P<0.001), symptoms of hyperphosphataemia (P<0.001) and increased use of Renal Patient View (P=0.002), <65 years old had significantly higher phosphate levels than those >65 years (P<0.001). No significant associations were found between phosphate control and the following factors: gender, timing of dialysis shift, years on dialysis or dialysis adequacy. CONCLUSIONS: In this population, despite better knowledge, younger patients have worse phosphate control than older patients. Using the same dietary education techniques may not be suitable for all ages, more innovative approaches supported by skilled health professionals are needed to motivate and engage with younger patients to promote self-management and adherence.

Characterizing the burden of invasive Pseudomonas infection on neonatal units in the UK between 2005 and 2011.

Concern about Pseudomonas infection in neonatal units has focused on outbreaks. This study analysed cases of invasive Pseudomonas infection in 18 UK neonatal units participating in the NeonIN Neonatal Infection Surveillance Network from January 2005 to December 2011. Forty-two cases were reported. The majority (35/42, 93%) of cases were late-onset (median 14 days, range 2-262 days), the highest incidence was seen in extremely-low-birthweight infants and all cases were sporadic. One-third of cases were known to be colonized prior to invasive disease. Attributable mortality was 18%. Opportunities for preventing invasive disease due to this important pathogen should be prioritized.

Missed opportunities for preventing group B streptococcus infection.

BACKGROUND: Group B streptococcus (GBS) is the most common cause of early onset (EO) neonatal infection in the UK. National guidelines for its prevention were introduced in 2003. We assessed the opportunities for prevention amongst cases of EO GBS using the electronic Neonatal Infection Surveillance Network (NeonIN). METHODS: Culture proven EO GBS cases occurring between 2004 and 2007 were identified prospectively in eight neonatal units participating in NeonIN. Data concerning risk factors, intrapartum antibiotic (IAP) use and infant outcome were collected retrospectively. RESULTS: There were 48 cases of GBS over the 4 years (0.52/1000 live-births); 22 male, median gestation 38 weeks. The most common clinical presentation was sepsis and the GBS-attributable mortality was 6%. Risk factors were present in 67% (32) and adequate IAP was given to six of these mothers (19%). If all women with risk factors received prophylaxis, 23 cases (48%) may have been prevented. CONCLUSIONS: Better GBS prevention strategies are required in the UK.

Safety and immunogenicity of AS03B adjuvanted split virion versus non-adjuvanted whole virion H1N1 influenza vaccine in UK children aged 6 months-12 years: open label, randomised, parallel group, multicentre study.

OBJECTIVES: To compare the safety, reactogenicity, and immunogenicity of an adjuvanted split virion H1N1 vaccine and a non-adjuvanted whole virion vaccine used in the pandemic immunisation programme in the United Kingdom. DESIGN: Open label, randomised, parallel group, phase II study. SETTING: Five UK centres (Oxford, Southampton, Bristol, Exeter, and London). PARTICIPANTS: Children aged 6 months to less than 13 years for whom a parent or guardian had provided written informed consent and who were able to comply with study procedures were eligible. Those with laboratory confirmed pandemic H1N1 influenza or clinically diagnosed disease meriting antiviral treatment, allergy to egg or any other vaccine components, or coagulation defects, or who were severely immunocompromised or had recently received blood products were excluded. Children were grouped by age: 6 months-<3 years (younger group) and 3-<13 years (older group). Recruitment was by media advertising and direct mailing. Recruitment visits were attended by 949 participants, of whom 943 were enrolled and 937 included in the per protocol analysis. INTERVENTIONS: Participants were randomised 1:1 to receive AS03(B) (tocopherol based oil in water emulsion) adjuvanted split virion vaccine derived from egg culture or non-adjuvanted whole virion vaccine derived from cell culture. Both were given as two doses 21 days apart. Reactogenicity data were collected for one week after immunisation by diary card. Serum samples were collected at baseline and after the second dose. MAIN OUTCOME MEASURES: Primary reactogenicity end points were frequency and severity of fever, tenderness, swelling, and erythema after vaccination. Immunogenicity was measured by microneutralisation and haemagglutination inhibition assays. The primary immunogenicity objective was a comparison between vaccines of the percentage of participants showing seroconversion by the microneutralisation assay (fourfold rise to a titre of >or=1:40 from before vaccination to three weeks after the second dose). RESULTS: Seroconversion rates were higher after the adjuvanted split virion vaccine than after the whole virion vaccine, most notably in the youngest children (163 of 166 participants with paired serum samples (98.2%, 95% confidence interval 94.8% to 99.6%) v 157 of 196 (80.1%, 73.8% to 85.5%), P<0.001) in children under 3 years and 226 of 228 (99.1%, 96.9% to 99.9%) v 95.9%, 92.4% to 98.1%, P=0.03) in those over 3 years). The adjuvanted split virion vaccine was more reactogenic than the whole virion vaccine, with more frequent systemic reactions and severe local reactions in children aged over 5 years after dose one (13 (7.2%, 3.9% to 12%) v 2 (1.1%, 0.1% to 3.9%), P<0.001) and dose two (15 (8.5%, 4.8% to 13.7%) v 2 (1.1%, 0.1% to 4.1%), P<0.002) and after dose two in those under 5 years (15 (5.9%, 3.3% to 9.6%) v 0 (0.0%, 0% to 1.4%), P<0.001). Dose two of the adjuvanted split virion vaccine was more reactogenic than dose one, especially for fever >or=38 masculineC in those aged under 5 (24 (8.9%, 5.8% to 12.9%) v 57 (22.4%, 17.5% to 28.1%), P<0.001). CONCLUSIONS: In this first direct comparison of an AS03(B) adjuvanted split virion versus whole virion non-adjuvanted H1N1 vaccine, the adjuvanted vaccine, while more reactogenic, was more immunogenic and, importantly, achieved high seroconversion rates in children aged less than 3 years. This indicates the potential for improved immunogenicity of influenza vaccines in this age group. TRIAL REGISTRATION: Clinical trials.gov NCT00980850; ISRCTN89141709.

An amplified assay for thiols based on reactivation of papain.

A sensitive spectrophotometric assay has been developed for thiol (sulfhydryl) groups using an inactive disulfide derivative of papain (papain-S-SCH3). The thiol-disulfide interchange reaction of a thiol with papain-S-SCH3 results in the stoichiometric formation of active papain (papain-SH). The reactivated papain catalyzes the hydrolysis of a chromogenic substrate, resulting in an amplified spectrophotometric signal proportional to the initial amount of thiol. A variety of thiols, e.g., cysteine, glutathione, penicillamine, cysteine methyl ester, and cysteamine, yield similar linear plots for the activity of papain vs the initial amount of thiol. An unknown concentration of a thiol is measured using a standard plot for the activity of papain vs the amount of thiol, obtained for the same thiol or for a similar thiol. Thiol groups on proteins and thiol groups of high values of pKa (2-mercaptoethanol, 3-mercaptopropanoic acid) can also be assayed using papain-S-SCH3 in the presence of excess cystamine. The assay is about 100-fold more sensitive than that using Ellman's reagent [5,5'-dithiobis(2-nitrobenzoic acid)]. A 0.4 microM solution of cysteine produces an absorbance change of 0.55 at 410 nm after 30 min in the assay, compared to a predicted change in absorbance of 0.0054 using Ellman's assay.

5'-Nucleotidases in rat heart. Evidence for the occurrence of two soluble enzymes with different substrate specificities.

Chromatography of soluble proteins from rat heart on phosphocellulose columns separates two 5'-nucleotidases. The first to emerge from the column shows a preference for AMP over IMP as substrate, whereas the second shows a preference for IMP over AMP. The properties of the IMP-preferring enzyme, including the conditions under which it is eluted from phosphocellulose columns, show it to be the enzyme studied by Itoh, Oka & Ozasa [Biochem. J. (1986) 235, 847-851]. The kinetic properties of the AMP-preferring enzyme indicate that it is likely to be the enzyme responsible for the production of adenosine under conditions of hypoxia and increased work load, and with metabolic stresses such as a high load of acetate.