Malignancy and mortality in paediatric-onset inflammatory bowel disease: a 3-year prospective, multinational study from the paediatric IBD Porto group of ESPGHAN.

BACKGROUND: Risk benefit strategies in managing inflammatory bowel diseases (IBD) are dependent upon understanding the risks of uncontrolled inflammation vs those of treatments. Malignancy and mortality in IBD have been associated with disease-related inflammation and immune suppression, but data are limited due to their rare occurrence. AIM: To identify and describe the most common causes of mortality, types of cancer and previous or current therapy among children and young adults with paediatric-onset IBD. METHODS: Information on paediatric-onset IBD patients diagnosed with malignancy or mortality was prospectively collected via a survey in 25 countries over a 42-month period. Patients were included if death or malignancy occurred after IBD diagnosis but before the age of 26 years. RESULTS: In total, 60 patients were identified including 43 malignancies and 26 fatal cases (9 due to cancer). Main causes of fatality were malignancies (n = 9), IBD or IBD-therapy related nonmalignant causes (n = 10; including 5 infections), and suicides (n = 3). Three cases, all fatal, of hepatosplenic T-cell lymphoma were identified, all were biologic-naïve but thiopurine-exposed. No other haematological malignancies were fatal. The 6 other fatal cancer cases included 3 colorectal adenocarcinomas and 3 cholangiocarcinomas (CCAs). Primary sclerosing cholangitis (PSC) was present in 5 (56%) fatal cancers (1 colorectal carcinoma, 3 CCAs and 1 hepatosplenic T-cell lymphoma). CONCLUSIONS: We report the largest number of paediatric-onset IBD patients with cancer and/or fatal outcomes to date. Malignancies followed by infections were the major causes of mortality. We identified PSC as a significant risk factor for cancer-associated mortality. Disease-related adenocarcinomas were a commoner cause of death than lymphomas.

A British Society of Paediatric Gastroenterology, Hepatology and Nutrition survey of the effectiveness and safety of adalimumab in children with inflammatory bowel disease.

BACKGROUND: Adalimumab is efficacious therapy for adults with Crohn's disease (CD). AIM: To summarise the United Kingdom and Republic of Ireland paediatric adalimumab experience. METHODS: British Society of Paediatric Gastroenterology, Hepatology and Nutrition (BSPGHAN) members with Inflammatory Bowel Disease (IBD) patients <18 years old commencing adalimumab with at least 4 weeks follow-up. Patient demographics and details of treatment were then collected. Response and remission was assessed using the Paediatric Crohn's Disease Activity Index (PCDAI)/Physicians Global Assessment (PGA). RESULTS: Seventy-two patients [70 CD, 1 ulcerative colitis (UC), 1 IBD unclassified (IBDU)] from 19 paediatric-centres received adalimumab at a median age of 14.8 (IQR 3.1, range 6.1-17.8) years; 66/70 CD (94%) had previously received infliximab. A dose of 80 mg then 40 mg was used for induction in 41(59%) and 40 mg fortnightly for maintenance in 61 (90%). Remission rates were 24%, 58% and 41% at 1, 6 and 12 months, respectively. Overall 43 (61%) went into remission at some point, with 24 (35%) requiring escalation of therapy. Remission rates were higher in those on concomitant immunosuppression cf. those not on immunosuppression [34/46 (74%) vs. 9/24 (37%), respectively, (χ(2) 8.8, P=0.003)]. There were 15 adverse events (21%) including four (6%) serious adverse events with two sepsis related deaths in patients who were also on immunosuppression and home parenteral nutrition (3% mortality rate). CONCLUSIONS: Adalimumab is useful in treatment of refractory paediatric patients with a remission rate of 61%. This treatment benefit should be balanced against side effects, including in this study a 3% mortality rate.

Priority water research questions as determined by UK practitioners and policy makers.

Several recent studies have emphasised the need for a more integrated process in which researchers, policy makers and practitioners interact to identify research priorities. This paper discusses such a process with respect to the UK water sector, detailing how questions were developed through inter-disciplinary collaboration using online questionnaires and a stakeholder workshop. The paper details the 94 key questions arising, and provides commentary on their scale and scope. Prioritization voting divided the nine research themes into three categories: (1) extreme events (primarily flooding), valuing freshwater services, and water supply, treatment and distribution [each >150/1109 votes]; (2) freshwater pollution and integrated catchment management [100-150 votes] and; (3) freshwater biodiversity, water industry governance, understanding and managing demand and communicating water research [50-100 votes]. The biggest demand was for research to improve understanding of intervention impacts in the water environment, while a need for improved understanding of basic processes was also clearly expressed, particularly with respect to impacts of pollution and aquatic ecosystems. Questions that addressed aspects of appraisal, particularly incorporation of ecological service values into decision making, were also strongly represented. The findings revealed that sustainability has entered the lexicon of the UK water sector, but much remains to be done to embed the concept operationally, with key sustainability issues such as resilience and interaction with related key sectors, such as energy and agriculture, relatively poorly addressed. However, the exercise also revealed that a necessary condition for sustainable development, effective communication between scientists, practitioners and policy makers, already appears to be relatively well established in the UK water sector.

Current diagnosis, management and morbidity in paediatric inflammatory bowel disease.

UNLABELLED: In the 1970s several reports highlighted the long delay in diagnosis often experienced by children with Crohn's disease. In recent years this disorder has attracted much publicity, and many believe that the incidence has increased substantially. The aim of this investigation was to determine whether heightened awareness had shortened the interval to diagnosis, improved clinical management and reduced morbidity. A retrospective study was therefore carried out on 112 children with inflammatory bowel disease (64 Crohn's disease, 41 ulcerative colitis, 7 indeterminate colitis) referred to a paediatric gastroenterology department in the UK between 1994 and 1998. In Crohn's disease the median interval to diagnosis was 47 wk (maximum 7 y). In those without diarrhoea this was longer (66 vs 28 wk; p = 0.005). In ulcerative colitis the median interval was 20 wk (maximum 3 y). Even in severe colitis the median interval was 5.5 wk (range 3-9 wk) and 4 required urgent colectomy soon after referral. Many with unrecognized Crohn's disease had undergone inappropriate treatments, such as growth hormone or psychiatric therapy. Nineteen (17%) had undergone endoscopic investigations in adult units prior to referral. Malnutrition was equally common in Crohn's disease and ulcerative colitis (11%). Short stature was present in 19% with Crohn's disease, and 5% with ulcerative colitis, and was severe in 8% with Crohn's disease. There was a significant correlation between symptom duration and the degree of growth impairment present (r(s) = -0.4; p = 0.004). CONCLUSION: This study suggests that late diagnosis and inappropriate investigation and management are still significant problems.

Goal orientations, self-determination and pupils' discipline in physical education.

In this study, we examined the patterns of goal orientations, perceived competence, reasons for behaving well and self-reported discipline in school physical education among 511 pupils from years 8 and 9 (mean age 14.2 years). Cluster analyses were conducted on two randomly split subsamples to identify homogeneous groups of pupils on these measures of achievement motivation and discipline. Three meaningful clusters emerged for the first subsample, which were then cross-validated for the second subsample. One group of pupils revealed low scores on task and ego orientations, perceived competence and feelings of self-determination about their behaviour in lessons. These perceptions were related to lower ratings of discipline in physical education than pupils who scored more highly on these variables. The highest discipline scores were reported by pupils with high task and ego orientations, perceived competence and feelings of autonomy. The results are useful for teachers and other physical activity leaders in enhancing motivation and disciplined behaviour in young people. Promoting more self-determined reasons for being disciplined, for example, could lead to more orderly classes.

Gibberellin biosynthesis: metabolic evidence for three steps in the early 13-hydroxylation pathway of rice.

[14C4]GA53, [14C4]GA44, and [2H2/14C4]GA19 were injected separately into seedlings of rice (Oryza sativa) using a dwarf mutant (d35) that has low levels of endogenous gibberellins (GAs). After 8 h incubation, the shoots were extracted and the labeled metabolites were identified by full-scan gas chromatography mass spectrometry (GC-MS) and Kovats retention indices (KRIs). Our results document the metabolic sequence, GA53-->GA44-->GA19-->GA20 and the presence of endogenous GA53, GA44, GA19, GA20 and GA1. Previous metabolic studies have shown the presence of the step, GA20-->GA1 in rice. Taken together, the data establish in vegetative shoots of rice the presence of the early 13-hydroxylation pathway, a pathway that originates from GA12 and leads to bioactive GA1.

Investigation and outcome of neonatal hepatitis in infants with hypopituitarism.

UNLABELLED: Congenital hypopituitarism is a recognized cause of neonatal hepatitis, but the diagnosis may be difficult to establish even if clinically suspected. In order to determine the natural history of this disorder, the outcome of 12 infants with neonatal hepatitis secondary to hypopituitarism is reviewed. The clinical diagnosis of hypopituitarism was established on a combination of features, which include dysmorphism (4 infants), optic nerve hypoplasia (8 infants), micropenis (5 male infants) and recurrent hypoglycaemia (blood glucose < 2.4 mmol/l (8 infants)). Endocrine investigation revealed low free thyroxine (T4) levels (< 10 pmol/l), with normal thyroid stimulating hormone (TSH) levels (0.4-4.5 mU/l) (11 infants), and serum cortisol levels which were inappropriately low (< 200 nmol/l). In 9 of 12 infants, liver disease resolved within 6 wk following treatment with thyroxine, hydrocortisone and, where appropriate, growth hormone, including Cases 9 and 1 in whom diagnosis and treatment were delayed until 3 mo and 3 y of age, respectively. Liver disease resolved spontaneously in two infants prior to starting hormone replacement therapy (Cases 11, 12), and one male infant (Case 10), in whom the diagnosis and hormone replacement therapy were delayed until 5 y of age, developed cirrhosis and portal hypertension and later underwent liver transplantation. CONCLUSION: The diagnosis of hypopituitarism should always be considered in infants with unexplained neonatal hepatitis. Delay in diagnosis and appropriate treatment was associated with persistently abnormal liver function tests and may lead to irreversible liver disease.

Surgical management of malignant melanoma using dynamic lymphoscintigraphy and gamma probe-guided sentinel lymph node biopsy: the Emory experience.

Sentinel lymph node (SLN) biopsy is revolutionizing the surgical management of primary malignant melanoma. It allows accurate nodal staging which targets patients who may benefit from regional lymphadenectomy and systemic therapy. This is a retrospective review of patients treated at Emory University for stage I and II malignant melanoma with gamma probe-guided SLN biopsy from 1/1/94 to 6/30/98. Three hundred sixty patients (males 228, females 132) were identified. Primary melanoma sites included: head and neck 58, trunk 148, and extremities 154 (upper 71, lower 83). Primary tumor staging was T1 9, T2 134, T3 153, and T4 64. SLNs were successfully identified in 99.7 per cent of patients and 98.9 per cent of nodal basins mapped. In 275 (76.6%) cases a single draining nodal basin was identified. In 84 (23.3%) cases there were multiple draining nodal basins. Positive SLNs were identified in 63 patients (17.5%). SLN positivity by tumor staging was T1 0 per cent, T2 9.0 per cent, T3 22.2 per cent, and T4 26.6 per cent. The overall recurrence rate was 11.9 per cent. Recurrences by SLN status were SLN+, 27 per cent, and SLN-, 8.8 per cent. Regional recurrence occurred in 7 (2.4%) of the 297 with negative SLN biopsies and 7 (11.1%) of the 63 with positive SLN biopsies. Dynamic lymphoscintigraphy and gamma probe-guided SLN localization was successful in more than 98 per cent of cases. Patients with negative SLN biopsies have a low risk of recurrence.

Predicting participation in noncompulsory physical education: do goal perspectives matter?

The purpose of this study was to examine relationships between personal and situational achievement goals and participation in an optional physical education program, utilising a prospective design. Pupils (N = 123) in their final year of compulsory schooling completed measures assessing goal orientations, perceptions of the motivational climate of their physical education class, perceived competence, and enjoyment of physical education. Those pupils (N = 76) who continued their full-time education at a sixth form college were later surveyed to determine whether they participated in physical education at college. Hierarchical discriminant function analysis showed that discrimination was possible between participants and nonparticipants in college physical education on perceptions of a prevailing mastery climate and enjoyment. Goal orientations, perceptions of a prevailing performance climate, and perceived competence did not contribute reliably to group separation. Participants at college were more likely to perceive a mastery environment and to enjoy lessons more than students who chose not to take part. These findings underscore the importance of promoting mastery-based criteria for success in classes, along with positive emotional reactions, if teachers wish to maximise involvement in noncompulsory physical education.

Management of malignant melanoma of the head and neck using dynamic lymphoscintigraphy and gamma probe-guided sentinel lymph node biopsy.

BACKGROUND: The sentinel lymph node (SLN) biopsy is revolutionizing the surgical management of primary malignant melanoma. It allows accurate nodal staging, and targets patients who may benefit from regional lymphadenectomy and systemic therapy; however, its use in the management of head and neck melanoma has not been widely accepted. METHODS: A retrospective review of patients treated for clinical stages I and II malignant melanoma of the head and neck with dynamic lymphoscintigraphy and gamma probe-guided SLN biopsy. RESULTS: Fifty-eight patients (47 male and 11 female) were identified. Primary melanoma sites included the scalp (21), ear (8), face (13), neck (15), and eyelid (1). Primary tumor staging was T2 (11), T3 (24), and T4 (23). Dynamic lymphoscintigraphy visualized SLNs in 57 patients (98.3%). In 43 cases (75%) a single draining nodal basin was identified, and in 14 cases there were multiple draining nodal basins. Sentinel lymph nodes were successfully identified in 72 (96%) of 75 nodal basins. Positive SLNs were identified in 10 patients (17.5%). Sentinal lymph node positivity by tumor staging was T3, 16.7% and T4, 27.3%. Completion lymphadenectomy revealed residual disease in 3 patients (30%). Relapse occurred in 10 (21.3%) of the 47 patients with negative SLN biopsy results and 7 (70%) of those with positive results. CONCLUSIONS: Gamma probe-guided SLN localization in the head and neck region was successful in 96% of draining nodal basins. It can target regional lymphadenectomy in patients who may benefit from regional nodal dissection.

Achievement goals, beliefs about the causes of success and reported emotion in post-16 physical education.

The main aim of this study was to examine whether goal orientations of male and female adolescents involved in an optional post-16 physical education (PE) programme were related in a conceptually consistent manner with their beliefs about the causes of success in PE. We also determined relationships between these achievement goal-belief dimensions and reported enjoyment and boredom within PE classes. Participants (n = 171) in a sixth-form college PE programme completed an inventory assessing their task and ego goal orientations, beliefs about the determinants of success in PE, and emotion in PE activities at college. Separate factor analyses of goal orientations and beliefs for male and female students revealed two goal-belief dimensions. The first dimension showed ego orientation was linked to the view that ability and deceptive tactics lead to success. The second dimension suggested task orientation was associated with the belief that success is the result of hard work and effort. This task goal-belief factor was found to be more strongly correlated with enjoyment in PE among female students than among males. For boys, the task goal-belief factor was correlated significantly and negatively with boredom in PE, but this was not the case for girls. No significant relationships emerged between the ego goal-belief factor and reported emotion in PE among the male and female participants. Facilitating task involvement and beliefs about causes of success that are fundamentally under personal control may, therefore, promote positive affective experiences in sixth-form PE, especially among female students.

The dwarf-1 (dt) Mutant of Zea mays blocks three steps in the gibberellin-biosynthetic pathway.

In plants, gibberellin (GA)-responding mutants have been used as tools to identify the genes that control specific steps in the GA-biosynthetic pathway. They have also been used to determine which native GAs are active per se, i.e., further metabolism is not necessary for bioactivity. We present metabolic evidence that the D1 gene of maize (Zea mays L.) controls the three biosynthetic steps: GA20 to GA1, Ga20 to GA5, and GA5 to GA3. We also present evidence that three gibberellins, GA1, GA5, and GA3, have per se activity in stimulating shoot elongation in maize. The metabolic evidence comes from the injection of [17-13C,3H]GA20 and [17-13C,3H]GA5 into seedlings of d1 and controls (normal and d5), followed by isolation and identification of the 13C-labeled metabolites by full-scan GC-MS and Kovats retention index. For the controls, GA20 was metabolized to GA1,GA3, and GA5; GA5 was metabolized to GA3. For the d1 mutant, GA20 was not metabolized to GA1, GA3, or to GA5, and GA5 was not metabolized to GA3. The bioassay evidence is based on dosage response curves using d1 seedlings for assay. GA1, GA3, and GA5 had similar bioactivities, and they were 10-times more active than GA20.

Gibberellin Metabolism in Maize (The Stepwise Conversion of Gibberellin A12-Aldehyde to Gibberellin A20.

The stepwise metabolism of gibberellin A12-aldehyde (GA12-aldehyde) to GA20 is demonstrated from seedling shoots of maize (Zea mays L.). The labeled substrates [13C,3H]GA12-aldehyde, [13C,3H]GA12, [14C4]GA53, [14C4/2H2]GA44, and [14C4/2H2]GA19 were fed individually to dwarf-5 vegetative shoots. Both [13C,3H]GA12-aldehyde and [13C,3H]GA12 were also added individually to normal shoots. The labeled metabolites were identified by full-scan gas chromatography-mass spectrometry and Kovats retention indices. GA12-aldehyde was metabolized to GA53-aldehyde, GA12, GA53, GA44, and GA19; GA12 was metabolized to 2[beta]-hydroxy-GA12, GA53, 2[beta]-hydroxyGA53, GA44, 2[beta]-hydroxyGA44, and GA19; GA53 was metabolized to GA44, GA19, GA20, and GA1; GA44 was metabolized to GA19; and GA19 was metabolized to GA20. These results, together with previously published data from this laboratory, document the most completely defined gibberellin pathway for the vegetative tissues of higher plants.

The Metabolism of Gibberellin A20 to Gibberellin A1 by Tall and Dwarf Mutants of Oryza sativa and Arabidopsis thaliana.

The purpose of this study was to demonstrate the metabolism of gibberellin A20 (GA20) to gibberellin A1 (GA1) by tall and mutant shoots of rice (Oryza sativa L.) and Arabidopsis thaliana (L.) Heynh. The data show that the tall and dx mutant of rice and the tall and ga5 mutant of Arabidopsis metabolize GA20 to GA1. The data also show that the dy mutant of rice and the ga4 mutant of Arabidopsis block the metabolism of GA20 to GA1. [17-13C,3H]GA20 was fed to tall and the dwarf mutants, dx and dy, of rice and tall and the dwarf mutants, ga5 and ga4, of Arabidopsis. The metabolites were analyzed by high-performance liquid chromatography and full-scan gas chromatography-mass spectrometry together with Kovats retention index data. For rice, the metabolite [13C]GA, was identified from tall and dx seedlings; [13C]GA1 was not identified from the dy seedlings. [13C]GA29 was identified from tall, dx, and dy seedlings. For Arabidopsis, the metabolite [13C]GA1 was identified from tall, ga5, and ga4 plants. The amount of [13C]GA1 from ga4 plants was less than 15% of that obtained from tall and ga5 plants. [13C]GA29 was identified from tall, ga5, and ga4 plants. [13C]GA5 and [13C]GA3 were not identified from any of the six types of plant material.

Metabolism and Biological Activity of Gibberellin A4 in Vegetative Shoots of Zea mays, Oryza sativa, and Arabidopsis thaliana.

[17-13C,3H]Gibberellin A4 (GA4) was injected into the shoots of tall (W23/L317), dwarf-1 (d1), and dwarf-5 (d5) Zea mays L. (maize); tall (cv Nipponbare), dwarf-x (dx), and dwarf-y (dy) Oryza sativa L. (rice); and tall (ecotype Landsberg erecta), ga4, and ga5 Arabidopsis thaliana (L.) Heynh. [13C]GA4 and its metabolites were identified from the shoots by full-scan gas chromatography-mass spectrometry and Kovats retention indices. GA4 was metabolized to GA1 in all nine genotypes. GA4 was also metabolized in some of the genotypes to 3-epi-GA1, GA2, 2[beta]-OH-GA2, 3-epi-GA2, endo-GA4, 16[alpha], 17-H2-16, 17-(OH)2-GA4, GA34, endo-GA34, GA58, 15-epi-GA63, GA71, and 16-epi-GA82. No evidence was found for the metabolism of GA4 to GA7 or of GA4 to GA3. The bioactivities of GA4 and GA1 were determined using the six dwarf mutants for assay. GA4 and GA1 had similar activities for the maize and rice mutants. For the Arabidopsis mutants, GA4 was more active than GA1 at low dosages; GA4 was less active than GA1 at higher dosages.

Hydrolysis and reconjugation of gibberellin A20 glucosyl ester by seedlings of Zea mays L.

The [6-2H]glucosyl ester of [17-13C,3H]gibberellin A20 (GA20) was injected into light-grown 14-day-old seedlings of normal, dwarf-1, and dwarf-5 maize (Zea mays L.). The plant material was extracted 24 h later, and the extracts were purified by solvent partitioning, column chromatography, and HPLC. 13C-labeled metabolites were identified from the purified extracts by full-scan gas chromatography/mass spectrometry and selected ion current monitoring in conjunction with Kovats retention indices. The metabolites, [13C]GA20, [13C]GA29, [13C]GA20-13-O-glucoside, and [13C]GA29-2-O-glucoside, were identified from normal, dwarf-1, and dwarf-5 seedlings. [13C]GA8 and [13C]GA8-2-O-glucoside were also identified from normal and dwarf-5 seedlings but not from dwarf-1 seedlings. The data provide definitive evidence for the endogenous hydrolysis by the seedlings of the introduced conjugate and its reconjugation to three glucosides.

Metabolism of ent-Kaurene to Gibberellin A(12)-Aldehyde in Young Shoots of Normal Maize.

Young shoots of normal maize (Zea mays L.) were used to determine both the stepwise metabolism of ent-kaurene to gibberellin A(12)-aldehyde and the endogenous presence of the members in this series. Each of the five steps in the sequence was established by feeds of 17-(13)C, (3)H-labeled kauranoids to cubes from the cortex of elongating internodes, to homogenates from the cortex of elongating internodes, and/or to homogenates from dark-grown seedlings. The (13)C-metabolites were identified by Kovats retention indices (KRI) and full-scan capillary gas chromatography-mass spectrometry (GC-MS). Five substrates and the final product in this sequence were shown to be native by the isotopic dilution of 17-(13)C, (3)H-labeled substrates added as internal standards to extracts obtained from elongating internodes. Evidence for the isotopic dilution was obtained by KRI and full-scan capillary GC-MS. Thus, we document the presence in young maize shoots of the metabolic steps, ent-kaurene --> ent-kaurenol --> ent-kaurenal --> ent-kaurenoic acid --> ent-7 alpha-hydroxykaurenoic acid --> gibberellin A(12)-aldehyde.

Gibberellin A(3) Is Biosynthesized from Gibberellin A(20) via Gibberellin A(5) in Shoots of Zea mays L.

[17-(13)C,(3)H]-Labeled gibberellin A(20) (GA(20)), GA(5), and GA(1) were fed to homozygous normal (+/+), heterozygous dominant dwarf (D8/+), and homozygous dominant dwarf (D8/D8) seedlings of Zea mays L. (maize). (13)C-Labeled GA(29), GA(8), GA(5), GA(1), and 3-epi-GA(1), as well as unmetabolized [(13)C]GA(20), were identified by gas chromatography-selected ion monitoring (GC-SIM) from feeds of [17-(13)C, (3)H]GA(20) to all three genotypes. (13)C-Labeled GA(8) and 3-epi-G(1), as well as unmetabolized [(13)C]GA(1), were identified by GC-SIM from feeds of [17-(13)C, (3)H]GA(1) to all three genotypes. From feeds of [17-(13)C, (3)H]GA(5), (13)C-labeled GA(3) and the GA(3)-isolactone, as well as unmetabolized [(13)C]GA(5), were identified by GC-SIM from +/+ and D8/D8, and by full scan GC-MS from D8/+. No evidence was found for the metabolism of [17-(13)C, (3)H]GA(5) to [(13)C]GA(1), either by full scan GC-mass spectrometry or by GC-SIM. The results demonstrate the presence in maize seedlings of three separate branches from GA(20), as follows: (a) GA(20) --> GA(1) --> GA(8); (b) GA(20) --> GA(5) --> GA(3); and (c) GA(20) --> GA(29). The in vivo biogenesis of GA(3) from GA(5), as well as the origin of GA(5) from GA(20), are conclusively established for the first time in a higher plant (maize shoots).

The dominant non-gibberellin-responding dwarf mutant (D8) of maize accumulates native gibberellins.

The endogenous gibberellins (GAs) were examined from young vegetative shoots of the dominant mutant, Dwarf-8, a GA-nonresponder, and normal maize; GA(44), GA(17), GA(19), GA(20), GA(29), GA(1), and GA(8), members of the early-13-hydroxylation pathway, were identified from both kinds of shoots by full-scan mass spectra and Kovats retention indices. In addition, we report the identification of 3-epi-GA(1), GA(3), GA(4), GA(5), GA(7), GA(9), GA(12), GA(15), GA(24), GA(34), and GA(53) by using the same criteria. [1,7,12,18-(14)C(4)]GA(53) and -GA(44), [17-(2)H(2)]GA(19), and [17-(13)C,(3)H(2)]GA(20), -GA(29), -GA(1), -GA(8), and -GA(5) were used as internal standards to determine the endogenous levels of these GAs by measurement of isotope dilution, using capillary gas chromatography and selected ion monitoring. Shoots of Dwarf-8 accumulate relatively high levels of GA(20), GA(1), and GA(8). The accumulation of GA(1) appears to be related to gene dosage. Since Dwarf-8 contains the same pattern of GAs as normals (including GA(1) and GA(3)), the genetic control point probably lies after GA(1) (and GA(3)). Thus Dwarf-8 may be a GA receptor mutant or a mutant that controls a product downstream from the binding of the bioactive GA to a receptor.

Qualitative and Quantitative Analyses of Gibberellins in Vegetative Shoots of Normal, dwarf-1, dwarf-2, dwarf-3, and dwarf-5 Seedlings of Zea mays L.

Gibberellins A(12) (GA(12)), GA(53), GA(44), GA(19), GA(17), GA(20), GA(29), GA(1), and GA(8) have been identified from extracts of vegetative shoots of normal (wild type) maize using full scan capillary gas chromatography-mass spectrometry and Kovats retention indices. Seven of these gibberellins (GAs) have been quantified by capillary gas chromatography-selected ion monitoring using internal standards of [(14)C(4)]GA(53), [(14)C(4)]GA(44), [(2)H(2)] GA(19), [(13)C(1)]GA(20), [(13)C(1)]GA(29), [(13)C(1)]GA(1), and [(13)C(1)]GA(8). Quantitative data from extracts of normal, dwarf-1, dwarf-2, dwarf-3, and dwarf-5 seedlings support the operation of the early 13-hydroxylation pathway in vegetative shoots of Zea mays. These data support the positions in the pathway blocked by the mutants, previously assigned by bioassay data and metabolic studies. The GA levels in dwarf-2, dwarf-3, and dwarf-5 were equal to, or less than, 2.0 nanograms per 100 grams fresh weight, showing that these mutants are blocked for steps early in the pathway. In dwarf-1, the level of GA(1) was very low (0.23 nanograms per 100 grams fresh weight) and less than 2% of that in normal shoots, while GA(20) and GA(29) accumulated to levels over 10 times those in normals; these results confirm that the dwarf-1 mutant blocks the conversion of GA(20) to GA(1). Since the level of GAs beyond the blocked step for each mutant is greater than zero, each mutated gene probably codes for an altered gene product, thus leading to impaired enzyme activities.