[A rare case of intraductal tumor of the pancreas in which an intraductal tubulopapillary neoplasm was mixed with a widely spreading gastric-type intraductal papillary-mucinous neoplasm].

A 70-year-old man receiving treatment for diabetes mellitus presented with a cystic mass in the border area of the pancreatic body and tail on plain computed tomography (CT) due to impaired glucose intolerance. Contrast-enhanced CT showed a faint hyperattenuated nodular mass extending from the dilated main pancreatic duct (MPD) to the branch duct. Endoscopic retrograde cholangiopancreatography revealed a mildly dilated orifice of the papilla of Vater and MPD stenosis with entire upstream and immediate downstream dilatations. The patient underwent distal pancreatectomy due to the suspicion of mixed-type intraductal papillary-mucinous carcinoma. A pathological examination showed an intraductal solid-nodular mass measuring 25mm in length, consisting of two types of neoplasms. One showed tubulopapillary growth with entirely high-grade (HG) atypical cuboidal epithelium, in which immunohistochemical examinations were positive for MUC6 but negative for human gastric mucin (HGM), MUC1, MUC2, and MUC5AC, fitting the concept of intraductal tubulopapillary neoplasm (ITPN). The other showed the same growth of low-grade (LG) atypical columnar cells positive for HGM and MUC5AC and negative for MUC1 and MUC2, which corresponded to gastric-type intraductal papillary-mucinous neoplasm (IPMN) -LG. The tumor had not invaded the duct walls, and no metastatic lymph nodes were observed. The ITPN was adjacent to the IPMN mainly composed of tubular glands mimicking pyloric glands with LG dysplasia that corresponded to the so-called IPMN-pyloric gland variant. Moreover, the proliferation of low-papillary gastric-type IPMN spread around the intraductal tumors. Consequently, the patient was diagnosed with an intraductal tubular neoplasm comprising a noninvasive ITPN and gastric-type IPMN-LG. ITPN is a recently identified intraductal neoplasm of the pancreas proposed by Yamaguchi et al. and is distinguished by intraductal tubulopapillary growth with HG cellular atypia without overt mucin production, in contrast to IPMN. To date, no cases of intraductal nodular tumors comprising ITPN and IPMN have been reported. We report this original case with imaging and pathological observations and discuss potential processes via which ITPN and IPMN may arise adjacent to each other in the same pancreatic duct.

Necessity of pharyngeal anesthesia during transoral gastrointestinal endoscopy: a randomized clinical trial.

BACKGROUND/AIMS: The necessity for pharyngeal anesthesia during upper gastrointestinal endoscopy is controversial. This study aimed to compare the observation ability with and without pharyngeal anesthesia under midazolam sedation. METHODS: This prospective, single-blinded, randomized study included 500 patients who underwent transoral upper gastrointestinal endoscopy under intravenous midazolam sedation. Patients were randomly allocated to pharyngeal anesthesia: PA+ or PA- groups (250 patients/group). The endoscopists obtained 10 images of the oropharynx and hypopharynx. The primary outcome was the non-inferiority of the PA- group in terms of the pharyngeal observation success rate. RESULTS: The pharyngeal observation success rates in the pharyngeal anesthesia with and without (PA+ and PA-) groups were 84.0% and 72.0%, respectively. The PA- group was inferior (p=0.707, non-inferiority) to the PA+ group in terms of observable parts (8.33 vs. 8.86, p=0.006), time (67.2 vs. 58.2 seconds, p=0.001), and pain (1.21±2.37 vs. 0.68±1.78, p=0.004, 0-10 point visual analog scale). Suitable quality images of the posterior wall of the oropharynx, vocal fold, and pyriform sinus were inferior in the PA- group. Subgroup analysis showed a higher sedation level (Ramsay score ≥5) with almost no differences in the pharyngeal observation success rate between the groups. CONCLUSION: Non-pharyngeal anesthesia showed no non-inferiority in pharyngeal observation ability. Pharyngeal anesthesia may improve pharyngeal observation ability in the hypopharynx and reduce pain. However, deeper anesthesia may reduce this difference.

Solitary extrahepatic hepatocellular carcinoma in vertebrae without a primary lesion in the liver might originate from bone marrow: a case report and new hypothesis based on a review of the literature and the latest findings.

Solitary extrahepatic hepatocellular carcinoma without a primary lesion in the liver is rare and unique. In such patients, in addition to hepatocellular carcinoma, hepatoid adenocarcinoma, hepatoid teratoma, and hepatoid yolk sac tumor must be considered as differential diagnoses, and patients must be investigated in detail by histopathological studies with immunohistochemistry, especially using epithelial markers for which tumor cells are generally negative in hepatocellular carcinoma. A case with a solitary neoplasm of the vertebrae, which was diagnosed histopathologically as hepatocellular carcinoma, without a primary lesion is presented. The primary lesion was not identified even on autopsy, and the liver was pathologically almost normal. Given the review of the literature and circumstantial evidence, we would like to propose a bold new hypothesis that hepatocellular carcinoma might primarily originate from bone marrow.

Safety and efficacy of sorafenib followed by regorafenib or lenvatinib in patients with hepatocellular carcinoma.

AIM: Sequential administration of sorafenib followed by regorafenib or lenvatinib is effective against advanced hepatocellular carcinoma (HCC). In this study, we compared the safety profiles and anti-tumor effects of sequential sorafenib and regorafenib or lenvatinib therapy in patients with HCC. METHODS: We investigated adverse events, treatment responses and dose intensities in patients with HCC who were consecutively treated with sorafenib followed by regorafenib or lenvatinib at the individual level. RESULTS: Each group included 20 patients. The safety profiles of regorafenib and sorafenib were similar. The severity of hypophosphatemia, palmar-plantar erythrodysesthesia syndrome, and decreased neutrophil counts associated with regorafenib or sorafenib was similar in 12 patients. Conversely, the incidences and grades of adverse events differed between sorafenib and lenvatinib treatment. The anti-tumor effects of regorafenib and lenvatinib compared with sorafenib were significantly different for each patient. The response to treatment and progression-free survival were comparable for regorafenib and lenvatinib. The median relative dose intensities during the first 56 days of regorafenib and lenvatinib treatment were 83.6 and 80.0%, respectively. CONCLUSIONS: Similar adverse events were experienced by patients during consecutive treatment with sorafenib and regorafenib, which was not observed during treatment with sorafenib and lenvatinib. The obtained safety profile of sorafenib provided meaningful insights for selecting sequential therapy for patients with advanced HCC.

Clinical outcomes of the "resect and discard" strategy using magnifying narrow-band imaging for small (< 10 mm) colorectal polyps.

Background and study aim The "resect and discard" strategy is a new paradigm for the management of small colorectal polyps that reduces the cost and effort related to pathological diagnosis after polypectomy. This retrospective study aimed to clarify the clinical outcome of the "resect and discard" strategy for small colorectal polyps. Patients and methods The clinical records were reviewed from 501 consecutive patients who underwent the "resect and discard" strategy for colorectal polyps smaller than 10 mm at our hospital between January 2008 and December 2010. All colorectal lesions were evaluated onsite under magnifying narrow-band imaging after careful conventional white-light imaging. In cases of low grade adenoma predicted with high confidence, colonoscopists selected the "resect and discard" option without formal histopathology. The mid-term outcomes were evaluated to validate the curability of the "resect and discard" strategy. Results The present study included 501 consecutive patients with 816 lesions. The mid-term outcomes were examined for 476 (95 %) patients who received follow-up for at least 1 year after undergoing the "resect and discard" strategy. The median observation period was 83 months (range 12 - 117 months). No patient died from colorectal cancer related to the procedure, resulting in a disease-specific survival rate of 100 %. There were no local and/or distant recurrences detected during follow-up. Conclusions The "resect and discard" strategy for small colorectal polyps under strict preoperative diagnosis achieves excellent mid-term outcome.

Delineation of the extent of early gastric cancer by magnifying narrow-band imaging and chromoendoscopy: a multicenter randomized controlled trial.

BACKGROUND: Accurate delineation of tumor margins is necessary for curative resection of early gastric cancer (EGC). The objective of this multicenter, randomized, controlled study was to compare the accuracy with which magnifying narrow-band imaging (M-NBI) and indigo carmine chromoendoscopy delineate EGC margins. METHODS: Patients with EGC ≥ 10 mm undergoing endoscopic or surgical resection were enrolled. The oral-side margins of the lesions were first evaluated with conventional white-light endoscopy in both groups and then delineated by either chromoendoscopy or M-NBI. Biopsies were taken from noncancerous and cancerous mucosa, each at 5 mm from the margin. Accurate delineation was judged to have been achieved when the histological findings in all biopsy samples were consistent with endoscopic diagnoses. The primary end point was the difference in rate of accurate delineation between the two techniques. RESULTS: Data on 343 patients were analyzed. The accurate delineation rate (95 % confidence interval) was 85.7 % (80.4 - 91.0) in the chromoendoscopy group (n = 168), and 88.0 % (83.2 - 92.8) in the M-NBI group (n = 175; P = 0.63). Lower third tumor location (odds ratio [OR] 2.9; P = 0.01), nonflat macroscopic type (OR 4.4; P < 0.01), and high diagnostic confidence (OR 3.6; P < 0.001) were associated with accurate delineation, whereas use of M-NBI was not (OR 1.2; P = 0.39). Even after adjustment for identified confounders, the difference in accurate delineation between the groups was not significant (OR 1.0; P = 0.82). CONCLUSIONS: M-NBI does not offer superior delineation of EGC margins compared with chromoendoscopy; the two methods appear to be clinically equivalent.

Clinical predictors of histologic type of gastric cancer.

BACKGROUND AND AIMS: Gastric cancer is classified into differentiated and undifferentiated types according to the degree of glandular differentiation. Undifferentiated-type early gastric cancer (EGC) carries a higher risk of lymph-node metastasis than differentiated type, and therefore the indication criteria for endoscopic resection differ. This study aimed to clarify the ability of clinical predictors to distinguish between differentiated-type and undifferentiated-type EGCs. METHODS: This was a post hoc study of a multicenter prospective trial carried out in 5 Japanese hospitals, including 343 patients with cT1 EGC of ≥10 mm. According to the protocol, age, sex, and endoscopic findings of cancer (diameter, location, macroscopic type, and invasion depth) were evaluated, and the final diagnosis was confirmed from resected specimens. We evaluated the associations between these clinical factors and the histologic type of cancer and calculated the ability of the factors to diagnose differentiated-type EGC. The diagnostic ability of forceps biopsy was also calculated as a reference. RESULTS: Multivariate analysis identified older age (≥72 years), male sex, larger tumor size (>30 mm), elevated type, and shallower invasion depth (cT1a) as independent significant predictors for differentiated-type EGC, with elevated type showing the highest positive likelihood ratio. The sensitivity, specificity, accuracy, and positive and negative likelihood ratios of elevated type for differentiated-type EGC were 24%, 99%, 38%, 15.7, and 0.77, respectively, compared with 96%, 86%, 95%, 7.0, and 0.04 for forceps biopsy. CONCLUSIONS: Endoscopic elevated type is a significant predictor for differentiated-type EGC and may exclude undifferentiated-type EGC without the need for forceps biopsy.

Effectiveness of clip-and-snare method using pre-looping technique for gastric endoscopic submucosal dissection.

AIM: To evaluate efficacy and safety of clip-and-snare method using pre-looping technique (CSM-PLT) for gastric endoscopic submucosal dissection (ESD). METHODS: In the CSM-PLT method, a clip attached to the lesion side was strangulated with a snare, followed by application of an appropriate tension to the lesion independent of an endoscope. Twenty consecutive lesions were resected by ESD using CSM-PLT (CSM-PLT group) and compared with a control group, including 20 lesions that were resected by conventional ESD. The control group was matched based on the size and location of the lesion, presence of pathologic fibrosis, and experience of endoscopists. Total procedure time of ESD, proportion of en bloc resection, and complications were analyzed. RESULTS: The total procedure time for the CSM-PLT group was significantly shorter than that for the control group (38.5 min vs 59.5 min, P = 0.023); all lesions were resected en bloc by ESD. There was no significant difference in complications between the two groups. Moreover, there was no complication in the CSM-PLT group. In one large lesion (size: 74 mm) that underwent extensive CSM-PLT during ESD, we used an additional CSM-PLT on another edge of the lesion after achieving submucosal resection to the maximum extent possible during initial CSM-PLT. In two lesions, the snare came off the lesion together with the clip after a sudden pull; nevertheless, ESD was successful in all lesions. CONCLUSION: CSM-PLT was an effective and safe method for gastric ESD.

Importance of colonoscopy in patients undergoing endoscopic resection for superficial esophageal squamous cell carcinoma.

BACKGROUND: The aim of the study was to clarify the frequency of colorectal neoplasm (CRN) complicating superficial esophageal squamous cell carcinoma (ESCC) and the need for colonoscopy. METHODS: We retrospectively reviewed 101 patients who had undergone initial endoscopic resection (ER) for superficial ESCC. Control group participants were age- and sex-matched asymptomatic subjects screened at our hospital over the same period of time. Advanced adenoma was defined as an adenoma ≥10 mm, with villous features, or high-grade dysplasia. Advanced CRN referred to advanced adenoma or cancer. We measured the incidence of advanced CRN in superficial ESCC and controls, and we compared the characteristics of superficial ESCC patients with and without advanced CRN. RESULTS: In the superficial ESCC group, advanced CRNs were found in 17 patients (16.8%). A history of smoking alone was found to be a significant risk factor of advanced CRN [odds ratio 6.02 (95% CI 1.30-27.8), P=0.005]. CONCLUSION: The frequency of synchronous advanced CRN is high in superficial ESCC patients subjected to ER. Colonoscopy should be highly considered for most patients who undergo ER for superficial ESCC with a history of smoking, and is recommended even in superficial ESCC patients.

Impact of the clip and snare method using the prelooping technique for colorectal endoscopic submucosal dissection.

BACKGROUND AND STUDY AIM: Endoscopic submucosal dissection (ESD) is more difficult to perform for colorectal tumors, but the use of traction to facilitate direct submucosal layer visualization is a promising method to reduce procedure times and complication rates. This study aimed to evaluate the efficacy and safety of the clip and snare method (CSM) with a prelooping technique (PLT) for colorectal tumors. PATIENTS AND METHODS: A total of 140 colorectal tumors were treated: 17 using the CSM with PLT; 123, used as controls, by standard ESD without these techniques. Therapeutic efficacy and safety were retrospectively assessed. RESULTS: All tumors were successfully resected en bloc using the CSM with PLT. Multisegment resection was found in four tumors in the control group.  There was a significant difference in the procedure time between the patients in the CSM with PLT group and the control group (45.6 vs. 70.1 minutes; P = 0.047). There were no significant differences in the complication rates (5.9 % vs. 8.1 %; P = 1.00). CONCLUSIONS: The CSM with PLT was effective and safe in this study.

The "white globe appearance" (WGA): a novel marker for a correct diagnosis of early gastric cancer by magnifying endoscopy with narrow-band imaging (M-NBI).

BACKGROUND AND STUDY AIMS: Although magnifying endoscopy with narrow-band imaging (M-NBI) is useful for the diagnosis of gastric mucosal lesions, differentiating between early cancer (EC) and low grade adenoma (LGA) remains a challenge. During M-NBI examination, we have noted the presence of a small, white lesion with a globular shape underneath cancerous gastric epithelium, and have termed this endoscopic finding the "white globe appearance" (WGA). The aim of this study was to determine whether or not the WGA could be an endoscopic marker for distinguishing EC from LGA. METHODS: We retrospectively analyzed both the M-NBI scans and resected specimens of a total of 111 gastric lesions from 95 consecutive patients. Our main outcome was a difference in the prevalence of the WGA in EC and LGA. RESULTS: The prevalence of the WGA in EC and LGA was 21.5 % (20 /93) and 0 % (0 /18), respectively (P = 0.039). The sensitivity, specificity, positive predictive value, and negative predictive value for differentiating between EC and LGA, according to the presence of the WGA, were 21.5, 100, 100, and 19.8 %, respectively. CONCLUSION: A positive WGA in a suspicious lesion on M-NBI would be an adjunct to the M-NBI diagnosis of possible EC because the specificity and positive predictive value of the WGA for differentiating between EC and LGA were extremely high. The WGA could be a novel endoscopic marker for differentiating between EC and LGA.

Does long-term use of proton pump inhibitors cause hypomagnesaemia in Japanese outpatients?

AIM: Hypomagnesaemia has been reported associated with long-term proton pump inhibitor (PPI) treatment. However, there have been no studies in Japanese patients. We therefore assessed the effects of long-term PPI use on serum magnesium concentrations in Japanese patients. METHODS: Data from 481 outpatients who visited our centre in October and November 2011 were reviewed to determine their underlying diseases, oral medications, including PPIs, and serum magnesium concentrations. The association between PPI use and serum Mg concentrations was assessed by multiple linear regression analysis. RESULTS: Serum magnesium concentrations were significantly lower in PPI users (n=199) than in PPI non-users (n=282; 1.86±0.21 vs 1.91±0.19 mg/dL, p<0.01). Multiple linear regression analysis showed that the presence or absence of cirrhosis and PPI use was a significant predictor of magnesium concentration. In evaluating magnesium concentrations among PPI users and non-users with and without cirrhosis, we found that mean serum magnesium concentrations in patients with cirrhosis were significantly lower in PPI users than non-users (1.78±0.22 vs 1.87±0.22 mg/dL, p=0.03). CONCLUSIONS: Outpatients receiving long-term PPI treatment had significantly lower serum magnesium concentrations than those not treated with PPI. To the best of our knowledge, this study is the first to show hypomagnesaemia in Japanese patients with cirrhosis receiving long-term PPI treatment.

Detection of pharyngeal cancer in the overall population undergoing upper GI endoscopy by using narrow-band imaging: a single-center experience, 2009-2012.

BACKGROUND: Nonmagnifying observation by using narrow-band imaging (NBI) is useful for detecting pharyngeal lesions. Magnifying observation by using NBI can distinguish between cancerous and noncancerous lesions and is therefore useful for the early detection of pharyngeal cancer. OBJECTIVE: To evaluate the usefulness of observation of the pharynx by using NBI in the overall population undergoing upper GI endoscopy. DESIGN: Retrospective study. SETTING: Single tertiary referral center. PATIENTS: A total of 11,050 upper GI endoscopies between January 2009 and December 2012. INTERVENTIONS: Observation of the pharynx by using NBI. MAIN OUTCOME MEASURES: The rate of detection of pharyngeal cancer, the rates of detection according to the reason for endoscopy, and the types of cancers detected. RESULTS: Thirty-eight cancerous lesions were detected in 29 patients (0.26%, 29/11,050). The rate of detection of pharyngeal cancer was significantly higher in patients with a history of head and neck cancer (9.7%, 3/31) or a history of esophageal cancer (3.5%, 10/282). In patients undergoing endoscopy for screening, pharyngeal discomfort, and a history of gastric cancer, the rates of detection of pharyngeal cancer were 0.11% (10/8872), 1.1% (3/265), and 0.19% (3/1600), respectively. Two patients (6.9%) were female. One had a history of esophageal cancer, and the other had pharyngeal discomfort. LIMITATIONS: Single-center, retrospective study. CONCLUSIONS: Observation of the pharynx by using NBI in patients with previous head and neck cancer or esophageal cancer or who have pharyngeal discomfort is very important. Moreover, pharyngeal cancer was certainly found in the male patients undergoing screening endoscopy, although the rate was lower.

Can flat-type brownish microlesions in the orohypopharynx be followed up without biopsy or endoscopic resection?

BACKGROUND: Narrow-band imaging (NBI) is useful for detecting superficial oropharyngeal lesions. However, the diagnostic and treatment guidelines for NBI are not established. The aim of the present study was to evaluate the treatment strategy for these microlesions. METHODS: From October 2008 to September 2009, 68 flat-type brownish microlesions were observed in the orohypopharynx using NBI. Lesions were examined via magnifying NBI (M-NBI) and followed up without biopsy or endoscopic resection for >12 months. To clarify the characteristics, lesions were compared with the endoscopic characteristics of flat-type lesions diagnosed by biopsy and endoscopic resection as squamous cell carcinoma and high-grade intraepithelial neoplasia. RESULTS: The average diameter of the 68 lesions was 1.6 mm (range, 0.5-5 mm). At the 1-year follow up, 19 lesions had disappeared. No size increases or morphological changes wereobserved among 49 lesions followed for >1 year. At 2 years, 10 patients had dropped out and 11 lesions had disappeared. No changes were observed among 28 lesions followed for >2 years. Of the flat-type lesions as squamous cell carcinoma and high-grade intraepithelial neoplasia, a distinct border and irregular distribution of atypical vessels were observed in all cases using M-NBI. These findings were observed in two of 68 flat-type brownish microlesions during follow up. CONCLUSION: Although there is some possibility of squamous cell carcinoma or high-grade intraepithelial neoplasia, flat-type microlesions of ≤5 mm diameter in the orohypopharynx may be followed for up to 2 years without biopsy or endoscopic resection.

Use of transoral endoscopy for pharyngeal examination: cross-sectional analysis.

BACKGROUND AND AIM: Transoral endoscopy with narrow band imaging (NBI) is useful for identifying early-stage head and neck cancer. However, the screening capability of transoral upper gastrointestinal endoscopy has not yet been systematically evaluated. We evaluated the usefulness of transoral upper gastrointestinal endoscopy for pharyngeal examination. METHODS: This cross-sectional study evaluated 480 patients. All endoscopic pharyngeal examinations with NBI were carried out in accordance with prescribed procedures, consisting of 10 images each and all images were assessed by a blinded reviewer. We examined the association between the diagnostic usefulness of pharyngeal examination and other factors. RESULTS: Median subject age was 64 years (range 22-90 years), and 64% were male. Almost all patients (98%) had an Eastern Cooperative Oncology Group Performance Status of 0 or 1.Butylscopolamine bromide was given to 382 patients (80%), and a sedative was given to 460 (96%) patients. Median observation time was 74 s (range, 16-362 s), resulting in a mean of 9.0 usable images per patient. However, photographs of the right and left pyriform sinuses were consistently poor. Ordered logistic regression analysis showed that quality images were positively correlated with increased patient age. CONCLUSIONS: Transoral endoscopic examination was possible in most patients for screening of the head and neck. However, results were poor in the pyriform sinuses, indicating that additional improvements of examination methods and instruments are needed to enhance screening accuracy.

Hypomagnesemia associated with a proton pump inhibitor.

Severe hypomagnesemia is a serious clinical condition. Proton pump inhibitor (PPI) induced hypomagnesemia has been recognized since 2006. In March 2011 the U.S. Food and Drug Administration advised that long-term use of PPI can induce hypomagnesemia. We report the first Japanese case of hypomagnesemia associated with chronic use of PPIs in a 64-year-old man hospitalized for nausea, bilateral ankle arthritis, and tremor of the extremities who had convulsions 3 days after admission. Blood analysis showed severe hypomagnesemia. He had been taking rabeprazole (10 mg/day) for 5 years. After stopping rabeprazole and correcting the electrolytes imbalances, his symptoms improved without recurrence.

[A case of disseminated carcinomatosis of bone marrow treated by S-1 and cisplatin after distal gastrectomy for early gastric cancer].

Patients with bone metastasis originating from gastric cancer experience complications from DIC. They are treated with anticoagulation therapy or platelet transfusion, but their prognosis is poor. Our case was a 50-year-old male who had undergone distal gastrectomy for early gastric cancer[pT1a(M)N0M0, pStage I a]ten years previously. He was admitted to our hospital complaining of backache. As a result of his examination, he was diagnosed with disseminated carcinosis of bone marrow with DIC as a postoperative recurrence of gastric cancer. The patient was treated with combination chemotherapy of S-1 and cisplatin(S-1 80 mg/body, po, day 1-21 and cisplatin 50mg/body, iv, day 8). After one course of treatment, DIC was resolved and his pain was relieved. He survived for about nine months. S-1 and cisplatin are considered to be effective for disseminated carcinosis of bone marrow.

Granule-bound starch synthase I is responsible for biosynthesis of extra-long unit chains of amylopectin in rice.

A rice Wx gene encoding a granule-bound starch synthase I (GBSSI) was introduced into the null-mutant waxy (wx) rice, and its effect on endosperm starches was examined. The apparent amylose content was increased from undetectable amounts for the non-transgenic wx cultivars to 21.6-22.2% of starch weight for the transgenic lines. The increase was in part due to a significant amount of extra-long unit chains (ELCs) of amylopectin (7.5-8.4% of amylopectin weight), that were absent in the non-transgenic wx cultivars. Thus, actual amylose content was calculated to be 14.9-16.0% for the transgenic lines. Only slight differences were found in chain-length distribution for the chains other than ELCs, indicating that the major effect of the Wx transgene on amylopectin structure was ELC formation. ELCs isolated from debranched amylopectin exhibited structures distinct from amylose. Structures of amylose from the transgenic lines were slightly different from those of cv. Labelle (Wx(a)) in terms of a higher degree of branching and size distribution. The amylose and ELC content of starches of the transgenic lines resulted in the elevation of pasting temperature, a 50% decrease in peak viscosity, a large decrease in breakdown and an increase in setback. As yet undetermined factors other than the GBSSI activity are thought to be involved in the control of formation and/or the amount of ELCs. Structural analysis of the Wx gene suggested that the presence of a tyrosine residue at position 224 of GBSSI correlates with the formation of large amounts of ELCs in cultivars carrying Wx(a).

The N-terminal region of the starch-branching enzyme from Phaseolus vulgaris L. is essential for optimal catalysis and structural stability.

Starch-branching enzymes (SBEs) play a pivotal role in determining the fine structure of starch by catalyzing the syntheses of alpha-1,6-branch points. They are the members of the alpha-amylase family and have four conserved regions in a central (beta/alpha)8 barrel, including the catalytic sites. Although the role of the catalytic barrel domain of an SBE is known, that of its N- and C-terminal regions remain unclear. We have previously shown that the C-terminal regions of the two SBE isozymes (designated as PvSBE1 and PvSBE2) from kidney bean (Phaseolus vulgaris L.) have different roles in branching enzyme activity. To understand the contribution of the N-terminal region to catalysis, six chimeric enzymes were constructed between PvSBE1 and PvSBE2. Only one enzyme (1Na/2Nb)-II, in which a portion of the N-terminal region of PvSBE2 was substituted by the corresponding region of PvSBE1, retained 6% of the PvSBE2 activity. The N-terminal truncated form (DeltaN46-PvSBE2), lacking 46 N-terminal residues of PvSBE2, lost enzyme activity and stability to proteolysis. To investigate the possible function of this region, three residues (Asp-15, His-24, and Arg-28) among these 46 residues were subjected to site-directed mutagenesis. The purified mutant enzymes showed nearly the same K(m) values as PvSBE2 but had lower V(max) values and heat stabilities than PvSBE2. These results suggest that the N-terminal region of the kidney bean SBE is essential for maximum enzyme activity and thermostability.

Angiotensin I-converting enzyme inhibitory peptides isolated from tofuyo fermented soybean food.

Angiotensin I-converting enzyme (ACE) inhibitory activity was observed in a tofuyo (fermented soybean food) extract with an IC(50) value of 1.77 mg/ml. Two ACE inhibitors were isolated to homogeneity from the extract by adsorption and gel filtration column chromatography, and by reverse-phase high-performance liquid chromatography (HPLC). The purified substances reacted with 2,4,6-trinitrobenzensulfonic acid sodium salt. The amino acid sequences of these inhibitors determined by Edman degradation were Ile-Phe-Leu (IC(50), 44.8 microM) and Trp-Leu (IC(50), 29.9 microM). The Ile-Phe-Leu sequence is found in the alpha- and beta-subunits of beta-conglycinin, while the Trp-Leu sequence is in the B-, B1A- and BX-subunits of glycinin from soybean. Both of the peptides are non-competitive inhibitors. The inhibitory activity of Trp-Leu was completely preserved after a treatment with pepsin, chymotrypsin or trypsin. Even after successive digestion by these gastrointestinal proteases, the activity remained at 29% of the original value.