Influence of Subunit Interface Mutations on Kinetic and Dynamic Properties of Alkaline Phosphatase from E. coli

Mutations, replacing amino acids involved in the formation of hydrogen bonds between subunits of dimeric alkaline phosphatase, have been introduced. Influence of mutations on kinetic properties and structural stability of mutant enzymes was established. In addition, alterations in protein dynamic properties have been studied using room temperature phosphorescence. Kinetic properties of both mutant enzymes were virtually the same, differing from the wild type enzyme in the kcat value that was almost twice lower. Changes in protein dynamic properties of mutant proteins, compared to the wild type enzyme, did not parallel changes in kinetic properties suggesting that an alteration in the rigidity of the Trp109 environment is not responsible for the reduction of kinetic properties. Instead, combined kinetic and dynamic consequences of introduced mutations suggest that breaking of specific links, involved in transmission of conformational change, could be responsible for altered kinetic properties. (doi: 10.5562/cca2168

Stabilnost ružmarinske kiseline u vodenim ekstraktima biljaka iz porodice Lamiaceae tijekom simuliranog procesa probave

The present study compares the gastrointestinal stability of rosmarinic acid in aqueous extracts of thyme, winter savory and lemon balm with the stability of pure rosmarinic acid. The stability of rosmarinic acid was detected after two-phase in vitro digestion process (gastric and duodenal) with human gastrointestinal enzymes. The concentration of rosmarinic acid in undigested and digested samples was detected using HPLC-DAD. Results showed that gastrointestinal stability of pure rosmarinic acid was significantly higher than that of rosmarinic acid from plant extracts after both gastric and intestinal phases of digestion. Among plant extracts, rosmarinic acid was the most stable in lemon balm after gastric (14.10 %) and intestinal digestion phases (6.5 %). The temperature (37 °C) and slightly alkaline medium (pH=7.5) did not aff ect the stability of rosmarinic acid, while acid medium (pH=2.5) significantly decreased its stability (≥50 %). In addition, the stability rate of rosmarinic acid is influenced by the concentration of human gastrointestinal juices.U ovom je istraživanju uspoređena gastrointestinalna stabilnost ružmarinske kiseline u vodenim ekstraktima timijana, primorskog vriska i matičnjaka s onom čistog kemijskog spoja. Stabilnost ružmarinske kiseline određena je simuliranjem dvofaznog procesa probave (u želucu i dvanaesniku) pomoću enzima izoliranih iz probavnog sustava čovjeka. Koncentracija ružmarinske kiseline određena je tehnikom HPLC-DAD prije i nakon simulirane probave. Rezultati pokazuju da je gastrointestinalna stabilnost čiste ružmarinske kiseline bila bitno veća od one u vodenim ekstraktima biljaka. Najveća stabilnost ovog spoja izmjerena je u ekstraktu matičnjaka, i to 14,10 % nakon probave u želucu, te 6,53 % nakon probave u tankom crijevu. Temperatura simuliranog procesa probave (37 °C) te blago lužnata sredina (pH=7,5) nisu utjecali na stabilnost ružmarinske kiseline. Kiseli medij želuca (pH=2,5) znatno je utjecao na smanjenje stabilnosti spoja (≥50 %). Koncentracija probavnih sokova također je utjecala na stabilnost ružmarinske kiseline

NeuroD1 Gene and Interleukin-18 Gene Polymorphisms in Type 1 Diabetes in Dalmatian Population of Southern Croatia

Aim: To evaluate the frequency of known polymorphisms in the exon 2 of the NeuroD1 gene and in the interleukin (IL)-18 promoter region in patients with type 1 diabetes mellitus (T1DM) and in healthy control subjects in Dalmatia, Southern Croatia. Methods: A total of 134 unrelated patients (73 men and 61 women) and 132 consecutive unrelated healthy controls (61 men and 71 women) from the Dalmatian region of southern Croatia were recruited for the study. NeuroD1 genotypes (GG, GA, AA) were identified by means of polymerase chain reaction followed by restriction fragment length polymorphism (PCR/RFLP). IL-18 polymorphism in the position –137 of the promoter region was detected by using PCR sequence-specific primers. Results: Genotype distributions of both genes did not show significant difference between patients and controls. Conclusion: Our results suggest that NeuroD1 exon 2 and IL-18 promoter gene polymorphisms are not associated with development of T1DM susceptibility in the population of South Croatia. In addition to previously published positive correlations of these polymorphisms with development of T1DM among different world populations, our findings indicate the existence of ethnic variations in the association of these genes with disease development

Bioavailability of Oral Curcumin in Systematic Reviews: A Methodological Study

Curcumin is a natural compound found in turmeric that exhibits diverse biological activities. However, its poor bioavailability limits its therapeutic application, which has led to the development of various bioavailability-improved formulations. In this methodological study, we analyzed whether systematic reviews on curcumin considered the bioavailability of systemic oral curcumin formulations when synthesizing evidence from human clinical trials. A total of 171 systematic reviews published between 2003 and 2022 were included in the study. From the included studies, we extracted data on study characteristics; type of curcumin; methods; and reporting regarding bioavailability, funding, and conflict of interest. Our results show that systematic reviews rarely consider the variable bioavailability of tested curcumin formulations. Relevant statistical subgroup and/or sensitivity analyses were reported in the methods and results of only 3.5% and 6.4% of reviews, respectively. However, more reviews mentioned bioavailability in their discussion (57%) or conclusion (13%). The detailed analysis of the included systematic reviews suggests that there is broad recognition of product bioavailability as a crucial factor affecting the health effects of curcumin, which is not accompanied by adequate evidence synthesis. Therefore, the results of most systematic reviews on orally administered curcumin should be taken with caution

Methods Used for Enhancing the Bioavailability of Oral Curcumin in Randomized Controlled Trials: A Meta-Research Study

It is unknown how randomized controlled trials (RCTs) approach the problem related to curcumin bioavailability. We analyzed methods and reporting regarding the bioavailability of systemic oral curcumin used in RCTs. We searched PubMed on 12 September 2020, to find articles reporting RCTs that used curcumin as an intervention. We extracted data about trial characteristics, curcumin products used, methods for improving curcumin bioavailability, and mentions of curcumin bioavailability. We included 165 RCTs. The most common category of intervention was simply described as “curcumin” or “curcuminoids” without a commercial name. There were 107 (64%) manuscripts that reported that they used methods to enhance the oral bioavailability of curcuminoids used in their intervention; 25 different methods were reported. The most common method was the addition of piperine (23%). Phospholipidated curcumin, a combination of curcumin and turmeric oils, nanomicellar curcumin, and colloidal dispersion of curcumin were the next most common methods. Fourteen trials (8.4%) compared more than one different curcumin product; nine (7.9%) trials compared the bioavailability/pharmacokinetics of curcumin products. In conclusion, a high number of diverse methods were used, and very few trials compared different curcumin products. More studies are needed to explore the comparative bioavailability and efficacy of different curcumin products

NeuroD1 Gene and Interleukin-18 Gene Polymorphisms in Type 1 Diabetes in the Dalmatian Population of Southern Croatia

Aim: To evaluate the frequency of known polymorphisms in the exon 2 of the NeuroD1 gene and in the interleukin (IL)-18 promoter region in patients with type 1 diabetes mellitus (T1DM) and in healthy control subjects in Dalmatia, Southern Croatia. Methods: A total of 134 unrelated patients (73 men and 61 women) and 132 consecutive unrelated healthy controls (61 men and 71 women) from the Dalmatian region of southern Croatia were recruited for the study. NeuroD1 genotypes (GG, GA, AA) were identified by means of polymerase chain reaction followed by restriction fragment length polymorphism (PCR/RFLP). IL-18 polymorphism in the position –137 of the promoter region was detected by using PCR sequence-specific primers. Results: Genotype distributions of both genes did not show significant difference between patients and controls. Conclusion: Our results suggest that NeuroD1 exon 2 and IL-18 promoter gene polymorphisms are not associated with development of T1DM susceptibility in the population of South Croatia. In addition to previously published positive correlations of these polymorphisms with development of T1DM among different world populations, our findings indicate the existence of ethnic variations in the association of these genes with disease development

Bemisia tabaci MED Population Density as Affected by Rootstock-Modified Leaf Anatomy and Amino Acid Profiles in Hydroponically Grown Tomato

Bemisia tabaci is one of the most devastating pests in tomato greenhouse production. Insecticide resistance management for B. tabaci requires a novel approach that maximizes non-chemical methods for pest control. The aim of this study was to test the effects of rootstocks on B. tabaci populations in hydroponically grown tomato plants. In order to contribute to the better understanding of the mechanisms defining the attractiveness of plant to the aerial pest, the effects of rootstocks on leaf anatomy and the amino acid composition of phloem sap were assessed. A two-factorial experimental design was adopted using cultivars (rootstock cultivars and Clarabella) grown as either non-grafted or grafted with cultivar Clarabella as a scion. The rootstock cultivars included Arnold, Buffon, Emperador, and Maxifort. A reduction in B. tabaci density was observed using all rootstock cultivars. The number of adult individuals per leaf was 2.7–5.4 times lower on rootstock cultivars than on Clarabella. The number of large nymphs per square centimeter was at least 24% higher on non–grafted Clarabella compared with all other treatments. The leaf lamina thickness and mesophyll thickness were lower in self-grafted Clarabella than in non-grafted or in one grafted on rootstock cultivars; however, the extent of this reduction depended on the rootstock. The leaves with thinner laminae were generally less attractive to B. tabaci. Eighteen amino acids were detected in the exudates of phloem sap. In all treatments, the most abundant amino acid was γ-aminobutyric acid (GABA), followed by proline, serine, alanine, and histidine. The scion cultivar Clarabella was the most attractive to B. tabaci and had a higher content of leucine than did rootstock cultivars, and a higher content of lysine compared to Buffon and Maxifort. The features modified by rootstock such are changes in leaf anatomy can affect the attractiveness of plants to B. tabaci. Thus, the grafting of tomato could constitute a valuable tool in an integrated management strategy against this aerial pest