A TrmBL2-like transcription factor mediates the growth phase-dependent expression of halolysin SptA in a concentration-dependent manner in Natrinema gari J7-2.

To cope with a high-salinity environment, haloarchaea generally employ the twin-arginine translocation (Tat) pathway to transport secretory proteins across the cytoplasm membrane in a folded state, including Tat-dependent extracellular subtilases (halolysins) capable of autocatalytic activation. Some halolysins, such as SptA of Natrinema gari J7-2, are produced at late-log phase to prevent premature enzyme activation and proteolytic damage of cellular proteins in haloarchaea; however, the regulation mechanism for growth phase-dependent expression of halolysins remains largely unknown. In this study, a DNA-protein pull-down assay was performed to identify the proteins binding to the 5'-flanking sequence of sptA encoding halolysin SptA in strain J7-2, revealing a TrmBL2-like transcription factor (NgTrmBL2). The ΔtrmBL2 mutant of strain J7-2 showed a sharp decrease in the production of SptA, suggesting that NgTrmBL2 positively regulates sptA expression. The purified recombinant NgTrmBL2 mainly existed as a dimer although monomeric and higher-order oligomeric forms were detected by native-PAGE analysis. The results of electrophoretic mobility shift assays (EMSAs) showed that NgTrmBL2 binds to the 5'-flanking sequence of sptA in a non-specific and concentration-dependent manner and exhibits an increased DNA-binding affinity with the increase in KCl concentration. Moreover, we found that a distal cis-regulatory element embedded in the neighboring upstream gene negatively regulates trmBL2 expression and thus participates in the growth phase-dependent biosynthesis of halolysin SptA. IMPORTANCE: Extracellular proteases play important roles in nutrient metabolism, processing of functional proteins, and antagonism of haloarchaea, but no transcription factor involved in regulating the expression of haloaechaeal extracellular protease has been reported yet. Here we report that a TrmBL2-like transcription factor (NgTrmBL2) mediates the growth phase-dependent expression of an extracellular protease, halolysin SptA, of haloarchaeon Natrinema gari J7-2. In contrast to its hyperthermophilic archaeal homologs, which are generally considered to be global transcription repressors, NgTrmBL2 functions as a positive regulator for sptA expression. This study provides new clues about the transcriptional regulation mechanism of extracellular protease in haloarchaea and the functional diversity of archaeal TrmBL2.

Isolation and quantification of a 17-epi-dammarane triterpenoid rhuslactone from roots of Rhus chinensis and its preventive effects on coronary heart disease and thrombosis in zebrafish / 中国中药杂志

Based on mass spectrometry(MS)-guided separation strategy, compound 1 was obtained from the roots of Rhus chinensis. By comprehensive analysis of high resolution-electrospray ionization-mass spectrometry(HR-ESI-MS), nuclear magnetic resonance(NMR) data, and quantum chemical calculation of NMR(qcc-NMR) parameters, compound 1 was elucidated as rhuslactone, a 17-epi-dammarane triterpenoid with a rare 17α-side chain. An HPLC-ELSD method for its quantification in R. chinensis was established and adopted for the quantification of rhuslactone in different batches of R. chinensis. Rhuslactone displayed a good linear relationship within the range of 0.021 3-1.07 μmol·mL~(-1 )(r=0.997 6), and the average recovery was 99.34% [relative standard deviation(RSD) 2.9%). Moreover, the results of the evaluation test of the preventive effects of rhusalctone on coronary heart disease(CHD) and thrombosis showed that rhuslactone(0.11 nmol·mL~(-1)) significantly alleviated heart enlargement and venous congestion and increased cardiac output(CO), blood flow velocity(BFV), and heart rate, thereby reducing thrombus formation in zebrafish with CHD. The effects of rhuslactone on CO and BFV were superior to that of digoxin(1.02 nmol·mL~(-1)), and its effect on improving heart rate was comparable to that of digoxin. This study provides experimental references for the isolation, identification, quality control, and application of rhuslactone from R. chinensis against CHD. It is worth mentioning that this study has discussed some omissions in the determination of the stereochemistry of C-17 in dammarane triterpenoids in the present coursebook Chemistry of Chinese Medicine and some research papers, that is, the compound may be 17-epi-dammarane triterpenoid. This paper has also proposed steps for the establishment of C-17 stereochemistry.

Sec-Dependent Secretion of Subtilase SptE in Haloarchaea Facilitates Its Proper Folding and Heterocatalytic Processing by Halolysin SptA Extracellularly.

In response to high-salt conditions, haloarchaea export most secretory proteins through the Tat pathway in folded states; however, it is unclear why some haloarchaeal proteins are still routed to the Sec pathway. SptE is an extracellular subtilase of Natrinema sp. strain J7-2. Here, we found that SptE precursor comprises a Sec signal peptide, an N-terminal propeptide, a catalytic domain, and a long C-terminal extension (CTE) containing seven domains (C1 to C7). SptE is produced extracellularly as a mature form (M180) in strain J7-2 and a proform (ΔS) in the ΔsptA mutant strain, indicating that halolysin SptA mediates the conversion of the secreted proform into M180. The proper folding of ΔS is more efficient in the presence of NaCl than KCl. ΔS requires SptA for cleavage of the N-terminal propeptide and C-terminal C6 and C7 domains to generate M180, accompanied by the appearance of autoprocessing product M120 lacking C5. At lower salinities or elevated temperatures, M180 and M120 could be autoprocessed into M90, which comprises the catalytic and C1 domains and has a higher activity than M180. When produced in Haloferax volcanii, SptE could be secreted as a properly folded proform, but its variant (TSptE) with a Tat signal peptide does not fold properly and suffers from severe proteolysis extracellularly; meanwhile, TSptE is more inclined to aggregate intracellularly than SptE. Systematic domain deletion analysis reveals that the long CTE is an important determinant for secretion of SptE via the Sec rather than Tat pathway to prevent enzyme aggregation before secretion. IMPORTANCE While Tat-dependent haloarchaeal subtilases (halolysins) have been extensively studied, the information about Sec-dependent subtilases of haloarchaea is limited. Our results demonstrate that proper maturation of Sec-dependent subtilase SptE of Natrinema sp. strain J7-2 depends on the action of halolysin SptA from the same strain, yielding multiple hetero- and autocatalytic mature forms. Moreover, we found that the different extra- and intracellular salt types (NaCl versus KCl) of haloarchaea and the long CTE are extrinsic and intrinsic factors crucial for routing SptE to the Sec rather than Tat pathway. This study provides new clues about the secretion and adaptation mechanisms of Sec substrates in haloarchaea.

Genetic Polymorphism of Y Chromosome Haplogroup D-M174 in East Asian Populations / 法医学杂志

Objective To explore the genetic polymorphism of Y chromosome D-M174 haplogroup and sub-haplogroups in East Asia. Methods The samples of 1 426 unrelated male individuals from East Asia were collected, and then 7 Y chromosome haplogroup D-M174 and the Y-SNP of its sub-haplogroups were detected with mini-sequencing. The 22 Y-STR genotypes were detected with DNA Typer™ Y26 kit. The haplogroup was analyzed using direct counting method, heatmap, phylogenetic cluster and network graph cluster, and then distribution of genetic polymorphism and the clustering relation between populations and samples of Y chromosome D haplogroup were discussed. Results Haplogroup D-M174 were distributed mostly among Tibetans （40.96%）and Japanese （35.71%）, while less or none were distributed among the surrounding areas of Tibet and other areas. Conclusion The geographical distribution of Y chromosome D-M174 haplogroup in East Asian populations has significant characteristics.

Application of the Plackett-Burman design to determine the main factors affecting the anti-oxidative activity of goat's milk casein hydrolyzed by Alcalase and papain.

BACKGROUND: Antioxidant peptides can scavenge excessive free radicals produced by the metabolism, main- tain a free radical balance and prevent aging and disease. Therefore, finding natural resources with strong antioxidant activity has become a new focus for research. METHODS: Single-factor experiments were used to investigate the effects of various factors (hydrolysis temperature, pH, substrate concentration, enzyme to substrate ratio (E/S), ratio of compound protease (EAlcalase/Epapain) and time on the preparation of antioxidant peptides by compound protease (Alcalase and papain) hydrolysis of goat’s milk casein. Then, a Plackett-Burman design was used to determine the sig- nificant factors for the preparation of antioxidant peptides by hydrolysis of goat’s milk casein with compound protease. RESULTS: The results of the single-factor experiments indicated that the highest anti-oxidative activity of pep- tides would be obtained at a hydrolysis temperature of 55°C, a pH of 7.5, a substrate concentration of 3.0%, an E/S of 4.0%, a ratio of compound protease of 1/3 and a hydrolysis time of 180 min. CONCLUSIONS: The main factors affecting the activity of anti-oxidative peptides hydrolyzed from goat’s milk casein with compound protease were determined by Plackett-Burman design and the results showed that temperature, E/S ratio and the ratio of compound protease had significant influences on the production of antioxidant peptides. This could provide a basis and reference for further optimization.

Halolysin SptA, a Serine Protease, Contributes to Growth-Phase Transition of Haloarchaeon Natrinema sp. J7-2, and Its Expression Involves Cooperative Action of Multiple Cis-Regulatory Elements.

Many haloarchaea produce extracellular subtilisin-like proteases (halolysins) during late log phase; however, the physiological function and regulatory mechanism of growth phase-dependent production of halolysins are unknown. Halolysin SptA, the major extracellular protease of Natrinema sp. J7-2, is capable of intracellular self-activation to affect haloarchaeal growth. Here, we report that deletion of sptA leads to loss of extracellular and intracellular protease activities against azocasein and/or suc-AAPF-pNA, as well as a change in growth-phase transition of the haloarchaeon. Our results suggest that SptA is important for strain J7-2 to enter the stationary and death phases. Deletion and mutational analyses of the 5'-flanking region of sptA revealed two partially overlapping, semi-palindromic sequences upstream of the TATA box act as positive and negative cis-regulatory elements, respectively, to mediate sptA expression in late log phase. Additionally, a negative cis-regulatory element covering WW motif and a distant enhancer contribute to the modulation of sptA expression. Our results demonstrate that SptA functions both extracellularly and intracellularly, and that sptA expression relies on the cooperative action of multiple cis-regulatory elements, allowing SptA to exert its function properly at different growth stages in strain J7-2.

Metabolite Profiling of a Zinc-Accumulating Rice Mutant.

Breeding crops with high zinc (Zn) density is an effective way to alleviate human dietary Zn deficiencies. We characterized a mutant Lilizhi (LLZ) accumulating at least 35% higher Zn concentration in grain than the wild type (WT) in hydroponic experiments. The mutant stored less Zn content in the root and transported more Zn to the grain. Metabolite profiling demonstrated that, with high Zn treatment, the contents of proline, asparagine, citric acid, and malic acid were enhanced in both LLZ and the WT, which were thought to be involved in Zn transport in rice. Furthermore, the contents of cysteine, allothreonine, alanine, tyrosine, homoserine, ß-alanine, and nicotianamine required for the production of many metal-binding proteins were specifically increased in LLZ. LLZ had higher capability of amino acid biosynthesis and metal cation transportation. The current research extends our understanding on the physiological mechanisms of Zn uploading into grain and provides references for further Zn biofortification breeding in rice.

Alternative Translation Initiation of a Haloarchaeal Serine Protease Transcript Containing Two In-Frame Start Codons.

UNLABELLED: Recent studies have shown that haloarchaea employ leaderless and Shine-Dalgarno (SD)-less mechanisms for translation initiation of leaderless transcripts with a 5' untranslated region (5' UTR) of <10 nucleotides (nt) and leadered transcripts with a 5' UTR of ≥10 nt, respectively. However, whether the two mechanisms can operate on the same naturally occurring haloarchaeal transcript carrying multiple potential start codons is unknown. In this study, the transcript of the sptA gene (encoding an extracellular serine protease of Natrinema sp. strain J7-2) was experimentally determined and found to contain two potential in-frame AUG codons (AUG(1) and AUG(2)) located 5 and 29 nt, respectively, downstream of the transcription start site. Mutational analysis revealed that both AUGs can function as the translation start codon for production of active SptA, although AUG(1) is more efficient than AUG(2) for translation initiation. Insertion of a stable stem-loop structure between the two AUGs completely abolished initiation at AUG(1) but did not affect initiation at AUG(2), indicating that AUG(2)-initiated translation does not involve ribosome scanning from the 5' end of the transcript. Furthermore, the efficiency of AUG(2)-initiated translation was not influenced by an upstream SD-like sequence. In addition, both AUG(1) and AUG(2) contribute to transcript stability, probably by recruiting ribosomes to protect the transcript against degradation. These data suggest that depending on which of two in-frame start codons is used, the sptA transcript can act as either a leaderless or a leadered transcript for SptA production in haloarchaea. IMPORTANCE: In eukaryotes and bacteria, alternative translation start sites contribute to proteome complexity and can be used as a functional mechanism to increase translation efficiency. However, little is known about alternative translation initiation in archaea. Our results demonstrate that leaderless and SD-less mechanisms can be used for translation initiation of the sptA transcript from two in-frame start codons, raising the possibility that in haloarchaea, alternative translation initiation on one transcript functions to increase translation efficiency and/or contribute to proteome complexity.

Optimization of the Purification Technology of Total Flavones in Ampelopsis grossedentala with Macropo-rous Resins by Central Composite Design-Response Surface Methodology / 中国药房

OBJECTIVE:To optimize the purification technology of total flavones from Ampelopsis grossedentala with macro-porous resins. METHODS:4 kinds of macroporous resins for the purification of total flavones from A. grossedentala were screened by using drug-loading amount,desorption rate,recovery and purification rate as indicators. Single factor test and central composite design-response surface methodology were used to optimized eluant mass fraction,adsorption time,flow rate of eluant,eluant pH and other factors of purification technology,and validation test was also conducted. RESULTS:D-101 macroporous resin was the best. The optimal condition was as follows as the concentration of sample solution 2 mg(by extract weight)/ml,the volume of sam-ple solution 1.1 BV,ethanol 86.0%,adsorption time 36.7 min,flow rate of eluant 3.81 BV/h,pH 7. In validation test,mass frac-tion of total flavones increased from 66.83% to 85.00% in validation test(RSD=0.15%,n=3),and were close to predicted val-ue(85.08%). CONCLUSIONS:Central composite design-response surface methodology is feasible and stable for the optimization of purification technology of total flavones from A. grossedentala with macroporous resins.

Synthetic multivalent glycopeptide-lipopeptide antitumor vaccines: impact of the cluster effect on the killing of tumor cells.

Multivalent synthetic vaccines were obtained by solid-phase synthesis of tumor-associated MUC1 glycopeptide antigens and their coupling to a Pam3 Cys lipopeptide through click reactions. These vaccines elicited immune responses in mice without the use of any external adjuvant. The vaccine containing four copies of a MUC1 sialyl-TN antigen showed a significant cluster effect. It induced in mice prevailing IgG2a antibodies, which bind to MCF-7 breast tumor cells and initiate the killing of these tumor cells by activation of the complement-dependent cytotoxicity complex.

Self-adjuvanting synthetic antitumor vaccines from MUC1 glycopeptides conjugated to T-cell epitopes from tetanus toxoid.

The T-helper epitope peptide P30 (green in the scheme) from tetanus toxoid was used as the immunostimulant in MUC1 glycopeptide antitumor vaccines and apparently also acts as a built-in adjuvant. P30-conjugated glycopeptide vaccines containing three glycans in the immunodominant motifs PDTRP and GSTAP induced much stronger immune responses and complement dependent cytotoxicity mediated killing of tumor cells when applied in plain PBS solution without complete Freund's adjuvant.

Characterizing the assembly behaviors of human amylin: a perspective derived from C-terminal variants.

The differences in the C-terminal domains of human amylin peptide variants initiate different aggregation processes and differences in the composition of the aggregates by affecting the hydrophobic cores, conformations, and intra-sheet interactions of the peptides, which have distinct effects on the cytotoxicity of the peptides.

Inflammatory-modulating mechanism of transcutaneous electrical acupoint stimulation combined with general anesthesia or controlled hypotension on postoperative hippocampal neuroprotection / 中国针灸

<p><b>OBJECTIVE</b>To observe the effects of transcutaneous electrical acupoint stimulation (TEAS) combined with general anesthesia or controlled hypotension on hippocampal neuronal damage and the inflammatory response in peripheral circulation and central nervous system (CNS) after surgery, and to investigate its brain protection mechanism.</p><p><b>METHODS</b>Eighteen healthy male beagles aged 6 - 8 months were randomly divided into a general anesthesia group (group G), a controlled hypotension group (group C) and a compound anesthesia acupuncture group (group A), 6 cases in each group. Dogs in group G was anesthetized by isoflurane inhalation, and group C was combined with intravenous infusion of sodium nitroprusside based on isoflurane inhalation to induce hypotension, and followed surgery after achieving the target blood pressure, and group A was combined with TEAS at "Quchi" (LI 11), "Hegu" (LI 4) "Zu sanli" (ST 36) and "Sanyinjiao" (SP 6) based on controlled hypotension, and then brain tissue was taken out on the 72 h after mean arterial pressure (MAP) was returned to baseline levels. The concentration of IL-1beta,TNF-alpha in serum at different time points were detected by ELISA. The expression of IL-1beta, TNF-alpha, Bcl-2, Bax and cleaved caspase-3 were measured by immunohistochemistry, and the apoptosis of hippocampus were detected by TUNEL.</p><p><b>RESULTS</b>(1) At different time points, the concentration of TNFalpha showed the trend of increase first and then decrease, while IL-1beta concentration represented a trend of decrease first and then increase in both group C and group A, but there were no significant differences in cytokine expression between the two groups (all P > 0.05). (2) The ratio of positive cells of IL-1beta, TNF-alpha and caspase-3 in CA1 and CA3 of hippocampus in both group C and A were higher than those in group G (all P < 0.01), and cytokines expression in group A were lower than those in group C (all P < 0.01), and caspase-3 in CA1 in group A was lower than that in group C (P < 0.01). The ratio of Bcl-2/Bax in both group C and A were lower than that in group G (all P < 0.01), and that in group A was higher than that in group C (P < 0.01 in CA1, P < 0.05 in CA3). (3) The apoptosis index (AI) of hippocampal neurons in both group C and A was significantly higher than that in group G (P < 0.01), while AI in CA1 in group A was lower than that in group C (P < 0.01).</p><p><b>CONCLUSION</b>The TEAS can regulate the expression of inflammatory factor in hippocampus in animals undergoing general anesthesia or con trolled hypotension surgery, further improving Bcl-2/Bax ratio, inhibiting the expression of caspase-3 and reducing neuron apoptosis in hippocampus so as to play a neuroprotection.</p>

Co-assembly of human islet amyloid polypeptide (hIAPP)/insulin.

The pathogenesis of type II diabetes can be linked to cosecreted hIAPP/insulin interacting with cell membranes. Here we investigate the nanostructures by co-assembling hIAPP and insulin on surfaces. By tuning the hIAPP/insulin ratio, atomic force microscopy reveals the resulting nanostructure morphology changes from fibrils to oligomers, to annular. Implications for in vivo studies are discussed.

Effect on therapeutic effect of inducing the formation of the post-moxibustion sore for bronchial asthma / 中国针灸

<p><b>OBJECTIVE</b>To observe the impact of inducing the formation of the post-moxibustion sore on the efficacy of bronchial asthma treated with scarring moxibustion.</p><p><b>METHODS</b>Three hundred and seventy-two cases diag nosed definitely as bronchial asthma at remission stage were randomly divided into a modified nursing group (248 cases) and a conventional nursing group (124 cases). The scarring moxibustion was applied at Dazhui (GV 14), Feishu (BL 13), Danzhong (CV 17) and Tiantu (CV 22) in either group. The direct moxibustion with moxa cone was adopted. In modified nursing group, 0.5% Iodine was used for the sterilization at moxa abscess. The herbal plaster was cut into an inverted triangle and compressed on the wound. After suppuration, the fester was not cleaned in each dressing change. Additionally, the patients were advised to have high-protein diets after moxibus tion till abscess dropped. In conventional nursing group, 0.5% Iodine was used for the sterilization at moxa abscess. The herbal plaster was cut into a round shape that could cover completely the moxa wound. After suppuration, in each dressing change, the wound was cleaned and sterilized. Additionally, the patients were advised to avoid any stimulating food after moxibustion till abscess dropped. The clinical efficacy, the change of C3 content in blood serum and the clinical symptom score were observed in two groups.</p><p><b>RESULTS</b>The total effective rate was 93.5% (232/248) in modified nursing group, which was better than 79.0% (98/124) in conventional nursing group (P < 0.01). C3 content increased apparently after treatment as compared with that before treatment in either group (both P < 0.01), but C3 level increased much more apparently in modified nursing group as compared with that in conventional nursing group (P < 0.01). The clinical symptom score reduced apparently after treatment in either group (both P < 0.01), but that reduced much more apparently in modified nursing group as compared with that in conventional nursing group (P < 0.01).</p><p><b>CONCLUSION</b>In the treatment of bronchial asthma with scarring moxibustion, to induce the formation of the post-moxibustion sore achieves the better clinical efficacy as compared with conventional nursing.</p>

Hybrid peptides attenuate cytotoxicity of beta-amyloid by inhibiting its oligomerization: implication from solvent effects.

Abnormal assembly of monomeric beta-amyloid (Abeta) in Alzheimer's disease leads to the formation of most neurotoxic oligomers in vivo. In this study, we explored a linking strategy to design hybrid peptides, by combining the Abeta recognition motif and the solvent disruptive sequences. We found that in vitro all synthetic peptides with the recognition motif can affect Abeta fibrillization and alter the morphology of Abeta aggregates variously, different from those without the recognition motif. The effects of peptides containing recognition motif on Abeta aggregation correlate with their abilities to change the surface tension of solutions. In addition, compounds with the recognition motif, not those without such motif, can inhibit cytotoxicity of Abeta in cell culture probably by decreasing the amount of toxic Abeta oligomers. These results indicate that recognition domain and solvent effect should be considered as important factors when designing molecules to target Abeta aggregation.

Sequestration of copper from beta-amyloid promotes selective lysis by cyclen-hybrid cleavage agents.

Decelerated degradation of beta-amyloid (Abeta) and its interaction with synaptic copper may be pathogenic in Alzheimer disease. Recently, Co(III)-cyclen tagged to an aromatic recognition motif was shown to degrade Abeta in vitro. Here, we report that apocyclen attached to selective Abeta recognition motifs (KLVFF or curcumin) can capture copper bound to Abeta and use the Cu(II) in place of Co(III) to become proteolytically active. The resultant complexes interfere with Abeta aggregation, degrade Abeta into fragments, preventing H2O2 formation and toxicity in neuronal cell culture. Because Abeta binds Cu in amyloid plaques, apocyclen-tagged targeting molecules may be a promising approach to the selective degradation of Abeta in Alzheimer disease. The principle of copper capture could generalize to other amyloidoses where copper is implicated.

Analysis on influence factors of dietary supplement used in population aged above 45 years in Beijing / 中华预防医学杂志

<p><b>OBJECTIVE</b>To explore influence factors of dietary supplement used in population aged above 45 years in Beijing.</p><p><b>METHODS</b>Utilizing the data of the survey of dietary supplement (DS) used in Beijing in 2006 was to investigate the influence factors by using multi-factorial logistic regression analysis.</p><p><b>RESULTS</b>All 2487 samples were included in the analysis. Sex, age, average income of each month for each member of the family, experiences of deficiency of nutrition, attitude to nutrition knowledge, attitude to the need of DS and city-or-rural resident had shown effects on DS using. The percentage of using DS in female was higher than that in male being 19.3% and 11.8% respectively. The percentage grew up as the age became older. The same trend appeared as the economic-status going up, but not as education level change. Use of DS was greater in urban population (16.9%) than in rural population (14.6%). The percentage of using DS in people who had or just been diagnosed as malnutrition/deficiency disease or chronic disease, or those who thought themselves having got these diseases only by their own feeling were higher than those did not.</p><p><b>CONCLUSION</b>People the female, the elder, or those having higher income, having experiences of deficiency of nutrition, are more interested in nutrition knowledge, and more positive in need of DS and those living in rural should be the target population of nutrition education for DS use.</p>

Effects of 2-n-nonyl-1,3-dioxolane as an enhanceron transdermal absorption of Salvia miltiorrhiza gel / 中国中药杂志

<p><b>OBJECTIVE</b>To choose the most suitable concentration of 2-n-nonyl-1,3-dioxolane as a penetration enhancer in tanshinone gel preparation.</p><p><b>METHOD</b>In vitro, transdermal absorption was studied using improved Frans equipment and rats skin. Tanshinone II A was tested by HPLC.</p><p><b>RESULT</b>The 4.0% concentration of 2-n-nonyl-1,3-dioxolane enhanced the transdermal absorption significantly in the preparation.</p><p><b>CONCLUSION</b>2-n-nonyl-1,3-dioxolane was a new effective permeaton enhancer.</p>