Withania somnifera is an important medicinal plant, which is used in traditional medicine to cure many diseases.Flavonoids were determined in the extracts of W. somnifera root (WSREt) and leaf (WSLEt). The amounts of total flavonoids found in WSREt and WSLEt were 530 and 520 mg/100 g dry weight (DW), respectively. Hypoglycaemic and hypolipidaemic effects of WSREt and WSLEt were also investigated in alloxan-induced diabetic rats. WSREt and WSLEt and the standard drug glibenclamide were orally administered daily to diabetic rats for eight weeks. After the treatment period, urine sugar, blood glucose, haemoglobin (Hb), glycosylated haemoglobin (HbA1C), liver glycogen, serum and tissues lipids, serum and tissues proteins, liver glucose-6-phosphatase (G6P) and serum enzymes like aspartate transaminase (AST), alanine transaminase (ALT), acid phosphatase (ACP) and alkaline phosphatase (ALP) levels were determined. The levels of urine sugar, blood glucose, HbA1C, G6P, AST, ALT, ACP, ALP, serum lipids except high density lipoprotein-bound cholesterol (HDL-c) and tissues like liver, kidney and heart lipids were significantly (p < 0.05) increased, however Hb, total protein, albumin, albumin:globulin (A:G) ratio, tissues protein and glycogen were significantly (p < 0.05) decreased in alloxan-induced diabetic rats. Treatment of the diabetic rats with WSREt, WSLEt and glibenclamide restored the changes of the above parameters to their normal level after eight weeks of treatment, indicating that WSREt and WSLEt possess hypoglycaemic and hypolipidaemic activities in alloxan-induced diabetes mellitus (DM) rats.

Staphylococcus aureusis a Gram-positive pathogen that causes a wide range of infections in humans and animals. Bacterial ghosts are nonliving, empty cell envelopes and are well represented as novel vaccine candidates. In this study, we examined the immunogenicity and protective efficacy ofS. aureusghosts (SAGs) against a virulent challenge in rats. Nonliving SAGs were generated by using the MIC of sodium hydroxide. The formation of a transmembrane lysis tunnel structure in SAGs was visualized by scanning electron microscopy. To investigate these SAGs as a vaccine candidate, rats were divided into four groups, A (nonimmunized control), B (orally immunized), C (subcutaneously immunized), and D (intravenously immunized). The IgG antibody responses were significantly stronger in the SAG-immunized groups than in the nonimmunized control group (P< 0.05). Moreover, a significant increase in the populations of CD4⁺and CD8⁺T cells was observed in all three immunized groups (P< 0.05). We also found that serum bactericidal antibodies were significantly elicited in the SAG-immunized groups (P< 0.05). Most importantly, the bacterial loads in the immunized groups were significantly lower than those in the nonimmunized control group (P< 0.01). These results suggest that immunization with SAGs induces immune responses and provides protection against a virulentS. aureuschallenge.

Identification of medicinal plants and naturally derived compounds as new natural antioxidant and antibacterial sources for topical acne treatment has long been important. To determine anti-Propionibacterium acnes activity and in vitro antioxidant activities, Sanguisorba officinalis L. root (SOR) was extracted with cold water (CWE), hot water (HWE), and methanol (ME), and each extract was fractionated successively with hexane, ethyl acetate (EA), and butanol to determine whether the activities could be attributed to the total phenolic, flavonoid, terpenoid, and condensed tannin contents. Pearson’s correlation coefficients were analyzed between the respective variables. The SOR CWE, HWE, ME, and their respective EA fractions showed anti-P. acnes activity based on the paper disc diffusion method on agar plates, minimum inhibitory concentration (MIC), and minimal bactericidal concentration (MBC). The MIC against P. acnes had a moderate (+) correlation with the total phenolic content, but not with the other measures. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging capacity (SC) had a strong (–) correlation with the total phenolic content and a moderate (–) correlation with the total flavonoid content. The total antioxidant capacity had a strong (+) correlation with the condensed tannin content. Linoleic acid peroxidation inhibition had a strong (–) correlation with the total phenolic content. To elucidate the major active phytochemicals in the CWE-EA, HWE-EA, and ME-EA fractions, high performance liquid chromatography-ultraviolet (HPLC-UV) and ultra high performance liquid chromatography coupled with hybrid triple quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) were performed. The HPLC-UV analysis showed the presence of nine compounds in common (arjunic acid and/or euscaphic acid, gallic acid, kaempferol, caffeic acid, ferulic acid, tannic acid, and coumarin, quercetin). The UHPLC-QTOF-MS analysis showed the presence of nine compounds in common (gallic acid; caffeic acid; umbelliferone; arjunic acid, euscaphic acid, and/or tormentic acid; pomolic acid; rosamultic acid; and benzoic acid). When standards of the identified phytochemicals were tested against the same bacterium, quercetin, coumarin, and euscaphic acid showed antibacterial activity against P. acnes.

Acellular bacterial ghosts (BGs) are empty non-living bacterial cell envelopes, commonly generated by controlled expression of the cloned lysis gene E of bacteriophage PhiX174. In this study, Vibrio parahaemolyticus ghosts (VPGs) were generated by chemically-induced lysis and the method is based on minimum inhibitory concentration (MIC) of sodium hydroxide (NaOH), acetic acid, boric acid, citric acid, maleic acid, hydrochloric acid, and sulfuric acid. The MIC values of the respective chemicals were 3.125, 6.25, <50.0, 25.0, 6.25, 1.56, and 0.781 mg/mL. Except for boric acid, the lysis efficiency reached more than 99.99% at 5 min after treatment of all chemicals. Among those chemicals, NaOH-induced VPGs appeared completely DNA-free, which was confirmed by quantitative real-time PCR. Besides, lipopolysaccharides (LPS) extracted from the NaOH-induced VPGs showed no distinctive band on SDS-PAGE gel after silver staining. On the other hand, LPS extracted from wild-type bacterial cells, as well as the organic acids-induced VPGs showed triple major bands and LPS extracted from the inorganic acids-induced VPGs showed double bands. It suggests that some surface structures in LPS of the NaOH-induced VPGs may be lost, weakened, or modified by the MIC of NaOH. Nevertheless, Limulus amoebocyte lysate assay revealed that there is no significant difference in endotoxic activity between the NaOH-induced VPGs and wild-type bacterial cells. Macrophages exposed to the NaOH-induced VPGs at 0.5 × 10⁶ CFU/mL showed cell viability of 97.9%, however, the MIC of NaOH did not reduce the cytotoxic effect of wild-type bacterial cells. Like Escherichia coli LPS, the NaOH-induced VPGs are an excellent activator of pro-inflammatory cytokines (IL-1β and iNOS), anti-inflammatory cytokine (IL-10), and dual activities (IL-6) in the stimulated macrophage cells. On the other hand, the induction of TNF-α mRNA was remarkable in the macrophages exposed with wild-type cells. Scanning electron microscopy showed the formation of trans-membrane lysis tunnel structures in the NaOH-induced VPGs. SDS-PAGE and agarose gel electrophoresis also confirmed that cytoplasmic proteins and genomic DNA released from the VPGs to culture medium through the lysis tunnel structures. Taken together, all these data indicate that the NaOH-induced VPGs show the potency of a safe, economical, and effective inactivated bacterial vaccine candidate.

Salmonella enteritidis and Salmonella typhimurium are important food-borne bacterial pathogens, which are responsible for diarrhea and gastroenteritis in humans and animals. In this study, S. typhimurium bacterial ghost (STG) was generated based on minimum inhibitory concentration (MIC) of sodium hydroxide (NaOH). Experimental studies performed using in vitro and in vivo experimental model systems to characterize effects of STG as a vaccine candidate. When compared with murine macrophages (RAW 264.7) exposed to PBS buffer (98.1%), the macrophages exposed to formalin-killed inactivated cells (FKC), live wild-type bacterial cells and NaOH-induced STG at 1 × 10⁸ CFU/mL showed 85.6%, 66.5% and 84.6% cell viability, respectively. It suggests that STG significantly reduces the cytotoxic effect of wild-type bacterial cells. Furthermore, STG is an excellent inducer for mRNAs of pro-inflammatory cytokine (TNF-α, IL-1β) and factor (iNOS), anti-inflammatory cytokine (IL-10) and dual activities (IL-6) in the stimulated macrophage cells. In vivo, STG vaccine induced humoral and cellular immune responses and protection against homologous and heterologous challenges in rats. Furthermore, the immunogenicity and protective efficacy of STG vaccine were compared with those of FKC and non-vaccinated PBS control groups. The vaccinated rats from STG group exhibited higher levels of serum IgG antibody responses, serum bactericidal antibodies, and CD4⁺ and CD8⁺ T-cell populations than those of the FKC and PBS control groups. Most importantly, after challenge with homologous and heterologous strains, the bacterial loads in the STG group were markedly lower than the FKC and PBS control groups. In conclusion, these findings suggest that the STG vaccine induces protective immunity against homologous and heterologous challenges.